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Journal of Herbs Spices amp Medicinal Plants
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Wild Goldenseal (Hydrastis canadensis) RhizomeRoot Alkaloid Content in Relation to Colony andHarvest Stage
Eric P Burkhart amp Grady H Zuiderveen
To cite this article Eric P Burkhart amp Grady H Zuiderveen (2019) Wild Goldenseal (Hydrastiscanadensis) RhizomeRoot Alkaloid Content in Relation to Colony and Harvest Stage Journal ofHerbs Spices amp Medicinal Plants 252 128-140 DOI 1010801049647520191577322
To link to this article httpsdoiorg1010801049647520191577322
Published online 22 Feb 2019
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Wild Goldenseal (Hydrastis canadensis) RhizomeRootAlkaloid Content in Relation to Colony and Harvest StageEric P Burkhartab and Grady H Zuiderveenb
aShaverrsquos Creek Environmental Center The Pennsylvania State University Petersburg PA USAbDepartment of Ecosystem Science and Management The Pennsylvania State University UniversityPark PA USA
ABSTRACTThis study examined the three major alkaloids (berberine hydras-tine and canadine) in wild goldenseal (Hydrastis canadensis) rootsand rhizomes in relation to plant colony and harvest stageGoldenseal colonies in central Pennsylvania USA were sampledon four dates (July 2 August 7 September 8 and October 12)corresponding with observable phenological stages between fruitmaturity and senescence Variation was observed for all threealkaloids with berberine and hydrastine present in all coloniesand samples while canadine was not detected during some lateseason sample dates Nineteen root samples (53) met the estab-lished United States Pharmacopeia (USP) standards for berberinecontent while only one sample (28) met USP standards forhydrastine All colonies and samples showed an increase in alkaloidlevels at the time of senescence which corroborated the industryguidance that rhizomesroots should be harvested at senescence(typically during the Fall season) Harvesting at senescence alsopermits fruit to mature and thereby facilitates sexual reproductionHowever alkaloid levels averaged the second highest at fruitmaturity (July 2) which suggested that alkaloids may fluctuateduring the growing season in response to or as a function ofkey reproductive events
ARTICLE HISTORYReceived 4 December 2017
KEYWORDSAlkaloids berberinemedicinal plant chemistryphytochemistry secondarymetabolites
Introduction
Goldenseal (Hydrastis canadensis) is an herbaceous perennial indigenous to east-ern North American forestlands The rhizomes and roots of this species are usedmedicinally for antimicrobial and digestive purposes (1) Significant medicinalchemical constituents identified to date in rhizomes and roots include the iso-quinaline alkaloids berberine berberastine tetrahydroberberastine hydrastinehydrastinine canadine and canalidine (23) Of these berberine hydrastine andcanadine have received the most attention in research and have confirmed anti-microbial properties (4) Berberine is often credited with many of the healthfulbenefits associated with goldenseal (1) Although berberine can be obtained fromother plant species such as Berberis spp (3) the additional alkaloids hydrastine
CONTACT Eric P Burkhart epb6psuedu Shaverrsquos Creek Environmental Center The Pennsylvania StateUniversity Petersburg PA 16669 USA
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS2019 VOL 25 NO 2 128ndash140httpsdoiorg1010801049647520191577322
copy 2019 Taylor amp Francis Group LLC
and canadine in goldenseal are also considered to be therapeutically important (4)This combined (ldquoholisticrdquo) chemistry drives continued interest and demand forgoldenseal despite alternative and in many cases cheaper and more readilyavailable (eg Berberis spp Mahonia spp) berberine containing plants
Goldenseal was introduced into cultivation in the United States more thana century ago (5) but the adoption of goldenseal as a specialty crop has beenlimited due to volatile prices and demand and profitability constraints (67)The available trade survey data suggest that most goldenseal in the herbalmarket today originates from wild harvesting in the Appalachian region ofthe eastern United States (89) It is the practice for some commercial rootdiggers and herb collectors in Appalachia to harvest plants at any time theyare requested by buyers do so andor plants are discovered during forestsearches with little if any attention paid to the influence of timing onconstituent levels (10) Harvesting without consideration of phytochemistryraises questions regarding the quality andor potency of wild collectedmedicinal products Studies of other wild-collected Appalachian medicinalforest plants such as bloodroot Sanguinaria canadensis (11) Americanmayapple Podophyllum peltatum (12) and American ginseng Panax quin-quefolius (13) for example have shown that there are often differences inchemistry resulting from when and where plants are harvested and that thesecan have important qualitative consequences for herb buyers and consumers
The present study examined alkaloid content in goldenseal rhizomes androots In particular berberine hydrastine and canadine were studied in wild-harvested rhizomes and roots in three plant colonies across four harvest datesand stages to examine (1) alkaloid variation in wild-harvested rhizomes androots between colonies occurring in a single forested area and (2) the preferredpost-reproductive phenological stage to harvest for purposes of maximizing rootalkaloid content The study sought to test the hypothesis that harvest location andtiming could influence constituent levels in wild goldenseal Given that thespecies is still largely harvested from the wild the results may have implicationsfor the collection and trade of this wild species and contribute to improvedunderstanding of the timing of medicinal root collection in general The results ofthis study can also be useful for the determination of harvest timing in agrofor-estry cultivation systems using a forest farming approach
Materials and Methods
Study Population and Associated Habitat Conditions
Goldenseal exhibits modular growth through asexual reproduction Specifically itis a clonal colonial plant that forms ldquopatchesrdquo over time which may be composedof many physically connected interdependent individuals (ie ramets) (1415)Accordingly it is difficult to distinguish between genets and ramets in populations
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 129
In this study no attempt was made to distinguish between the two instead threespatially separate and distinct clonal ldquopatchesrdquo (hereafter referred to as lsquocoloniesrsquo)were sampled within a forested drainage covering an area of approximately sixhectares Colony-1 was situated at the lowermost topographic position in theforested drainage (elevation 305 m) colony-2 at a middle-upper location andcolony-3 occurred at the upper end of the drainage (elevation 370m) Each colonywas spatially distinct with approximately 450m between colonies 1 and 3 The sizeof each colonywithin the population rootweight and associated soil chemistry areprovided in Table 1
Owing to conservation concerns surrounding goldenseal the exact location ofthis study is withheld from this paper but Global Positioning System (GPS)coordinates are available from the authors and are on-file with the PennsylvaniaDepartment of Conservation and Natural Resources (DCNR) Wild PlantManagement Program (Harrisburg PA) Voucher specimens for the study weredeposited in the following herbaria the Pennsylvania State University in StateCollege PA (PAC) the Carnegie Museum of Natural History in Pittsburgh PA(CM) and the Morris Arboretum of the University of Pennsylvania inPhiladelphia PA (MOAR)
RhizomeRoot Sampling and Processing
Goldenseal produces short stout rhizomes covered with irregularly spacedfine roots along with more slender rhizomes produced for colonialexpansion In this study the entire rhizome and roots were gathered atsampling and were prepared and analyzed collectively Hence the termldquorhizomerootrdquo is used throughout This was done to remain consistentwith post-harvest handling practices in the trade where there is generallylittle effort to differentiate rhizomes from roots amongst collectors orbuyers
Goldenseal rhizomesroots were harvested during the summer of 2012from mature reproductive stems (ie stems bearing 2ndash3 leaves) on fourdates corresponding with the following phenological stages (1) July 2 fruitpresent and fully mature foliage green (2) August 7 post-fruit bearingfoliage green (3) September 8 post-fruit bearing some foliage beginning toyellow but not senescing and (4) October 12 90ndash100 yellow foliage ineach colony around 50 of each colony already senesced For each sampledate three rhizomeroot samples were collected from each of the threecolonies for a total of nine samples per date and 36 samples totalRhizomesroots were washed and dried at 35degC for 24ndash36 h until theywere dry enough to break cleanly Dried rhizomesroots were weighed andshipped overnight from Pennsylvania to North Carolina where they wereprepared for analysis
130 E P BURKHART AND G H ZUIDERVEEN
Table1
Wild
goldenseal
(HydrastiscanadensisL)colony
characteristicsandassociated
soilcond
ition
sforsampleplotsinclud
edin
thisstud
yCo
lony
characteristics
Associated
soilcond
ition
s(n
=5)a
Num
berof
ramets
Dry
root
weigh
tsa
(n=50)
pHPh
osph
atekg
haminus1
Potash
kgha
minus1
Magnesium
kgha
minus1
Calcium
kgha
minus1
Catio
nExchange
Capacity
Acidity
perm
2total
Colony-1
28ndash53
1000ndash1500
18g(07)
63(02)
83(20)
202(47)
536(62)
7364
(1026)
17(18)
38(10)
Colony-2
24ndash45
1500ndash3000
17g(04)
70(03)
155(9)
259(52)
615(164)
9518
(1451)
164
(17)
06(13)
Colony-3
19ndash31
100ndash150
09g(02)
66(01)
89(10)
298(27)
492(91)
6064
(1884)
135
(27)
24(13)
a Stand
arddeviations
areinclud
edin
parentheses
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 131
RhizomeRoot Sample Preparation and Chemical Analysis
Dried whole rhizomeroot samples were ground to approximately 60 mesh witha Foss Cyclotectrade mill prior to extraction For each sample about 250 mg ofground material was weighed and placed into a 50 mL centrifuge tube to which250 mL of extraction solvent (waterndashacetonitrilendashphosphoric acid 70 + 30 + 01vvv) was added The sample was capped and mixed using a Vortexreg mixer for10 sec sonicated in a Bransonic 2510 ultrasonic cleaner for 10 min and thenvortexed again following sonication The solid material was settled by centrifu-ging the sample preparation for 5 min at 4000 rpm in a Sorvallreg Legend tabletopcentrifuge An aliquot of the supernatant was then filtered with a 045 microm PTFEsyringe filter In an amber HPLC vial 200 microL of filtered sample was added to800 microL diluent (90 acetonitrile 10 water) and vortexed prior to analysis
Calibration curves were prepared from five mixed standard solutions contain-ing hydrastine berberine palmatine and canadine Standards were prepared inlow-actinic volumetric flasks and diluted with 90 acetonitrile10 waterConcentrations of individual stock standards were calculated based on thereported purity and corrected for chloride content in the case of berberine andpalmatine The linear ranges of the calibration curves were as follows hydrastine103ndash155 microgmLminus1 palmatine 367ndash551 microgmLminus1 berberine 794ndash119 microgmLminus1 andcanadine 564ndash846 microgmLminus1 Curves were all linear with an r2 value of above 0999
High Pressure Liquid Chromatography (HPLC) analysis was performed usingChromeleonreg chromatography management software on a Dionex ICS-3000system equipped with dual pump automatic sample injector and variable wave-length detector The column temperature was maintained at 40degC The analyticalcolumn was an Agilent Zorbax Eclipse XDB-C18 column (150 times 46 mm 35 microm)The mobile phase consisted of 25 mM ammonium formate pH 38 (C) and 01triethylamine in acetonitrile (D) at a flow rate of 075 mL minminus1 The isocraticeluting mobile phase was 70 C and 30 D and injection volume was 10 microLDetection was at 230 nm and total run time was 13 min per injection
Reagents and standards HPLC-grade acetonitrile formic acid (ge98)o-phosphoric acid (85) triethylamine (99) and ammonium formate (99)were obtained from Fisher Scientific HPLC-grade chemical reference standardsberberine chloride (886) (1R 9S) -(-)-B-hydrastine (995) palmatine chlor-ide (tetramethoxyprotoberberine chloride 792) and DL-canadine (tetrahy-droberberine 972) were purchased from Chromadex (Santa Ana CA)
Statistical Analysis
In addition to basic statistical analyses Pearsonrsquos correlation analysis wasused to examine rhizomeroot phytochemistry in relation to soil chemistryand colony Linear regression was used to examine rhizomeroot phytochem-istry in relation to rhizomeroot weight
