Investigation of common butterbur

(Petasytes hybridus L., Asteraceae) non-polar extract

Dušanka Runjaić-Antić1, Vanja Tadić2, Ivana Arsic3, Zorica Vujić3

2Alternativa medica, Manufacturer of phytopreparations, Loznica, Serbia;

4Institute for Medicinal Plant Research „Dr Josif Pančić“, Belgrade, Serbia; 1Faculty of Medicine,Department of Pharmacy, University of Niš, Serbia;

3Faculty of Pharmacy, University of Belgrade, Serbia;

Common butterbur (Petasytes hybridus L., Asteraceae) has

numerous applications and expanding clinical interest in

the treatment of migraines, allergic rhinitis, and asthma

(spasmolytic action). The combined anti-spasmodic, anti-

inflammatory and calcium-channel blocking effects of P.

hybridus proved a rationale base for its use in the

prophylaxis of migraine. The rhizomes (Petasitidis

rhizoma), and leaves (Petasitidis folium) are used for

medicinal purposes. Butterbur extracts contain

sesquiterpene esters, polifenolic acids, flavonoids, lignans,

essential oils and in the minor concentration pyrrolizidine

alkaloids. The literature data revealed that the main

constituents of butterbur are sesquiterpene esters of

petasin and isopetasin.

The plant material was collected on the mountain Jagodnja (Serbia).

The extraction procedure was performed with methylene chloride, and obtained

extract was analyzed for total polifenolic compounds (mg GA/g dry mass),

flavonoids (%), tannins (%) and petasine (%) content. The total phenolics content-TF

was determined by the Folin-Ciocalteu method. The percentage content of flavonoids

CF was calculated using the method described in DAB 10. The percentage content of

tannins-TC ware investigated using the method described in the Ph. Eur. 6.0. (Table

1). The total sesquiterpene content were determined according to Ph. Eor. 6.0, taking

S-petasin as reference standard. HPLC fingerprint of the extract (Figure 2 and 3) was

achieved by HPLC (Agilent Technologies 1200). Detection was performed using

Diode Array Detector (DAD), and the chromatograms were recorded at λ = 260 and

325 nm (for sesquiterpene lactones – petasines, and poliphenolic compounds,

respectively). HPLC separation of components was achieved using a LiChrospher

100 RP 18e (5 μm), 250 × 4 mm i.d. column, with a flow rate of 1 mL/min and

mobile phase, A [500 mL of H2O plus 9.8 mL of 85% H3PO4 (w/w)], B (MeCN),

elution, combination of gradient mode: 90-75% A, 0-25 min; isocratic 75% A, 25-30

min; 75-55% A, 30-46 min). The sample was prepared dissolving 118.6mg of the

extract in 10 mL of MeOH, filtered through 0.2 μm PTFE filters prior to HPLC

analysis. The injected volume was 4 μL. Standard solution of S-petasin was prepared

at a final concentration of 0.01 mg/mL.

Figure. 2. Comparative HPLC spectra of

S-petasin (a), MeOH (b) and CH2Cl2 (c)

common butterbur extracts (from 55-

65min). UV spectra of S-petasin (d), and

the compounds with Rt close to Rt of S-

petasin (e and f, from MeOH and CH2Cl2

extracts respectively)

Conclusion Further investigation should proceed in aim to determine the main

sesquiterpene lacones present in the species from this geografic region.

Besides, the profile of phenolic compounds would be a challenge to

determined, as well.

For more information please do not hesitate to contact us at:

E-mail: amedica2@opentelekom.com.rs

Phone: +381 11 7120 982

Fax.: + 381 11 3120 381

The aim of our investigation was to determine the chemical

profile of Petasitides rhizoma non-polar extract regarding

the content of total polifenolic compounds, including

flavonoids and tannins and total petasine content.

Investigated extract subjected to HPLC analysis revealed the absence of S-

petasine, as well as in the MeOH extracts, made to check the obtained

results (Figure 2).

min56 58 60 62 64

mAU

0

200

400

DAD1 C, Sig=260,8 Ref=600,100 (VMT\VT_01042013 2013-04-01 14-14-25\PETASH000100.D)

57.93

5 58.38

3

59.45

7

59.84

8

min56 58 60 62 64

mAU

0

25

50

DAD1 C, Sig=260,8 Ref=600,100 (VMT\VT_01042013 2013-04-01 14-14-25\PETASH000106.D)

55.27

2

56.53

0

57.06

7

58.18

2

58.89

2 59.61

4

60.46

1

61.02

0

62.36

6

62.81

9

64.44

8

min56 58 60 62 64

mAU

0

50

100

DAD1 C, Sig=260,8 Ref=600,100 (VMT\VT_01042013 2013-04-01 14-14-25\PETASH000112.D)

54.50

1

55.15

5

56.40

8

57.13

4

57.73

2

58.23

4

58.98

0 59

.207

59.59

9

60.05

8

60.53

6

61.10

9

62.00

4

62.48

2

63.02

1

63.35

7

64.53

0

nm250 300 350 400 450 500 550 600

mAU

0

50

100

150

200

DAD1, 57.886 (192 mAU, - ) of PETASH000100.D DAD1, 57.933 (235 mAU, - ) of PETASH000100.D

a

b

c

d

e

f

Introduction

Aim

Material and method

Results

Petsytidis rhizoma (Petasytes hybridus L., Asteraceae)

Total phenols (mg GA/g ekstrakta) 29.00

Flavonoids content (%) 0.10

Tannins content (%) 0.89

Sesquiterpene lactones content 2.10

Table 1. Total phenolic, tannins, flavonoid and

sesquiterpene lactones content in non-polar extract of

common butterbur root (Petasites hybridus, L.,

Asteraceae). The results were expressed by dry

weight.

Literature:

A. Bodensieck, O. Kunert, E. Haslinger, R. Bauer. Helv Chim Acta (2007), 90, 183-194; 2. R.

Chizzola, T. Langer, C. Franz. Planta Med. 2006, 72, 1254-1256

Blois, M.S., 1958. Antioxidant determinations by the use of a stable free radical. Nature. 181, 1199-

1200.

European Pharmacopoeia. 6th edition. 2004. Strasbourg: Council of Europe.

Weißdornblätter mit Blüten. In: Deutsches Arzneibuch, Band 2, Monographien R-Z; Amlitche

Ausgabe Deutcher Apotheker Verlag. Stuttgart, Frankfurt: Govi-Verlag GmbH; 1991

min10 20 30 40 50 60

mAU

0

20

40

60

80

100

120

DAD1 A, Sig=260,4 Ref=550,100 (VMT\VT_02042013 2013-04-02 13-12-42\PETASH000132.D)

min10 20 30 40 50 60 70

mAU

0

50

100

150

200

250

300

350

DAD1 B, Sig=325,4 Ref=600,100 (VMT\VT_01042013 2013-04-01 14-14-25\PETASH000106.D)

Figure. 1. HPLC fingerprint of P. hybridus root non-polar extract

recorded on 260 and 325 nm

HPLC chromatogram of the investigated extract poinetd that the P.

hybridus root extract was rich in sesquiterpene lactopnes (Figure 1). The

preliminary investigation indicated phenolic compounds presence, as

well. Namely, moderate phenolic (mg GA/g) and tannins (%) content

were found (29.0 and 0.89, respectively). Flavonoid content was low

(0.1%). Contrary to this, high sesquiterpene lactones content was

revealed (Table 1).