132 E P BURKHART AND G H ZUIDERVEEN
Analysis of variance was used to compare mean values of total alkaloidconcentration as well as the three individual alkaloids separately for eachharvest time and location Differences between individual harvest data andsites were analyzed using Bonferroni Pairwise Comparisons
Statistical analyses were conducted using SPSS (version 24) and Minitab173 (Minitab 17 Statistical Software 2010)
Results
Variation was observed both within and between colonies for all three alkaloids(Table 2) Total rhizomeroot alkaloid content varied from a low of 32 inAugust (represented by a single sample from Colony-1) to a high of 48 in Julyand October (samples both collected from Colony-3) Both total and individualalkaloid content was associated with harvest timing with the highest alkaloidsobserved at the senescent stage in October (Fig 1) Berberine and hydrastinewere detected in all samples and colonies Canadine however was not detectedin Colony-3 samples from two dates (Aug 7 and Sept 8) Since each sample wasreplicated (n = 3) on each date and in each colony sampled it is unlikely thatthe absence of canadine in colony 3 on two sample dates was the result ofsample processing or analytical errors
Correlation results were insignificant for all soil chemistry parameters exam-ined except for soil Ca and P which were correlated with alkaloid content on twoharvest dates (Table 3) These correlations were both positive (Ca P and cana-dine) and negative (Ca and hydrastine) however revealed no clear trend Themost significant and perhaps interesting correlations were for total alkaloid con-tent root weight and colony at the final harvest date (October) In the case of rootweight there was a negative correlation between rhizomeroot weight and totalalkaloid content indicating that smaller rhizomesroots contained higher totalalkaloid concentrations (Fig 2) For the effect of colony there was a positivecorrelation between colony sampled and total alkaloid content Average rootalkaloid content was the highest in colony-3 on all sample dates
Discussion
The Influence of Harvest Stage and Timing on Quality
The World Health Organization (WHO) Guidelines on Good Agriculturaland Collection Practices (GACP) for Medicinal Plants provide the followingguidance with respect to harvest timing of wild-collected medicinalplants (16)
ldquoMedicinal plant materials should be collected during the appropriateseason or time period to ensure the best possible quality of both sourcematerials and finished products It is well known that the quantitative
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 133
Table2
Alkaloid
levelsin
wild-harvested
goldensealrootsrhizom
esin
relatio
nto
colony
andharveststage(n
=3foreach
plot
oneach
harvestdaten
=36
totalsam
ples)Observatio
nsregardingph
enolog
ical
stageat
thetim
eof
harvestareinclud
edbelow
theharvestdate
July2
2012
Augu
st7
2012
Septem
ber8
2012
Octob
er122
012
Fruitpresentandfully
maturefoliage
green
Post
fruitbearing
foliage
green
Post
fruitbearing
foliage
beginn
ingto
yellow
Foliage
yellowp
lants
senescing
Colony
1Berberineww
25σ02(23ndash27)
21σ02(19ndash23)
23σ02(21ndash26)
25σ05(22ndash31)
Hydrastineww
14σ02(13ndash16)
16σ03(13ndash19)
15σ00(15ndash15)
15σ01(14ndash16)
Canadine
ww
008
σ001
(07-09)
003
σ001
(03-04)
011
σ001
(11-12)
018
σ003
(14-20)
Totalalkaloids
ww
39σ03(36ndash41)
37σ05(32ndash42)
38σ03(35ndash41)
40σ06(37ndash47)
Colony
2Berberineww
23σ01(22ndash25)
22σ01(22ndash23)
25σ01(23ndash26)
29σ01(28ndash30)
Hydrastineww
13σ01(11ndash14)
14σ01(13ndash15)
13σ01(12ndash14)
15σ00(15ndash16)
Canadine
ww
009
σ02(07-09)
008
σ004
(03-11)
009
σ001
(ND-09)
018
σ002
(17-20)
Totalalkaloids
ww
36σ03(34ndash39)
37σ01(35ndash37)
38σ03(35ndash39)
44σ01(44ndash45)
Colony
3Berberineww
25σ02(23ndash27)
22σ03(20ndash25)
26σ01(25ndash27)
28σ01(28ndash29)
Hydrastineww
18σ03(15ndash21)
16σ01(15ndash16)
17σ01(17ndash18)
19σ01(18ndash20)
Canadine
ww
007
σ001
(07-08)
Not
detected
(ND)
Not
detected
(ND)
023
σ004
(20-27)
Totalalkaloids
ww
43σ05(38ndash48)
38σ03(35ndash41)
43σ01(42ndash45)
47σ01(46ndash48)
134 E P BURKHART AND G H ZUIDERVEEN
concentration of biologically active constituents varies with the stage ofplant growth and development [hellip] The best time for collection (qualitypeak season or time of day) should be determined according to thequality and quantity of biologically active constituents rather than thetotal vegetative yield of the targeted medicinal plant partsrdquo [bold empha-sis authorsrsquo]
Goldenseal is included in the United States Pharmacopeia (USP) and USPstandards require a minimum alkaloid content of 25 berberine and 20hydrastine for dried goldenseal rhizomes and roots (17) Out of the 36 wild-harvested goldenseal rhizomeroot samples analyzed in this study only abouthalf (53 n = 19) met this threshold for berberine content while only onesample (28) met the hydrastine threshold (Fig 3) Nearly half of wildrhizomeroots samples harvested before senescence and nearly all harvestedin July and August failed to meet USP minimums for alkaloid content
Figure 1 Alkaloid levels in wild-harvested goldenseal rhizomesroots in relation to colony andharvest date Clockwise from top-left total alkaloid content (berberine hydrastine and canadine)berberine canadine and hydrastine Note the different scales on the vertical axes Harvest datescorresponded with the following phenological stages 0702 = fruit present and fully ripe foliagegreen 0807 = fruit gone foliage green 0908 = foliage beginning to yellow 1012 = foliageyellow plants senescing
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 135
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
Wild Goldenseal (Hydrastis canadensis) RhizomeRootAlkaloid Content in Relation to Colony and Harvest StageEric P Burkhartab and Grady H Zuiderveenb
aShaverrsquos Creek Environmental Center The Pennsylvania State University Petersburg PA USAbDepartment of Ecosystem Science and Management The Pennsylvania State University UniversityPark PA USA
ABSTRACTThis study examined the three major alkaloids (berberine hydras-tine and canadine) in wild goldenseal (Hydrastis canadensis) rootsand rhizomes in relation to plant colony and harvest stageGoldenseal colonies in central Pennsylvania USA were sampledon four dates (July 2 August 7 September 8 and October 12)corresponding with observable phenological stages between fruitmaturity and senescence Variation was observed for all threealkaloids with berberine and hydrastine present in all coloniesand samples while canadine was not detected during some lateseason sample dates Nineteen root samples (53) met the estab-lished United States Pharmacopeia (USP) standards for berberinecontent while only one sample (28) met USP standards forhydrastine All colonies and samples showed an increase in alkaloidlevels at the time of senescence which corroborated the industryguidance that rhizomesroots should be harvested at senescence(typically during the Fall season) Harvesting at senescence alsopermits fruit to mature and thereby facilitates sexual reproductionHowever alkaloid levels averaged the second highest at fruitmaturity (July 2) which suggested that alkaloids may fluctuateduring the growing season in response to or as a function ofkey reproductive events
ARTICLE HISTORYReceived 4 December 2017
KEYWORDSAlkaloids berberinemedicinal plant chemistryphytochemistry secondarymetabolites
Introduction
Goldenseal (Hydrastis canadensis) is an herbaceous perennial indigenous to east-ern North American forestlands The rhizomes and roots of this species are usedmedicinally for antimicrobial and digestive purposes (1) Significant medicinalchemical constituents identified to date in rhizomes and roots include the iso-quinaline alkaloids berberine berberastine tetrahydroberberastine hydrastinehydrastinine canadine and canalidine (23) Of these berberine hydrastine andcanadine have received the most attention in research and have confirmed anti-microbial properties (4) Berberine is often credited with many of the healthfulbenefits associated with goldenseal (1) Although berberine can be obtained fromother plant species such as Berberis spp (3) the additional alkaloids hydrastine
CONTACT Eric P Burkhart epb6psuedu Shaverrsquos Creek Environmental Center The Pennsylvania StateUniversity Petersburg PA 16669 USA
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS2019 VOL 25 NO 2 128ndash140httpsdoiorg1010801049647520191577322
copy 2019 Taylor amp Francis Group LLC
and canadine in goldenseal are also considered to be therapeutically important (4)This combined (ldquoholisticrdquo) chemistry drives continued interest and demand forgoldenseal despite alternative and in many cases cheaper and more readilyavailable (eg Berberis spp Mahonia spp) berberine containing plants
Goldenseal was introduced into cultivation in the United States more thana century ago (5) but the adoption of goldenseal as a specialty crop has beenlimited due to volatile prices and demand and profitability constraints (67)The available trade survey data suggest that most goldenseal in the herbalmarket today originates from wild harvesting in the Appalachian region ofthe eastern United States (89) It is the practice for some commercial rootdiggers and herb collectors in Appalachia to harvest plants at any time theyare requested by buyers do so andor plants are discovered during forestsearches with little if any attention paid to the influence of timing onconstituent levels (10) Harvesting without consideration of phytochemistryraises questions regarding the quality andor potency of wild collectedmedicinal products Studies of other wild-collected Appalachian medicinalforest plants such as bloodroot Sanguinaria canadensis (11) Americanmayapple Podophyllum peltatum (12) and American ginseng Panax quin-quefolius (13) for example have shown that there are often differences inchemistry resulting from when and where plants are harvested and that thesecan have important qualitative consequences for herb buyers and consumers
The present study examined alkaloid content in goldenseal rhizomes androots In particular berberine hydrastine and canadine were studied in wild-harvested rhizomes and roots in three plant colonies across four harvest datesand stages to examine (1) alkaloid variation in wild-harvested rhizomes androots between colonies occurring in a single forested area and (2) the preferredpost-reproductive phenological stage to harvest for purposes of maximizing rootalkaloid content The study sought to test the hypothesis that harvest location andtiming could influence constituent levels in wild goldenseal Given that thespecies is still largely harvested from the wild the results may have implicationsfor the collection and trade of this wild species and contribute to improvedunderstanding of the timing of medicinal root collection in general The results ofthis study can also be useful for the determination of harvest timing in agrofor-estry cultivation systems using a forest farming approach
Materials and Methods
Study Population and Associated Habitat Conditions
Goldenseal exhibits modular growth through asexual reproduction Specifically itis a clonal colonial plant that forms ldquopatchesrdquo over time which may be composedof many physically connected interdependent individuals (ie ramets) (1415)Accordingly it is difficult to distinguish between genets and ramets in populations
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 129
In this study no attempt was made to distinguish between the two instead threespatially separate and distinct clonal ldquopatchesrdquo (hereafter referred to as lsquocoloniesrsquo)were sampled within a forested drainage covering an area of approximately sixhectares Colony-1 was situated at the lowermost topographic position in theforested drainage (elevation 305 m) colony-2 at a middle-upper location andcolony-3 occurred at the upper end of the drainage (elevation 370m) Each colonywas spatially distinct with approximately 450m between colonies 1 and 3 The sizeof each colonywithin the population rootweight and associated soil chemistry areprovided in Table 1
Owing to conservation concerns surrounding goldenseal the exact location ofthis study is withheld from this paper but Global Positioning System (GPS)coordinates are available from the authors and are on-file with the PennsylvaniaDepartment of Conservation and Natural Resources (DCNR) Wild PlantManagement Program (Harrisburg PA) Voucher specimens for the study weredeposited in the following herbaria the Pennsylvania State University in StateCollege PA (PAC) the Carnegie Museum of Natural History in Pittsburgh PA(CM) and the Morris Arboretum of the University of Pennsylvania inPhiladelphia PA (MOAR)
RhizomeRoot Sampling and Processing
Goldenseal produces short stout rhizomes covered with irregularly spacedfine roots along with more slender rhizomes produced for colonialexpansion In this study the entire rhizome and roots were gathered atsampling and were prepared and analyzed collectively Hence the termldquorhizomerootrdquo is used throughout This was done to remain consistentwith post-harvest handling practices in the trade where there is generallylittle effort to differentiate rhizomes from roots amongst collectors orbuyers
Goldenseal rhizomesroots were harvested during the summer of 2012from mature reproductive stems (ie stems bearing 2ndash3 leaves) on fourdates corresponding with the following phenological stages (1) July 2 fruitpresent and fully mature foliage green (2) August 7 post-fruit bearingfoliage green (3) September 8 post-fruit bearing some foliage beginning toyellow but not senescing and (4) October 12 90ndash100 yellow foliage ineach colony around 50 of each colony already senesced For each sampledate three rhizomeroot samples were collected from each of the threecolonies for a total of nine samples per date and 36 samples totalRhizomesroots were washed and dried at 35degC for 24ndash36 h until theywere dry enough to break cleanly Dried rhizomesroots were weighed andshipped overnight from Pennsylvania to North Carolina where they wereprepared for analysis
130 E P BURKHART AND G H ZUIDERVEEN
Table1
Wild
goldenseal
(HydrastiscanadensisL)colony
characteristicsandassociated
soilcond
ition
sforsampleplotsinclud
edin
thisstud
yCo
lony
characteristics
Associated
soilcond
ition
s(n
=5)a
Num
berof
ramets
Dry
root
weigh
tsa
(n=50)
pHPh
osph
atekg
haminus1
Potash
kgha
minus1
Magnesium
kgha
minus1
Calcium
kgha
minus1
Catio
nExchange
Capacity
Acidity
perm
2total
Colony-1
28ndash53
1000ndash1500
18g(07)
63(02)
83(20)
202(47)
536(62)
7364
(1026)
17(18)
38(10)
Colony-2
24ndash45
1500ndash3000
17g(04)
70(03)
155(9)
259(52)
615(164)
9518
(1451)
164
(17)
06(13)
Colony-3
19ndash31
100ndash150
09g(02)
66(01)
89(10)
298(27)
492(91)
6064
(1884)
135
(27)
24(13)
a Stand
arddeviations
areinclud
edin
parentheses
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 131
RhizomeRoot Sample Preparation and Chemical Analysis
Dried whole rhizomeroot samples were ground to approximately 60 mesh witha Foss Cyclotectrade mill prior to extraction For each sample about 250 mg ofground material was weighed and placed into a 50 mL centrifuge tube to which250 mL of extraction solvent (waterndashacetonitrilendashphosphoric acid 70 + 30 + 01vvv) was added The sample was capped and mixed using a Vortexreg mixer for10 sec sonicated in a Bransonic 2510 ultrasonic cleaner for 10 min and thenvortexed again following sonication The solid material was settled by centrifu-ging the sample preparation for 5 min at 4000 rpm in a Sorvallreg Legend tabletopcentrifuge An aliquot of the supernatant was then filtered with a 045 microm PTFEsyringe filter In an amber HPLC vial 200 microL of filtered sample was added to800 microL diluent (90 acetonitrile 10 water) and vortexed prior to analysis
Calibration curves were prepared from five mixed standard solutions contain-ing hydrastine berberine palmatine and canadine Standards were prepared inlow-actinic volumetric flasks and diluted with 90 acetonitrile10 waterConcentrations of individual stock standards were calculated based on thereported purity and corrected for chloride content in the case of berberine andpalmatine The linear ranges of the calibration curves were as follows hydrastine103ndash155 microgmLminus1 palmatine 367ndash551 microgmLminus1 berberine 794ndash119 microgmLminus1 andcanadine 564ndash846 microgmLminus1 Curves were all linear with an r2 value of above 0999
High Pressure Liquid Chromatography (HPLC) analysis was performed usingChromeleonreg chromatography management software on a Dionex ICS-3000system equipped with dual pump automatic sample injector and variable wave-length detector The column temperature was maintained at 40degC The analyticalcolumn was an Agilent Zorbax Eclipse XDB-C18 column (150 times 46 mm 35 microm)The mobile phase consisted of 25 mM ammonium formate pH 38 (C) and 01triethylamine in acetonitrile (D) at a flow rate of 075 mL minminus1 The isocraticeluting mobile phase was 70 C and 30 D and injection volume was 10 microLDetection was at 230 nm and total run time was 13 min per injection
Reagents and standards HPLC-grade acetonitrile formic acid (ge98)o-phosphoric acid (85) triethylamine (99) and ammonium formate (99)were obtained from Fisher Scientific HPLC-grade chemical reference standardsberberine chloride (886) (1R 9S) -(-)-B-hydrastine (995) palmatine chlor-ide (tetramethoxyprotoberberine chloride 792) and DL-canadine (tetrahy-droberberine 972) were purchased from Chromadex (Santa Ana CA)
Statistical Analysis
In addition to basic statistical analyses Pearsonrsquos correlation analysis wasused to examine rhizomeroot phytochemistry in relation to soil chemistryand colony Linear regression was used to examine rhizomeroot phytochem-istry in relation to rhizomeroot weight
132 E P BURKHART AND G H ZUIDERVEEN
Analysis of variance was used to compare mean values of total alkaloidconcentration as well as the three individual alkaloids separately for eachharvest time and location Differences between individual harvest data andsites were analyzed using Bonferroni Pairwise Comparisons
Statistical analyses were conducted using SPSS (version 24) and Minitab173 (Minitab 17 Statistical Software 2010)
Results
Variation was observed both within and between colonies for all three alkaloids(Table 2) Total rhizomeroot alkaloid content varied from a low of 32 inAugust (represented by a single sample from Colony-1) to a high of 48 in Julyand October (samples both collected from Colony-3) Both total and individualalkaloid content was associated with harvest timing with the highest alkaloidsobserved at the senescent stage in October (Fig 1) Berberine and hydrastinewere detected in all samples and colonies Canadine however was not detectedin Colony-3 samples from two dates (Aug 7 and Sept 8) Since each sample wasreplicated (n = 3) on each date and in each colony sampled it is unlikely thatthe absence of canadine in colony 3 on two sample dates was the result ofsample processing or analytical errors
Correlation results were insignificant for all soil chemistry parameters exam-ined except for soil Ca and P which were correlated with alkaloid content on twoharvest dates (Table 3) These correlations were both positive (Ca P and cana-dine) and negative (Ca and hydrastine) however revealed no clear trend Themost significant and perhaps interesting correlations were for total alkaloid con-tent root weight and colony at the final harvest date (October) In the case of rootweight there was a negative correlation between rhizomeroot weight and totalalkaloid content indicating that smaller rhizomesroots contained higher totalalkaloid concentrations (Fig 2) For the effect of colony there was a positivecorrelation between colony sampled and total alkaloid content Average rootalkaloid content was the highest in colony-3 on all sample dates
Discussion
The Influence of Harvest Stage and Timing on Quality
The World Health Organization (WHO) Guidelines on Good Agriculturaland Collection Practices (GACP) for Medicinal Plants provide the followingguidance with respect to harvest timing of wild-collected medicinalplants (16)
ldquoMedicinal plant materials should be collected during the appropriateseason or time period to ensure the best possible quality of both sourcematerials and finished products It is well known that the quantitative
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 133
Table2
Alkaloid
levelsin
wild-harvested
goldensealrootsrhizom
esin
relatio
nto
colony
andharveststage(n
=3foreach
plot
oneach
harvestdaten
=36
totalsam
ples)Observatio
nsregardingph
enolog
ical
stageat
thetim
eof
harvestareinclud
edbelow
theharvestdate
July2
2012
Augu
st7
2012
Septem
ber8
2012
Octob
er122
012
Fruitpresentandfully
maturefoliage
green
Post
fruitbearing
foliage
green
Post
fruitbearing
foliage
beginn
ingto
yellow
Foliage
yellowp
lants
senescing
Colony
1Berberineww
25σ02(23ndash27)
21σ02(19ndash23)
23σ02(21ndash26)
25σ05(22ndash31)
Hydrastineww
14σ02(13ndash16)
16σ03(13ndash19)
15σ00(15ndash15)
15σ01(14ndash16)
Canadine
ww
008
σ001
(07-09)
003
σ001
(03-04)
011
σ001
(11-12)
018
σ003
(14-20)
Totalalkaloids
ww
39σ03(36ndash41)
37σ05(32ndash42)
38σ03(35ndash41)
40σ06(37ndash47)
Colony
2Berberineww
23σ01(22ndash25)
22σ01(22ndash23)
25σ01(23ndash26)
29σ01(28ndash30)
Hydrastineww
13σ01(11ndash14)
14σ01(13ndash15)
13σ01(12ndash14)
15σ00(15ndash16)
Canadine
ww
009
σ02(07-09)
008
σ004
(03-11)
009
σ001
(ND-09)
018
σ002
(17-20)
Totalalkaloids
ww
36σ03(34ndash39)
37σ01(35ndash37)
38σ03(35ndash39)
44σ01(44ndash45)
Colony
3Berberineww
25σ02(23ndash27)
22σ03(20ndash25)
26σ01(25ndash27)
28σ01(28ndash29)
Hydrastineww
18σ03(15ndash21)
16σ01(15ndash16)
17σ01(17ndash18)
19σ01(18ndash20)
Canadine
ww
007
σ001
(07-08)
Not
detected
(ND)
Not
detected
(ND)
023
σ004
(20-27)
Totalalkaloids
ww
43σ05(38ndash48)
38σ03(35ndash41)
43σ01(42ndash45)
47σ01(46ndash48)
134 E P BURKHART AND G H ZUIDERVEEN
concentration of biologically active constituents varies with the stage ofplant growth and development [hellip] The best time for collection (qualitypeak season or time of day) should be determined according to thequality and quantity of biologically active constituents rather than thetotal vegetative yield of the targeted medicinal plant partsrdquo [bold empha-sis authorsrsquo]
Goldenseal is included in the United States Pharmacopeia (USP) and USPstandards require a minimum alkaloid content of 25 berberine and 20hydrastine for dried goldenseal rhizomes and roots (17) Out of the 36 wild-harvested goldenseal rhizomeroot samples analyzed in this study only abouthalf (53 n = 19) met this threshold for berberine content while only onesample (28) met the hydrastine threshold (Fig 3) Nearly half of wildrhizomeroots samples harvested before senescence and nearly all harvestedin July and August failed to meet USP minimums for alkaloid content
Figure 1 Alkaloid levels in wild-harvested goldenseal rhizomesroots in relation to colony andharvest date Clockwise from top-left total alkaloid content (berberine hydrastine and canadine)berberine canadine and hydrastine Note the different scales on the vertical axes Harvest datescorresponded with the following phenological stages 0702 = fruit present and fully ripe foliagegreen 0807 = fruit gone foliage green 0908 = foliage beginning to yellow 1012 = foliageyellow plants senescing
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 135
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
and canadine in goldenseal are also considered to be therapeutically important (4)This combined (ldquoholisticrdquo) chemistry drives continued interest and demand forgoldenseal despite alternative and in many cases cheaper and more readilyavailable (eg Berberis spp Mahonia spp) berberine containing plants
Goldenseal was introduced into cultivation in the United States more thana century ago (5) but the adoption of goldenseal as a specialty crop has beenlimited due to volatile prices and demand and profitability constraints (67)The available trade survey data suggest that most goldenseal in the herbalmarket today originates from wild harvesting in the Appalachian region ofthe eastern United States (89) It is the practice for some commercial rootdiggers and herb collectors in Appalachia to harvest plants at any time theyare requested by buyers do so andor plants are discovered during forestsearches with little if any attention paid to the influence of timing onconstituent levels (10) Harvesting without consideration of phytochemistryraises questions regarding the quality andor potency of wild collectedmedicinal products Studies of other wild-collected Appalachian medicinalforest plants such as bloodroot Sanguinaria canadensis (11) Americanmayapple Podophyllum peltatum (12) and American ginseng Panax quin-quefolius (13) for example have shown that there are often differences inchemistry resulting from when and where plants are harvested and that thesecan have important qualitative consequences for herb buyers and consumers
The present study examined alkaloid content in goldenseal rhizomes androots In particular berberine hydrastine and canadine were studied in wild-harvested rhizomes and roots in three plant colonies across four harvest datesand stages to examine (1) alkaloid variation in wild-harvested rhizomes androots between colonies occurring in a single forested area and (2) the preferredpost-reproductive phenological stage to harvest for purposes of maximizing rootalkaloid content The study sought to test the hypothesis that harvest location andtiming could influence constituent levels in wild goldenseal Given that thespecies is still largely harvested from the wild the results may have implicationsfor the collection and trade of this wild species and contribute to improvedunderstanding of the timing of medicinal root collection in general The results ofthis study can also be useful for the determination of harvest timing in agrofor-estry cultivation systems using a forest farming approach
Materials and Methods
Study Population and Associated Habitat Conditions
Goldenseal exhibits modular growth through asexual reproduction Specifically itis a clonal colonial plant that forms ldquopatchesrdquo over time which may be composedof many physically connected interdependent individuals (ie ramets) (1415)Accordingly it is difficult to distinguish between genets and ramets in populations
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 129
In this study no attempt was made to distinguish between the two instead threespatially separate and distinct clonal ldquopatchesrdquo (hereafter referred to as lsquocoloniesrsquo)were sampled within a forested drainage covering an area of approximately sixhectares Colony-1 was situated at the lowermost topographic position in theforested drainage (elevation 305 m) colony-2 at a middle-upper location andcolony-3 occurred at the upper end of the drainage (elevation 370m) Each colonywas spatially distinct with approximately 450m between colonies 1 and 3 The sizeof each colonywithin the population rootweight and associated soil chemistry areprovided in Table 1
Owing to conservation concerns surrounding goldenseal the exact location ofthis study is withheld from this paper but Global Positioning System (GPS)coordinates are available from the authors and are on-file with the PennsylvaniaDepartment of Conservation and Natural Resources (DCNR) Wild PlantManagement Program (Harrisburg PA) Voucher specimens for the study weredeposited in the following herbaria the Pennsylvania State University in StateCollege PA (PAC) the Carnegie Museum of Natural History in Pittsburgh PA(CM) and the Morris Arboretum of the University of Pennsylvania inPhiladelphia PA (MOAR)
RhizomeRoot Sampling and Processing
Goldenseal produces short stout rhizomes covered with irregularly spacedfine roots along with more slender rhizomes produced for colonialexpansion In this study the entire rhizome and roots were gathered atsampling and were prepared and analyzed collectively Hence the termldquorhizomerootrdquo is used throughout This was done to remain consistentwith post-harvest handling practices in the trade where there is generallylittle effort to differentiate rhizomes from roots amongst collectors orbuyers
Goldenseal rhizomesroots were harvested during the summer of 2012from mature reproductive stems (ie stems bearing 2ndash3 leaves) on fourdates corresponding with the following phenological stages (1) July 2 fruitpresent and fully mature foliage green (2) August 7 post-fruit bearingfoliage green (3) September 8 post-fruit bearing some foliage beginning toyellow but not senescing and (4) October 12 90ndash100 yellow foliage ineach colony around 50 of each colony already senesced For each sampledate three rhizomeroot samples were collected from each of the threecolonies for a total of nine samples per date and 36 samples totalRhizomesroots were washed and dried at 35degC for 24ndash36 h until theywere dry enough to break cleanly Dried rhizomesroots were weighed andshipped overnight from Pennsylvania to North Carolina where they wereprepared for analysis
130 E P BURKHART AND G H ZUIDERVEEN
Table1
Wild
goldenseal
(HydrastiscanadensisL)colony
characteristicsandassociated
soilcond
ition
sforsampleplotsinclud
edin
thisstud
yCo
lony
characteristics
Associated
soilcond
ition
s(n
=5)a
Num
berof
ramets
Dry
root
weigh
tsa
(n=50)
pHPh
osph
atekg
haminus1
Potash
kgha
minus1
Magnesium
kgha
minus1
Calcium
kgha
minus1
Catio
nExchange
Capacity
Acidity
perm
2total
Colony-1
28ndash53
1000ndash1500
18g(07)
63(02)
83(20)
202(47)
536(62)
7364
(1026)
17(18)
38(10)
Colony-2
24ndash45
1500ndash3000
17g(04)
70(03)
155(9)
259(52)
615(164)
9518
(1451)
164
(17)
06(13)
Colony-3
19ndash31
100ndash150
09g(02)
66(01)
89(10)
298(27)
492(91)
6064
(1884)
135
(27)
24(13)
a Stand
arddeviations
areinclud
edin
parentheses
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 131
RhizomeRoot Sample Preparation and Chemical Analysis
Dried whole rhizomeroot samples were ground to approximately 60 mesh witha Foss Cyclotectrade mill prior to extraction For each sample about 250 mg ofground material was weighed and placed into a 50 mL centrifuge tube to which250 mL of extraction solvent (waterndashacetonitrilendashphosphoric acid 70 + 30 + 01vvv) was added The sample was capped and mixed using a Vortexreg mixer for10 sec sonicated in a Bransonic 2510 ultrasonic cleaner for 10 min and thenvortexed again following sonication The solid material was settled by centrifu-ging the sample preparation for 5 min at 4000 rpm in a Sorvallreg Legend tabletopcentrifuge An aliquot of the supernatant was then filtered with a 045 microm PTFEsyringe filter In an amber HPLC vial 200 microL of filtered sample was added to800 microL diluent (90 acetonitrile 10 water) and vortexed prior to analysis
Calibration curves were prepared from five mixed standard solutions contain-ing hydrastine berberine palmatine and canadine Standards were prepared inlow-actinic volumetric flasks and diluted with 90 acetonitrile10 waterConcentrations of individual stock standards were calculated based on thereported purity and corrected for chloride content in the case of berberine andpalmatine The linear ranges of the calibration curves were as follows hydrastine103ndash155 microgmLminus1 palmatine 367ndash551 microgmLminus1 berberine 794ndash119 microgmLminus1 andcanadine 564ndash846 microgmLminus1 Curves were all linear with an r2 value of above 0999
High Pressure Liquid Chromatography (HPLC) analysis was performed usingChromeleonreg chromatography management software on a Dionex ICS-3000system equipped with dual pump automatic sample injector and variable wave-length detector The column temperature was maintained at 40degC The analyticalcolumn was an Agilent Zorbax Eclipse XDB-C18 column (150 times 46 mm 35 microm)The mobile phase consisted of 25 mM ammonium formate pH 38 (C) and 01triethylamine in acetonitrile (D) at a flow rate of 075 mL minminus1 The isocraticeluting mobile phase was 70 C and 30 D and injection volume was 10 microLDetection was at 230 nm and total run time was 13 min per injection
Reagents and standards HPLC-grade acetonitrile formic acid (ge98)o-phosphoric acid (85) triethylamine (99) and ammonium formate (99)were obtained from Fisher Scientific HPLC-grade chemical reference standardsberberine chloride (886) (1R 9S) -(-)-B-hydrastine (995) palmatine chlor-ide (tetramethoxyprotoberberine chloride 792) and DL-canadine (tetrahy-droberberine 972) were purchased from Chromadex (Santa Ana CA)
Statistical Analysis
In addition to basic statistical analyses Pearsonrsquos correlation analysis wasused to examine rhizomeroot phytochemistry in relation to soil chemistryand colony Linear regression was used to examine rhizomeroot phytochem-istry in relation to rhizomeroot weight
132 E P BURKHART AND G H ZUIDERVEEN
Analysis of variance was used to compare mean values of total alkaloidconcentration as well as the three individual alkaloids separately for eachharvest time and location Differences between individual harvest data andsites were analyzed using Bonferroni Pairwise Comparisons
Statistical analyses were conducted using SPSS (version 24) and Minitab173 (Minitab 17 Statistical Software 2010)
Results
Variation was observed both within and between colonies for all three alkaloids(Table 2) Total rhizomeroot alkaloid content varied from a low of 32 inAugust (represented by a single sample from Colony-1) to a high of 48 in Julyand October (samples both collected from Colony-3) Both total and individualalkaloid content was associated with harvest timing with the highest alkaloidsobserved at the senescent stage in October (Fig 1) Berberine and hydrastinewere detected in all samples and colonies Canadine however was not detectedin Colony-3 samples from two dates (Aug 7 and Sept 8) Since each sample wasreplicated (n = 3) on each date and in each colony sampled it is unlikely thatthe absence of canadine in colony 3 on two sample dates was the result ofsample processing or analytical errors
Correlation results were insignificant for all soil chemistry parameters exam-ined except for soil Ca and P which were correlated with alkaloid content on twoharvest dates (Table 3) These correlations were both positive (Ca P and cana-dine) and negative (Ca and hydrastine) however revealed no clear trend Themost significant and perhaps interesting correlations were for total alkaloid con-tent root weight and colony at the final harvest date (October) In the case of rootweight there was a negative correlation between rhizomeroot weight and totalalkaloid content indicating that smaller rhizomesroots contained higher totalalkaloid concentrations (Fig 2) For the effect of colony there was a positivecorrelation between colony sampled and total alkaloid content Average rootalkaloid content was the highest in colony-3 on all sample dates
Discussion
The Influence of Harvest Stage and Timing on Quality
The World Health Organization (WHO) Guidelines on Good Agriculturaland Collection Practices (GACP) for Medicinal Plants provide the followingguidance with respect to harvest timing of wild-collected medicinalplants (16)
ldquoMedicinal plant materials should be collected during the appropriateseason or time period to ensure the best possible quality of both sourcematerials and finished products It is well known that the quantitative
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 133
Table2
Alkaloid
levelsin
wild-harvested
goldensealrootsrhizom
esin
relatio
nto
colony
andharveststage(n
=3foreach
plot
oneach
harvestdaten
=36
totalsam
ples)Observatio
nsregardingph
enolog
ical
stageat
thetim
eof
harvestareinclud
edbelow
theharvestdate
July2
2012
Augu
st7
2012
Septem
ber8
2012
Octob
er122
012
Fruitpresentandfully
maturefoliage
green
Post
fruitbearing
foliage
green
Post
fruitbearing
foliage
beginn
ingto
yellow
Foliage
yellowp
lants
senescing
Colony
1Berberineww
25σ02(23ndash27)
21σ02(19ndash23)
23σ02(21ndash26)
25σ05(22ndash31)
Hydrastineww
14σ02(13ndash16)
16σ03(13ndash19)
15σ00(15ndash15)
15σ01(14ndash16)
Canadine
ww
008
σ001
(07-09)
003
σ001
(03-04)
011
σ001
(11-12)
018
σ003
(14-20)
Totalalkaloids
ww
39σ03(36ndash41)
37σ05(32ndash42)
38σ03(35ndash41)
40σ06(37ndash47)
Colony
2Berberineww
23σ01(22ndash25)
22σ01(22ndash23)
25σ01(23ndash26)
29σ01(28ndash30)
Hydrastineww
13σ01(11ndash14)
14σ01(13ndash15)
13σ01(12ndash14)
15σ00(15ndash16)
Canadine
ww
009
σ02(07-09)
008
σ004
(03-11)
009
σ001
(ND-09)
018
σ002
(17-20)
Totalalkaloids
ww
36σ03(34ndash39)
37σ01(35ndash37)
38σ03(35ndash39)
44σ01(44ndash45)
Colony
3Berberineww
25σ02(23ndash27)
22σ03(20ndash25)
26σ01(25ndash27)
28σ01(28ndash29)
Hydrastineww
18σ03(15ndash21)
16σ01(15ndash16)
17σ01(17ndash18)
19σ01(18ndash20)
Canadine
ww
007
σ001
(07-08)
Not
detected
(ND)
Not
detected
(ND)
023
σ004
(20-27)
Totalalkaloids
ww
43σ05(38ndash48)
38σ03(35ndash41)
43σ01(42ndash45)
47σ01(46ndash48)
134 E P BURKHART AND G H ZUIDERVEEN
concentration of biologically active constituents varies with the stage ofplant growth and development [hellip] The best time for collection (qualitypeak season or time of day) should be determined according to thequality and quantity of biologically active constituents rather than thetotal vegetative yield of the targeted medicinal plant partsrdquo [bold empha-sis authorsrsquo]
Goldenseal is included in the United States Pharmacopeia (USP) and USPstandards require a minimum alkaloid content of 25 berberine and 20hydrastine for dried goldenseal rhizomes and roots (17) Out of the 36 wild-harvested goldenseal rhizomeroot samples analyzed in this study only abouthalf (53 n = 19) met this threshold for berberine content while only onesample (28) met the hydrastine threshold (Fig 3) Nearly half of wildrhizomeroots samples harvested before senescence and nearly all harvestedin July and August failed to meet USP minimums for alkaloid content
Figure 1 Alkaloid levels in wild-harvested goldenseal rhizomesroots in relation to colony andharvest date Clockwise from top-left total alkaloid content (berberine hydrastine and canadine)berberine canadine and hydrastine Note the different scales on the vertical axes Harvest datescorresponded with the following phenological stages 0702 = fruit present and fully ripe foliagegreen 0807 = fruit gone foliage green 0908 = foliage beginning to yellow 1012 = foliageyellow plants senescing
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 135
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
In this study no attempt was made to distinguish between the two instead threespatially separate and distinct clonal ldquopatchesrdquo (hereafter referred to as lsquocoloniesrsquo)were sampled within a forested drainage covering an area of approximately sixhectares Colony-1 was situated at the lowermost topographic position in theforested drainage (elevation 305 m) colony-2 at a middle-upper location andcolony-3 occurred at the upper end of the drainage (elevation 370m) Each colonywas spatially distinct with approximately 450m between colonies 1 and 3 The sizeof each colonywithin the population rootweight and associated soil chemistry areprovided in Table 1
Owing to conservation concerns surrounding goldenseal the exact location ofthis study is withheld from this paper but Global Positioning System (GPS)coordinates are available from the authors and are on-file with the PennsylvaniaDepartment of Conservation and Natural Resources (DCNR) Wild PlantManagement Program (Harrisburg PA) Voucher specimens for the study weredeposited in the following herbaria the Pennsylvania State University in StateCollege PA (PAC) the Carnegie Museum of Natural History in Pittsburgh PA(CM) and the Morris Arboretum of the University of Pennsylvania inPhiladelphia PA (MOAR)
RhizomeRoot Sampling and Processing
Goldenseal produces short stout rhizomes covered with irregularly spacedfine roots along with more slender rhizomes produced for colonialexpansion In this study the entire rhizome and roots were gathered atsampling and were prepared and analyzed collectively Hence the termldquorhizomerootrdquo is used throughout This was done to remain consistentwith post-harvest handling practices in the trade where there is generallylittle effort to differentiate rhizomes from roots amongst collectors orbuyers
Goldenseal rhizomesroots were harvested during the summer of 2012from mature reproductive stems (ie stems bearing 2ndash3 leaves) on fourdates corresponding with the following phenological stages (1) July 2 fruitpresent and fully mature foliage green (2) August 7 post-fruit bearingfoliage green (3) September 8 post-fruit bearing some foliage beginning toyellow but not senescing and (4) October 12 90ndash100 yellow foliage ineach colony around 50 of each colony already senesced For each sampledate three rhizomeroot samples were collected from each of the threecolonies for a total of nine samples per date and 36 samples totalRhizomesroots were washed and dried at 35degC for 24ndash36 h until theywere dry enough to break cleanly Dried rhizomesroots were weighed andshipped overnight from Pennsylvania to North Carolina where they wereprepared for analysis
130 E P BURKHART AND G H ZUIDERVEEN
Table1
Wild
goldenseal
(HydrastiscanadensisL)colony
characteristicsandassociated
soilcond
ition
sforsampleplotsinclud
edin
thisstud
yCo
lony
characteristics
Associated
soilcond
ition
s(n
=5)a
Num
berof
ramets
Dry
root
weigh
tsa
(n=50)
pHPh
osph
atekg
haminus1
Potash
kgha
minus1
Magnesium
kgha
minus1
Calcium
kgha
minus1
Catio
nExchange
Capacity
Acidity
perm
2total
Colony-1
28ndash53
1000ndash1500
18g(07)
63(02)
83(20)
202(47)
536(62)
7364
(1026)
17(18)
38(10)
Colony-2
24ndash45
1500ndash3000
17g(04)
70(03)
155(9)
259(52)
615(164)
9518
(1451)
164
(17)
06(13)
Colony-3
19ndash31
100ndash150
09g(02)
66(01)
89(10)
298(27)
492(91)
6064
(1884)
135
(27)
24(13)
a Stand
arddeviations
areinclud
edin
parentheses
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 131
RhizomeRoot Sample Preparation and Chemical Analysis
Dried whole rhizomeroot samples were ground to approximately 60 mesh witha Foss Cyclotectrade mill prior to extraction For each sample about 250 mg ofground material was weighed and placed into a 50 mL centrifuge tube to which250 mL of extraction solvent (waterndashacetonitrilendashphosphoric acid 70 + 30 + 01vvv) was added The sample was capped and mixed using a Vortexreg mixer for10 sec sonicated in a Bransonic 2510 ultrasonic cleaner for 10 min and thenvortexed again following sonication The solid material was settled by centrifu-ging the sample preparation for 5 min at 4000 rpm in a Sorvallreg Legend tabletopcentrifuge An aliquot of the supernatant was then filtered with a 045 microm PTFEsyringe filter In an amber HPLC vial 200 microL of filtered sample was added to800 microL diluent (90 acetonitrile 10 water) and vortexed prior to analysis
Calibration curves were prepared from five mixed standard solutions contain-ing hydrastine berberine palmatine and canadine Standards were prepared inlow-actinic volumetric flasks and diluted with 90 acetonitrile10 waterConcentrations of individual stock standards were calculated based on thereported purity and corrected for chloride content in the case of berberine andpalmatine The linear ranges of the calibration curves were as follows hydrastine103ndash155 microgmLminus1 palmatine 367ndash551 microgmLminus1 berberine 794ndash119 microgmLminus1 andcanadine 564ndash846 microgmLminus1 Curves were all linear with an r2 value of above 0999
High Pressure Liquid Chromatography (HPLC) analysis was performed usingChromeleonreg chromatography management software on a Dionex ICS-3000system equipped with dual pump automatic sample injector and variable wave-length detector The column temperature was maintained at 40degC The analyticalcolumn was an Agilent Zorbax Eclipse XDB-C18 column (150 times 46 mm 35 microm)The mobile phase consisted of 25 mM ammonium formate pH 38 (C) and 01triethylamine in acetonitrile (D) at a flow rate of 075 mL minminus1 The isocraticeluting mobile phase was 70 C and 30 D and injection volume was 10 microLDetection was at 230 nm and total run time was 13 min per injection
Reagents and standards HPLC-grade acetonitrile formic acid (ge98)o-phosphoric acid (85) triethylamine (99) and ammonium formate (99)were obtained from Fisher Scientific HPLC-grade chemical reference standardsberberine chloride (886) (1R 9S) -(-)-B-hydrastine (995) palmatine chlor-ide (tetramethoxyprotoberberine chloride 792) and DL-canadine (tetrahy-droberberine 972) were purchased from Chromadex (Santa Ana CA)
Statistical Analysis
In addition to basic statistical analyses Pearsonrsquos correlation analysis wasused to examine rhizomeroot phytochemistry in relation to soil chemistryand colony Linear regression was used to examine rhizomeroot phytochem-istry in relation to rhizomeroot weight
132 E P BURKHART AND G H ZUIDERVEEN
Analysis of variance was used to compare mean values of total alkaloidconcentration as well as the three individual alkaloids separately for eachharvest time and location Differences between individual harvest data andsites were analyzed using Bonferroni Pairwise Comparisons
Statistical analyses were conducted using SPSS (version 24) and Minitab173 (Minitab 17 Statistical Software 2010)
Results
Variation was observed both within and between colonies for all three alkaloids(Table 2) Total rhizomeroot alkaloid content varied from a low of 32 inAugust (represented by a single sample from Colony-1) to a high of 48 in Julyand October (samples both collected from Colony-3) Both total and individualalkaloid content was associated with harvest timing with the highest alkaloidsobserved at the senescent stage in October (Fig 1) Berberine and hydrastinewere detected in all samples and colonies Canadine however was not detectedin Colony-3 samples from two dates (Aug 7 and Sept 8) Since each sample wasreplicated (n = 3) on each date and in each colony sampled it is unlikely thatthe absence of canadine in colony 3 on two sample dates was the result ofsample processing or analytical errors
Correlation results were insignificant for all soil chemistry parameters exam-ined except for soil Ca and P which were correlated with alkaloid content on twoharvest dates (Table 3) These correlations were both positive (Ca P and cana-dine) and negative (Ca and hydrastine) however revealed no clear trend Themost significant and perhaps interesting correlations were for total alkaloid con-tent root weight and colony at the final harvest date (October) In the case of rootweight there was a negative correlation between rhizomeroot weight and totalalkaloid content indicating that smaller rhizomesroots contained higher totalalkaloid concentrations (Fig 2) For the effect of colony there was a positivecorrelation between colony sampled and total alkaloid content Average rootalkaloid content was the highest in colony-3 on all sample dates
Discussion
The Influence of Harvest Stage and Timing on Quality
The World Health Organization (WHO) Guidelines on Good Agriculturaland Collection Practices (GACP) for Medicinal Plants provide the followingguidance with respect to harvest timing of wild-collected medicinalplants (16)
ldquoMedicinal plant materials should be collected during the appropriateseason or time period to ensure the best possible quality of both sourcematerials and finished products It is well known that the quantitative
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 133
Table2
Alkaloid
levelsin
wild-harvested
goldensealrootsrhizom
esin
relatio
nto
colony
andharveststage(n
=3foreach
plot
oneach
harvestdaten
=36
totalsam
ples)Observatio
nsregardingph
enolog
ical
stageat
thetim
eof
harvestareinclud
edbelow
theharvestdate
July2
2012
Augu
st7
2012
Septem
ber8
2012
Octob
er122
012
Fruitpresentandfully
maturefoliage
green
Post
fruitbearing
foliage
green
Post
fruitbearing
foliage
beginn
ingto
yellow
Foliage
yellowp
lants
senescing
Colony
1Berberineww
25σ02(23ndash27)
21σ02(19ndash23)
23σ02(21ndash26)
25σ05(22ndash31)
Hydrastineww
14σ02(13ndash16)
16σ03(13ndash19)
15σ00(15ndash15)
15σ01(14ndash16)
Canadine
ww
008
σ001
(07-09)
003
σ001
(03-04)
011
σ001
(11-12)
018
σ003
(14-20)
Totalalkaloids
ww
39σ03(36ndash41)
37σ05(32ndash42)
38σ03(35ndash41)
40σ06(37ndash47)
Colony
2Berberineww
23σ01(22ndash25)
22σ01(22ndash23)
25σ01(23ndash26)
29σ01(28ndash30)
Hydrastineww
13σ01(11ndash14)
14σ01(13ndash15)
13σ01(12ndash14)
15σ00(15ndash16)
Canadine
ww
009
σ02(07-09)
008
σ004
(03-11)
009
σ001
(ND-09)
018
σ002
(17-20)
Totalalkaloids
ww
36σ03(34ndash39)
37σ01(35ndash37)
38σ03(35ndash39)
44σ01(44ndash45)
Colony
3Berberineww
25σ02(23ndash27)
22σ03(20ndash25)
26σ01(25ndash27)
28σ01(28ndash29)
Hydrastineww
18σ03(15ndash21)
16σ01(15ndash16)
17σ01(17ndash18)
19σ01(18ndash20)
Canadine
ww
007
σ001
(07-08)
Not
detected
(ND)
Not
detected
(ND)
023
σ004
(20-27)
Totalalkaloids
ww
43σ05(38ndash48)
38σ03(35ndash41)
43σ01(42ndash45)
47σ01(46ndash48)
134 E P BURKHART AND G H ZUIDERVEEN
concentration of biologically active constituents varies with the stage ofplant growth and development [hellip] The best time for collection (qualitypeak season or time of day) should be determined according to thequality and quantity of biologically active constituents rather than thetotal vegetative yield of the targeted medicinal plant partsrdquo [bold empha-sis authorsrsquo]
Goldenseal is included in the United States Pharmacopeia (USP) and USPstandards require a minimum alkaloid content of 25 berberine and 20hydrastine for dried goldenseal rhizomes and roots (17) Out of the 36 wild-harvested goldenseal rhizomeroot samples analyzed in this study only abouthalf (53 n = 19) met this threshold for berberine content while only onesample (28) met the hydrastine threshold (Fig 3) Nearly half of wildrhizomeroots samples harvested before senescence and nearly all harvestedin July and August failed to meet USP minimums for alkaloid content
Figure 1 Alkaloid levels in wild-harvested goldenseal rhizomesroots in relation to colony andharvest date Clockwise from top-left total alkaloid content (berberine hydrastine and canadine)berberine canadine and hydrastine Note the different scales on the vertical axes Harvest datescorresponded with the following phenological stages 0702 = fruit present and fully ripe foliagegreen 0807 = fruit gone foliage green 0908 = foliage beginning to yellow 1012 = foliageyellow plants senescing
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 135
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
Table1
Wild
goldenseal
(HydrastiscanadensisL)colony
characteristicsandassociated
soilcond
ition
sforsampleplotsinclud
edin
thisstud
yCo
lony
characteristics
Associated
soilcond
ition
s(n
=5)a
Num
berof
ramets
Dry
root
weigh
tsa
(n=50)
pHPh
osph
atekg
haminus1
Potash
kgha
minus1
Magnesium
kgha
minus1
Calcium
kgha
minus1
Catio
nExchange
Capacity
Acidity
perm
2total
Colony-1
28ndash53
1000ndash1500
18g(07)
63(02)
83(20)
202(47)
536(62)
7364
(1026)
17(18)
38(10)
Colony-2
24ndash45
1500ndash3000
17g(04)
70(03)
155(9)
259(52)
615(164)
9518
(1451)
164
(17)
06(13)
Colony-3
19ndash31
100ndash150
09g(02)
66(01)
89(10)
298(27)
492(91)
6064
(1884)
135
(27)
24(13)
a Stand
arddeviations
areinclud
edin
parentheses
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 131
RhizomeRoot Sample Preparation and Chemical Analysis
Dried whole rhizomeroot samples were ground to approximately 60 mesh witha Foss Cyclotectrade mill prior to extraction For each sample about 250 mg ofground material was weighed and placed into a 50 mL centrifuge tube to which250 mL of extraction solvent (waterndashacetonitrilendashphosphoric acid 70 + 30 + 01vvv) was added The sample was capped and mixed using a Vortexreg mixer for10 sec sonicated in a Bransonic 2510 ultrasonic cleaner for 10 min and thenvortexed again following sonication The solid material was settled by centrifu-ging the sample preparation for 5 min at 4000 rpm in a Sorvallreg Legend tabletopcentrifuge An aliquot of the supernatant was then filtered with a 045 microm PTFEsyringe filter In an amber HPLC vial 200 microL of filtered sample was added to800 microL diluent (90 acetonitrile 10 water) and vortexed prior to analysis
Calibration curves were prepared from five mixed standard solutions contain-ing hydrastine berberine palmatine and canadine Standards were prepared inlow-actinic volumetric flasks and diluted with 90 acetonitrile10 waterConcentrations of individual stock standards were calculated based on thereported purity and corrected for chloride content in the case of berberine andpalmatine The linear ranges of the calibration curves were as follows hydrastine103ndash155 microgmLminus1 palmatine 367ndash551 microgmLminus1 berberine 794ndash119 microgmLminus1 andcanadine 564ndash846 microgmLminus1 Curves were all linear with an r2 value of above 0999
High Pressure Liquid Chromatography (HPLC) analysis was performed usingChromeleonreg chromatography management software on a Dionex ICS-3000system equipped with dual pump automatic sample injector and variable wave-length detector The column temperature was maintained at 40degC The analyticalcolumn was an Agilent Zorbax Eclipse XDB-C18 column (150 times 46 mm 35 microm)The mobile phase consisted of 25 mM ammonium formate pH 38 (C) and 01triethylamine in acetonitrile (D) at a flow rate of 075 mL minminus1 The isocraticeluting mobile phase was 70 C and 30 D and injection volume was 10 microLDetection was at 230 nm and total run time was 13 min per injection
Reagents and standards HPLC-grade acetonitrile formic acid (ge98)o-phosphoric acid (85) triethylamine (99) and ammonium formate (99)were obtained from Fisher Scientific HPLC-grade chemical reference standardsberberine chloride (886) (1R 9S) -(-)-B-hydrastine (995) palmatine chlor-ide (tetramethoxyprotoberberine chloride 792) and DL-canadine (tetrahy-droberberine 972) were purchased from Chromadex (Santa Ana CA)
Statistical Analysis
In addition to basic statistical analyses Pearsonrsquos correlation analysis wasused to examine rhizomeroot phytochemistry in relation to soil chemistryand colony Linear regression was used to examine rhizomeroot phytochem-istry in relation to rhizomeroot weight
132 E P BURKHART AND G H ZUIDERVEEN
Analysis of variance was used to compare mean values of total alkaloidconcentration as well as the three individual alkaloids separately for eachharvest time and location Differences between individual harvest data andsites were analyzed using Bonferroni Pairwise Comparisons
Statistical analyses were conducted using SPSS (version 24) and Minitab173 (Minitab 17 Statistical Software 2010)
Results
Variation was observed both within and between colonies for all three alkaloids(Table 2) Total rhizomeroot alkaloid content varied from a low of 32 inAugust (represented by a single sample from Colony-1) to a high of 48 in Julyand October (samples both collected from Colony-3) Both total and individualalkaloid content was associated with harvest timing with the highest alkaloidsobserved at the senescent stage in October (Fig 1) Berberine and hydrastinewere detected in all samples and colonies Canadine however was not detectedin Colony-3 samples from two dates (Aug 7 and Sept 8) Since each sample wasreplicated (n = 3) on each date and in each colony sampled it is unlikely thatthe absence of canadine in colony 3 on two sample dates was the result ofsample processing or analytical errors
Correlation results were insignificant for all soil chemistry parameters exam-ined except for soil Ca and P which were correlated with alkaloid content on twoharvest dates (Table 3) These correlations were both positive (Ca P and cana-dine) and negative (Ca and hydrastine) however revealed no clear trend Themost significant and perhaps interesting correlations were for total alkaloid con-tent root weight and colony at the final harvest date (October) In the case of rootweight there was a negative correlation between rhizomeroot weight and totalalkaloid content indicating that smaller rhizomesroots contained higher totalalkaloid concentrations (Fig 2) For the effect of colony there was a positivecorrelation between colony sampled and total alkaloid content Average rootalkaloid content was the highest in colony-3 on all sample dates
Discussion
The Influence of Harvest Stage and Timing on Quality
The World Health Organization (WHO) Guidelines on Good Agriculturaland Collection Practices (GACP) for Medicinal Plants provide the followingguidance with respect to harvest timing of wild-collected medicinalplants (16)
ldquoMedicinal plant materials should be collected during the appropriateseason or time period to ensure the best possible quality of both sourcematerials and finished products It is well known that the quantitative
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 133
Table2
Alkaloid
levelsin
wild-harvested
goldensealrootsrhizom
esin
relatio
nto
colony
andharveststage(n
=3foreach
plot
oneach
harvestdaten
=36
totalsam
ples)Observatio
nsregardingph
enolog
ical
stageat
thetim
eof
harvestareinclud
edbelow
theharvestdate
July2
2012
Augu
st7
2012
Septem
ber8
2012
Octob
er122
012
Fruitpresentandfully
maturefoliage
green
Post
fruitbearing
foliage
green
Post
fruitbearing
foliage
beginn
ingto
yellow
Foliage
yellowp
lants
senescing
Colony
1Berberineww
25σ02(23ndash27)
21σ02(19ndash23)
23σ02(21ndash26)
25σ05(22ndash31)
Hydrastineww
14σ02(13ndash16)
16σ03(13ndash19)
15σ00(15ndash15)
15σ01(14ndash16)
Canadine
ww
008
σ001
(07-09)
003
σ001
(03-04)
011
σ001
(11-12)
018
σ003
(14-20)
Totalalkaloids
ww
39σ03(36ndash41)
37σ05(32ndash42)
38σ03(35ndash41)
40σ06(37ndash47)
Colony
2Berberineww
23σ01(22ndash25)
22σ01(22ndash23)
25σ01(23ndash26)
29σ01(28ndash30)
Hydrastineww
13σ01(11ndash14)
14σ01(13ndash15)
13σ01(12ndash14)
15σ00(15ndash16)
Canadine
ww
009
σ02(07-09)
008
σ004
(03-11)
009
σ001
(ND-09)
018
σ002
(17-20)
Totalalkaloids
ww
36σ03(34ndash39)
37σ01(35ndash37)
38σ03(35ndash39)
44σ01(44ndash45)
Colony
3Berberineww
25σ02(23ndash27)
22σ03(20ndash25)
26σ01(25ndash27)
28σ01(28ndash29)
Hydrastineww
18σ03(15ndash21)
16σ01(15ndash16)
17σ01(17ndash18)
19σ01(18ndash20)
Canadine
ww
007
σ001
(07-08)
Not
detected
(ND)
Not
detected
(ND)
023
σ004
(20-27)
Totalalkaloids
ww
43σ05(38ndash48)
38σ03(35ndash41)
43σ01(42ndash45)
47σ01(46ndash48)
134 E P BURKHART AND G H ZUIDERVEEN
concentration of biologically active constituents varies with the stage ofplant growth and development [hellip] The best time for collection (qualitypeak season or time of day) should be determined according to thequality and quantity of biologically active constituents rather than thetotal vegetative yield of the targeted medicinal plant partsrdquo [bold empha-sis authorsrsquo]
Goldenseal is included in the United States Pharmacopeia (USP) and USPstandards require a minimum alkaloid content of 25 berberine and 20hydrastine for dried goldenseal rhizomes and roots (17) Out of the 36 wild-harvested goldenseal rhizomeroot samples analyzed in this study only abouthalf (53 n = 19) met this threshold for berberine content while only onesample (28) met the hydrastine threshold (Fig 3) Nearly half of wildrhizomeroots samples harvested before senescence and nearly all harvestedin July and August failed to meet USP minimums for alkaloid content
Figure 1 Alkaloid levels in wild-harvested goldenseal rhizomesroots in relation to colony andharvest date Clockwise from top-left total alkaloid content (berberine hydrastine and canadine)berberine canadine and hydrastine Note the different scales on the vertical axes Harvest datescorresponded with the following phenological stages 0702 = fruit present and fully ripe foliagegreen 0807 = fruit gone foliage green 0908 = foliage beginning to yellow 1012 = foliageyellow plants senescing
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 135
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
RhizomeRoot Sample Preparation and Chemical Analysis
Dried whole rhizomeroot samples were ground to approximately 60 mesh witha Foss Cyclotectrade mill prior to extraction For each sample about 250 mg ofground material was weighed and placed into a 50 mL centrifuge tube to which250 mL of extraction solvent (waterndashacetonitrilendashphosphoric acid 70 + 30 + 01vvv) was added The sample was capped and mixed using a Vortexreg mixer for10 sec sonicated in a Bransonic 2510 ultrasonic cleaner for 10 min and thenvortexed again following sonication The solid material was settled by centrifu-ging the sample preparation for 5 min at 4000 rpm in a Sorvallreg Legend tabletopcentrifuge An aliquot of the supernatant was then filtered with a 045 microm PTFEsyringe filter In an amber HPLC vial 200 microL of filtered sample was added to800 microL diluent (90 acetonitrile 10 water) and vortexed prior to analysis
Calibration curves were prepared from five mixed standard solutions contain-ing hydrastine berberine palmatine and canadine Standards were prepared inlow-actinic volumetric flasks and diluted with 90 acetonitrile10 waterConcentrations of individual stock standards were calculated based on thereported purity and corrected for chloride content in the case of berberine andpalmatine The linear ranges of the calibration curves were as follows hydrastine103ndash155 microgmLminus1 palmatine 367ndash551 microgmLminus1 berberine 794ndash119 microgmLminus1 andcanadine 564ndash846 microgmLminus1 Curves were all linear with an r2 value of above 0999
High Pressure Liquid Chromatography (HPLC) analysis was performed usingChromeleonreg chromatography management software on a Dionex ICS-3000system equipped with dual pump automatic sample injector and variable wave-length detector The column temperature was maintained at 40degC The analyticalcolumn was an Agilent Zorbax Eclipse XDB-C18 column (150 times 46 mm 35 microm)The mobile phase consisted of 25 mM ammonium formate pH 38 (C) and 01triethylamine in acetonitrile (D) at a flow rate of 075 mL minminus1 The isocraticeluting mobile phase was 70 C and 30 D and injection volume was 10 microLDetection was at 230 nm and total run time was 13 min per injection
Reagents and standards HPLC-grade acetonitrile formic acid (ge98)o-phosphoric acid (85) triethylamine (99) and ammonium formate (99)were obtained from Fisher Scientific HPLC-grade chemical reference standardsberberine chloride (886) (1R 9S) -(-)-B-hydrastine (995) palmatine chlor-ide (tetramethoxyprotoberberine chloride 792) and DL-canadine (tetrahy-droberberine 972) were purchased from Chromadex (Santa Ana CA)
Statistical Analysis
In addition to basic statistical analyses Pearsonrsquos correlation analysis wasused to examine rhizomeroot phytochemistry in relation to soil chemistryand colony Linear regression was used to examine rhizomeroot phytochem-istry in relation to rhizomeroot weight
132 E P BURKHART AND G H ZUIDERVEEN
Analysis of variance was used to compare mean values of total alkaloidconcentration as well as the three individual alkaloids separately for eachharvest time and location Differences between individual harvest data andsites were analyzed using Bonferroni Pairwise Comparisons
Statistical analyses were conducted using SPSS (version 24) and Minitab173 (Minitab 17 Statistical Software 2010)
Results
Variation was observed both within and between colonies for all three alkaloids(Table 2) Total rhizomeroot alkaloid content varied from a low of 32 inAugust (represented by a single sample from Colony-1) to a high of 48 in Julyand October (samples both collected from Colony-3) Both total and individualalkaloid content was associated with harvest timing with the highest alkaloidsobserved at the senescent stage in October (Fig 1) Berberine and hydrastinewere detected in all samples and colonies Canadine however was not detectedin Colony-3 samples from two dates (Aug 7 and Sept 8) Since each sample wasreplicated (n = 3) on each date and in each colony sampled it is unlikely thatthe absence of canadine in colony 3 on two sample dates was the result ofsample processing or analytical errors
Correlation results were insignificant for all soil chemistry parameters exam-ined except for soil Ca and P which were correlated with alkaloid content on twoharvest dates (Table 3) These correlations were both positive (Ca P and cana-dine) and negative (Ca and hydrastine) however revealed no clear trend Themost significant and perhaps interesting correlations were for total alkaloid con-tent root weight and colony at the final harvest date (October) In the case of rootweight there was a negative correlation between rhizomeroot weight and totalalkaloid content indicating that smaller rhizomesroots contained higher totalalkaloid concentrations (Fig 2) For the effect of colony there was a positivecorrelation between colony sampled and total alkaloid content Average rootalkaloid content was the highest in colony-3 on all sample dates
Discussion
The Influence of Harvest Stage and Timing on Quality
The World Health Organization (WHO) Guidelines on Good Agriculturaland Collection Practices (GACP) for Medicinal Plants provide the followingguidance with respect to harvest timing of wild-collected medicinalplants (16)
ldquoMedicinal plant materials should be collected during the appropriateseason or time period to ensure the best possible quality of both sourcematerials and finished products It is well known that the quantitative
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 133
Table2
Alkaloid
levelsin
wild-harvested
goldensealrootsrhizom
esin
relatio
nto
colony
andharveststage(n
=3foreach
plot
oneach
harvestdaten
=36
totalsam
ples)Observatio
nsregardingph
enolog
ical
stageat
thetim
eof
harvestareinclud
edbelow
theharvestdate
July2
2012
Augu
st7
2012
Septem
ber8
2012
Octob
er122
012
Fruitpresentandfully
maturefoliage
green
Post
fruitbearing
foliage
green
Post
fruitbearing
foliage
beginn
ingto
yellow
Foliage
yellowp
lants
senescing
Colony
1Berberineww
25σ02(23ndash27)
21σ02(19ndash23)
23σ02(21ndash26)
25σ05(22ndash31)
Hydrastineww
14σ02(13ndash16)
16σ03(13ndash19)
15σ00(15ndash15)
15σ01(14ndash16)
Canadine
ww
008
σ001
(07-09)
003
σ001
(03-04)
011
σ001
(11-12)
018
σ003
(14-20)
Totalalkaloids
ww
39σ03(36ndash41)
37σ05(32ndash42)
38σ03(35ndash41)
40σ06(37ndash47)
Colony
2Berberineww
23σ01(22ndash25)
22σ01(22ndash23)
25σ01(23ndash26)
29σ01(28ndash30)
Hydrastineww
13σ01(11ndash14)
14σ01(13ndash15)
13σ01(12ndash14)
15σ00(15ndash16)
Canadine
ww
009
σ02(07-09)
008
σ004
(03-11)
009
σ001
(ND-09)
018
σ002
(17-20)
Totalalkaloids
ww
36σ03(34ndash39)
37σ01(35ndash37)
38σ03(35ndash39)
44σ01(44ndash45)
Colony
3Berberineww
25σ02(23ndash27)
22σ03(20ndash25)
26σ01(25ndash27)
28σ01(28ndash29)
Hydrastineww
18σ03(15ndash21)
16σ01(15ndash16)
17σ01(17ndash18)
19σ01(18ndash20)
Canadine
ww
007
σ001
(07-08)
Not
detected
(ND)
Not
detected
(ND)
023
σ004
(20-27)
Totalalkaloids
ww
43σ05(38ndash48)
38σ03(35ndash41)
43σ01(42ndash45)
47σ01(46ndash48)
134 E P BURKHART AND G H ZUIDERVEEN
concentration of biologically active constituents varies with the stage ofplant growth and development [hellip] The best time for collection (qualitypeak season or time of day) should be determined according to thequality and quantity of biologically active constituents rather than thetotal vegetative yield of the targeted medicinal plant partsrdquo [bold empha-sis authorsrsquo]
Goldenseal is included in the United States Pharmacopeia (USP) and USPstandards require a minimum alkaloid content of 25 berberine and 20hydrastine for dried goldenseal rhizomes and roots (17) Out of the 36 wild-harvested goldenseal rhizomeroot samples analyzed in this study only abouthalf (53 n = 19) met this threshold for berberine content while only onesample (28) met the hydrastine threshold (Fig 3) Nearly half of wildrhizomeroots samples harvested before senescence and nearly all harvestedin July and August failed to meet USP minimums for alkaloid content
Figure 1 Alkaloid levels in wild-harvested goldenseal rhizomesroots in relation to colony andharvest date Clockwise from top-left total alkaloid content (berberine hydrastine and canadine)berberine canadine and hydrastine Note the different scales on the vertical axes Harvest datescorresponded with the following phenological stages 0702 = fruit present and fully ripe foliagegreen 0807 = fruit gone foliage green 0908 = foliage beginning to yellow 1012 = foliageyellow plants senescing
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 135
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
Analysis of variance was used to compare mean values of total alkaloidconcentration as well as the three individual alkaloids separately for eachharvest time and location Differences between individual harvest data andsites were analyzed using Bonferroni Pairwise Comparisons
Statistical analyses were conducted using SPSS (version 24) and Minitab173 (Minitab 17 Statistical Software 2010)
Results
Variation was observed both within and between colonies for all three alkaloids(Table 2) Total rhizomeroot alkaloid content varied from a low of 32 inAugust (represented by a single sample from Colony-1) to a high of 48 in Julyand October (samples both collected from Colony-3) Both total and individualalkaloid content was associated with harvest timing with the highest alkaloidsobserved at the senescent stage in October (Fig 1) Berberine and hydrastinewere detected in all samples and colonies Canadine however was not detectedin Colony-3 samples from two dates (Aug 7 and Sept 8) Since each sample wasreplicated (n = 3) on each date and in each colony sampled it is unlikely thatthe absence of canadine in colony 3 on two sample dates was the result ofsample processing or analytical errors
Correlation results were insignificant for all soil chemistry parameters exam-ined except for soil Ca and P which were correlated with alkaloid content on twoharvest dates (Table 3) These correlations were both positive (Ca P and cana-dine) and negative (Ca and hydrastine) however revealed no clear trend Themost significant and perhaps interesting correlations were for total alkaloid con-tent root weight and colony at the final harvest date (October) In the case of rootweight there was a negative correlation between rhizomeroot weight and totalalkaloid content indicating that smaller rhizomesroots contained higher totalalkaloid concentrations (Fig 2) For the effect of colony there was a positivecorrelation between colony sampled and total alkaloid content Average rootalkaloid content was the highest in colony-3 on all sample dates
Discussion
The Influence of Harvest Stage and Timing on Quality
The World Health Organization (WHO) Guidelines on Good Agriculturaland Collection Practices (GACP) for Medicinal Plants provide the followingguidance with respect to harvest timing of wild-collected medicinalplants (16)
ldquoMedicinal plant materials should be collected during the appropriateseason or time period to ensure the best possible quality of both sourcematerials and finished products It is well known that the quantitative
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 133
Table2
Alkaloid
levelsin
wild-harvested
goldensealrootsrhizom
esin
relatio
nto
colony
andharveststage(n
=3foreach
plot
oneach
harvestdaten
=36
totalsam
ples)Observatio
nsregardingph
enolog
ical
stageat
thetim
eof
harvestareinclud
edbelow
theharvestdate
July2
2012
Augu
st7
2012
Septem
ber8
2012
Octob
er122
012
Fruitpresentandfully
maturefoliage
green
Post
fruitbearing
foliage
green
Post
fruitbearing
foliage
beginn
ingto
yellow
Foliage
yellowp
lants
senescing
Colony
1Berberineww
25σ02(23ndash27)
21σ02(19ndash23)
23σ02(21ndash26)
25σ05(22ndash31)
Hydrastineww
14σ02(13ndash16)
16σ03(13ndash19)
15σ00(15ndash15)
15σ01(14ndash16)
Canadine
ww
008
σ001
(07-09)
003
σ001
(03-04)
011
σ001
(11-12)
018
σ003
(14-20)
Totalalkaloids
ww
39σ03(36ndash41)
37σ05(32ndash42)
38σ03(35ndash41)
40σ06(37ndash47)
Colony
2Berberineww
23σ01(22ndash25)
22σ01(22ndash23)
25σ01(23ndash26)
29σ01(28ndash30)
Hydrastineww
13σ01(11ndash14)
14σ01(13ndash15)
13σ01(12ndash14)
15σ00(15ndash16)
Canadine
ww
009
σ02(07-09)
008
σ004
(03-11)
009
σ001
(ND-09)
018
σ002
(17-20)
Totalalkaloids
ww
36σ03(34ndash39)
37σ01(35ndash37)
38σ03(35ndash39)
44σ01(44ndash45)
Colony
3Berberineww
25σ02(23ndash27)
22σ03(20ndash25)
26σ01(25ndash27)
28σ01(28ndash29)
Hydrastineww
18σ03(15ndash21)
16σ01(15ndash16)
17σ01(17ndash18)
19σ01(18ndash20)
Canadine
ww
007
σ001
(07-08)
Not
detected
(ND)
Not
detected
(ND)
023
σ004
(20-27)
Totalalkaloids
ww
43σ05(38ndash48)
38σ03(35ndash41)
43σ01(42ndash45)
47σ01(46ndash48)
134 E P BURKHART AND G H ZUIDERVEEN
concentration of biologically active constituents varies with the stage ofplant growth and development [hellip] The best time for collection (qualitypeak season or time of day) should be determined according to thequality and quantity of biologically active constituents rather than thetotal vegetative yield of the targeted medicinal plant partsrdquo [bold empha-sis authorsrsquo]
Goldenseal is included in the United States Pharmacopeia (USP) and USPstandards require a minimum alkaloid content of 25 berberine and 20hydrastine for dried goldenseal rhizomes and roots (17) Out of the 36 wild-harvested goldenseal rhizomeroot samples analyzed in this study only abouthalf (53 n = 19) met this threshold for berberine content while only onesample (28) met the hydrastine threshold (Fig 3) Nearly half of wildrhizomeroots samples harvested before senescence and nearly all harvestedin July and August failed to meet USP minimums for alkaloid content
Figure 1 Alkaloid levels in wild-harvested goldenseal rhizomesroots in relation to colony andharvest date Clockwise from top-left total alkaloid content (berberine hydrastine and canadine)berberine canadine and hydrastine Note the different scales on the vertical axes Harvest datescorresponded with the following phenological stages 0702 = fruit present and fully ripe foliagegreen 0807 = fruit gone foliage green 0908 = foliage beginning to yellow 1012 = foliageyellow plants senescing
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 135
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
Table2
Alkaloid
levelsin
wild-harvested
goldensealrootsrhizom
esin
relatio
nto
colony
andharveststage(n
=3foreach
plot
oneach
harvestdaten
=36
totalsam
ples)Observatio
nsregardingph
enolog
ical
stageat
thetim
eof
harvestareinclud
edbelow
theharvestdate
July2
2012
Augu
st7
2012
Septem
ber8
2012
Octob
er122
012
Fruitpresentandfully
maturefoliage
green
Post
fruitbearing
foliage
green
Post
fruitbearing
foliage
beginn
ingto
yellow
Foliage
yellowp
lants
senescing
Colony
1Berberineww
25σ02(23ndash27)
21σ02(19ndash23)
23σ02(21ndash26)
25σ05(22ndash31)
Hydrastineww
14σ02(13ndash16)
16σ03(13ndash19)
15σ00(15ndash15)
15σ01(14ndash16)
Canadine
ww
008
σ001
(07-09)
003
σ001
(03-04)
011
σ001
(11-12)
018
σ003
(14-20)
Totalalkaloids
ww
39σ03(36ndash41)
37σ05(32ndash42)
38σ03(35ndash41)
40σ06(37ndash47)
Colony
2Berberineww
23σ01(22ndash25)
22σ01(22ndash23)
25σ01(23ndash26)
29σ01(28ndash30)
Hydrastineww
13σ01(11ndash14)
14σ01(13ndash15)
13σ01(12ndash14)
15σ00(15ndash16)
Canadine
ww
009
σ02(07-09)
008
σ004
(03-11)
009
σ001
(ND-09)
018
σ002
(17-20)
Totalalkaloids
ww
36σ03(34ndash39)
37σ01(35ndash37)
38σ03(35ndash39)
44σ01(44ndash45)
Colony
3Berberineww
25σ02(23ndash27)
22σ03(20ndash25)
26σ01(25ndash27)
28σ01(28ndash29)
Hydrastineww
18σ03(15ndash21)
16σ01(15ndash16)
17σ01(17ndash18)
19σ01(18ndash20)
Canadine
ww
007
σ001
(07-08)
Not
detected
(ND)
Not
detected
(ND)
023
σ004
(20-27)
Totalalkaloids
ww
43σ05(38ndash48)
38σ03(35ndash41)
43σ01(42ndash45)
47σ01(46ndash48)
134 E P BURKHART AND G H ZUIDERVEEN
concentration of biologically active constituents varies with the stage ofplant growth and development [hellip] The best time for collection (qualitypeak season or time of day) should be determined according to thequality and quantity of biologically active constituents rather than thetotal vegetative yield of the targeted medicinal plant partsrdquo [bold empha-sis authorsrsquo]
Goldenseal is included in the United States Pharmacopeia (USP) and USPstandards require a minimum alkaloid content of 25 berberine and 20hydrastine for dried goldenseal rhizomes and roots (17) Out of the 36 wild-harvested goldenseal rhizomeroot samples analyzed in this study only abouthalf (53 n = 19) met this threshold for berberine content while only onesample (28) met the hydrastine threshold (Fig 3) Nearly half of wildrhizomeroots samples harvested before senescence and nearly all harvestedin July and August failed to meet USP minimums for alkaloid content
Figure 1 Alkaloid levels in wild-harvested goldenseal rhizomesroots in relation to colony andharvest date Clockwise from top-left total alkaloid content (berberine hydrastine and canadine)berberine canadine and hydrastine Note the different scales on the vertical axes Harvest datescorresponded with the following phenological stages 0702 = fruit present and fully ripe foliagegreen 0807 = fruit gone foliage green 0908 = foliage beginning to yellow 1012 = foliageyellow plants senescing
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 135
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
concentration of biologically active constituents varies with the stage ofplant growth and development [hellip] The best time for collection (qualitypeak season or time of day) should be determined according to thequality and quantity of biologically active constituents rather than thetotal vegetative yield of the targeted medicinal plant partsrdquo [bold empha-sis authorsrsquo]
Goldenseal is included in the United States Pharmacopeia (USP) and USPstandards require a minimum alkaloid content of 25 berberine and 20hydrastine for dried goldenseal rhizomes and roots (17) Out of the 36 wild-harvested goldenseal rhizomeroot samples analyzed in this study only abouthalf (53 n = 19) met this threshold for berberine content while only onesample (28) met the hydrastine threshold (Fig 3) Nearly half of wildrhizomeroots samples harvested before senescence and nearly all harvestedin July and August failed to meet USP minimums for alkaloid content
Figure 1 Alkaloid levels in wild-harvested goldenseal rhizomesroots in relation to colony andharvest date Clockwise from top-left total alkaloid content (berberine hydrastine and canadine)berberine canadine and hydrastine Note the different scales on the vertical axes Harvest datescorresponded with the following phenological stages 0702 = fruit present and fully ripe foliagegreen 0807 = fruit gone foliage green 0908 = foliage beginning to yellow 1012 = foliageyellow plants senescing
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 135
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
Table3
Pearsonrsquoscorrelationcoefficient
(r)betweensoilnu
trient
elem
entsrootweigh
tcolonyand
rootrhizomealkaloid
levelsin
wild
goldensealin
relatio
nto
harvestdate
(n=9)
Rootrhizomealkaloid
Berberine
Hydrastine
Canadine
Totalalkaloids
a
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
July
Aug
Sept
Oct
SoilpH
minus0359
0372
minus0072
0390
minus0179
minus0108
minus0492
0047
0478
0479
minus0073
minus0107
minus0282
0128
minus0276
0328
SoilP
minus0625
0300
minus0021
0310
minus0374
minus0340
minus0627
minus0229
0435
073
0minus0263
minus0224
minus0457
minus0081
minus0388
0141
SoilK
minus0629
minus0030
0333
0253
minus0085
minus0619
0063
0376
0018
0241
minus0631
0454
minus0269
minus0447
0231
0335
SoilCa
minus0580
0022
minus0110
0342
minus0410
minus0436
minus074
5minus0317
0541
078
20121
minus0452
minus0507
minus0253
minus0484
0114
SoilMg
minus0476
0000
minus0132
0258
minus0298
minus0393
minus0393
minus0140
0164
0590
0101
minus0109
minus0422
minus0225
minus0316
0115
Root
weigh
tminus0325
minus0034
minus0632
minus0396
minus0383
0096
minus0635
minus090
3
0256
0651
0505
minus085
7
minus0371
0929
minus082
5
minus068
8Co
lony
072
0327
0480
0426
0566
minus0080
0514
082
0
minus0195
minus0351
minus089
2
0632
0469
0095
0616
069
6a Totalalkaloidsisthesum
ofthethreeindividu
alalkaloids
Bolded
values
indicate
thecorrelationissign
ificant
atthe005
level()
or001
level()respectively
136 E P BURKHART AND G H ZUIDERVEEN
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
Figure 2 Goldenseal rhizome and root total alkaloid content for October harvest date in relationto root weight
Figure 3 Effects of Harvest Date on alkaloid levels in goldenseal rootrhizome Bars in eachgraph represent 95 confidence intervals which were calculated using individual standarddeviations Dotted lines represent standards for minimum alkaloid content in goldenseal rootrhizome as listed by the United States Pharmacopeia
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 137
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
The Influence of Environmental Factors on Phytochemistry
A variety of environmental factors influence wild plant phytochemistry instudies of other wild occurring North American medicinal forest plants(1213) Of the factors examined in this study rhizomeroot size and colonywere both correlated with total alkaloid content suggesting that phytochemistrycan vary both within and between populations These differences may be due toenvironmental conditions andor underlying genetic differences not measuredor controlled for in this study The smaller rhizomeroot weight and slightlyhigher alkaloid levels associated with colony-3 may also reflect some underlyingenvironmental influence resulting in higher phytochemical constituentsEnvironmental stress (eg drought light intensity) for example was importantfactor causing higher secondary metabolites in plants (1819)
There was little support for any influence of soil chemistry on alkaloidlevels in this study However the study sample size was small and themethodology was limited to correlation rather than experimental manipula-tion Additional studies of wild goldenseal phytochemistry with increasedreplication may provide better understanding of the influence of soil condi-tions and other environmental factors on rhizomeroot chemistry Theseshould ideally be paired with experimental manipulation of environmentalconditions (eg factorial plots) to provide a more complete understanding ofany underlying interrelationship(s)
Although these findings were limited to a single location in Pennsylvaniathese phytochemistry results are of potential significance throughout the wildgoldenseal harvest region of the United States Total alkaloid content washighest at plant senescence corroborating industry guidance (20) that rootdiggers should harvest roots and rhizomes at senescence (typically during latesummer and fall months) However rhizomeroot collectors will frequentlyharvest wild goldenseal at any time during the growing season a behaviorthat is increasingly visible in part because collectors post pictures of theirharvests on social media platforms (eg Facebook) in ldquoroot diggerrdquo groupsAppalachian root buyers begin advertising prices as early as April and manycollectors begin harvesting goldenseal as soon as plants emerge in the springIn many cases no guidance appears to be offered from buyers to collectors asto when to harvest wild rhizomesroots
Results obtained here support a late season or Fall harvest of goldensealrhizomesroots at the stage when plants are senescing (ie turning yellow) ordormant Findings suggest that scientifically based harvest timing guidanceshould be provided to buyers and in turn goldenseal collectors in order tomaximize quality In doing so harvest timing could also be shifted to a plantstage (eg post-fruit maturation) that allows for plant sexual reproductionThe timing of harvests to permit fruit maturation allowing an adequate
138 E P BURKHART AND G H ZUIDERVEEN
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
recovery interval and attention to site influences are all important compo-nents to sustainable harvesting from wild goldenseal populations (141521)
An unexpected finding in this study was that alkaloid levels were nearly ashigh at fruit maturity (early July) as they were at plant senescence (earlyOctober) This suggests further research is needed to examine early seasonalkaloid levels particularly at flowering and fruiting and the period in betweenIt may be that alkaloid levels fluctuate during the growing season in relation tokey reproductive phenological stages as has been observed in other wild-collected medicinal plants (112223) While early (pre-fruit maturation) seasonharvests would negatively affect sexual reproduction in wild populations earlyseason harvests from cultivated or forest-farmed populations would not neces-sarily present such an ethical dilemma
Acknowledgments
Thanks to Marie Knight (BioNetwork Candler NC) for the laboratory phytochemistrypartnership
Funding
This work was supported by the Pennsylvania Department of Conservation and NaturalResources (PA DCNR) Wild Resource Conservation Program [08321]
References
1 Braun L and M Cohen 2010 Herbs and Natural Supplements An Evidence-BasedGuide 3rd ed Elsevier Chatswood Australia 1221 pp
2 Avula B Y-H Wang and I S Khan 2012 Quantitative determination of alkaloidsfrom roots of Hydrastis canadensis L and dietary supplements using ultra-performanceliquid chromatography with UV detection J AOAC Int 95(5)1398ndash1405
3 Villinski J R E R Dumas H-B Chai J M Pezzuto C K Angerhofer andS Gafner 2003 Antibacterial activity and alkaloid content of Berberis thunbergiiBerberis vulgaris and Hydrastis canadensis Pharm Biol 41(8)551ndash557 doi10108013880200390500768
4 Scazzocchio F M F Cometa L Tomassini and M Palmeri 2001 Antibacterialactivity of Hydrastis canadensis extract and its major isolated alkaloids Planta Med67561ndash564 doi101055s-2001-16493
5 Van Fleet W 1914 (Revised 1936) Goldenseal Under Cultivation United StatesDepartment of Agriculture Farmersrsquo Bulletin No 613 United States Department ofAgriculture Washington DC
6 Burkhart E P and M G Jacobson 2009 Transitioning from wild collection to forestcultivation of indigenous medicinal forest plants in eastern North America is con-strained by lack of profitability Agroforest Syst 76(2)437ndash453 doi101007s10457-008-9173-y
JOURNAL OF HERBS SPICES amp MEDICINAL PLANTS 139
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-
7 Persons W S and J M Davis 2014 Growing and Marketing Ginseng Goldenseal andOther Woodland Medicinals Revised and Expanded New Society Publishers GabriolaIsland BC 508 pp
8 AmericanHerbal Products Association 1997-2005Tonnage Survey of Select North AmericanWild-Harvested Plants American Herbal Products Association Silver Spring MD
9 Kruger S and J Munsell 2014 Root Report Preliminary Results for 2014 VA TechRetrieved November 2018 from httpwwwrootreportfrecvteduindexhtml
10 Crellin J K and J Philpott 1990 A Reference Guide to Medicinal Plants HerbalMedicine Past and Present Duke University Press Durham NC 552pp
11 Salmore A K and M D Hunter 2001 Elevational trends in defense chemistryvegetation and reproduction in Sanguinaria canadensis J Chem Ecol 27(9)1713ndash1727
12 Zheljazkov V D A M Jones B Avula V Maddox and D E Rowe 2009 Lignan andnutrient concentrations in American mayapple (Podophyllum peltatum L) in the east-ern United States HortScience 44(2)349ndash353 doi1021273HORTSCI442349
13 Lim W K W Mudge and F Vermeylen 2005 Effects of population age andcultivation methods on ginsenoside content of wild American ginseng (Panaxquinquefolium) J Agr Food Chem 538498ndash8505 doi101021jf051070y
14 Sanders S and J B McGraw 2005 Harvest recovery of goldenseal Hydrastis canadensisL Am Midl Nat 15387ndash94 doi1016740003-0031(2005)153[0087HROGHC]20CO2
15 Sinclair A and P M Catling 2004 Restoration of Hydrastis canadensis experimentaltest of a disturbance hypothesis after two growing seasons Restor Ecol 12(2)184ndash189doi101111j1061-2971200400297x
16 World Health Organization 2003 Guidelines on good agricultural and collectionpractices (GACP) for medicinal plants Retrieved November 2018 from httpappswhointmedicinedocsendJs4928e5html
17 United States Pharmacopeia 2018 Goldenseal Retrieved November 2018 from httpwwwpharmacopeiacnv29240usp29nf24s0_m35762html
18 Yang L K S Wen X Ruan Y X Zhao F Wei and Q Wang 2018 Response ofplant secondary metabolites to environmental factors Molecules 23(4)762doi103390molecules23040762
19 Akula R and G A Ravishankar 2011 Influence of abiotic stress signals on secondarymetabolites in plants Plant Signaling Behav 6(11)1720ndash1731 doi104161psb61117613
20 American Herbal Products Association 2017 Good agricultural and collection prac-tices and good manufacturing practices for botanical materials American HerbalProducts Association Silver Spring MD Retrieved November 2018 from httpahpaorgResourcesGoodAgriculturalandCollectionPractices(GACP)aspx
21 Albrecht M A and B C McCarthy 2006 Comparative analysis of goldenseal(Hydrastis canadensis L) population re-growth following human harvest implicationsfor conservation Am Midl Nat 156229ndash236 doi1016740003-0031(2006)156[229CAOGHC]20CO2
22 Graf TN KE Levine ME Andrews JS Perlmutter SJ Nielson JM Davis MCWani and NH Oberlies 2007 Variability in the yield of benzophenanthridine alka-loids in wildcrafted vs cultivated bloodroot (Sanguinaria canadensis L) J Agr FoodChem 551205ndash1211 doi101021jf062498f
23 Konchar H L Xiao-Li Y Yong-Ping and E Emshwiller 2011 Phytochemical varia-tion in Fritillaria cirrhosa D Don (Chuan Bei Mu) in relation to plant reproductivestage and timing of harvest Econ Bot 65(3)283ndash294 doi101007s12231-011-9170-3
140 E P BURKHART AND G H ZUIDERVEEN
- Abstract
- Introduction
- Materials and Methods
-
- Study Population and Associated Habitat Conditions
- RhizomeRoot Sampling and Processing
- RhizomeRoot Sample Preparation and Chemical Analysis
- Statistical Analysis
-
- Results
- Discussion
-
- The Influence of Harvest Stage and Timing on Quality
- The Influence of Environmental Factors on Phytochemistry
-
- Acknowledgments
- Funding
- References
-