..\/lima/ Feed Science and Techn%gy, 13 (1985) 203-214 203Elsevier Science Publishers B. V., Amsterdam - Printed in The Netherlands

", I-ed. ~ NUTRIENT AND TRYPSIN INHIBITOR CONTENT OF

!nco :y HYDROPONICALL Y SPROUTED SOY A BEANS;:

,

and I D.J. PEER and S. LEESON

and Department of Anima/ and Poultry Science, University of Gue/ph, Gue/ph,'; Ontario N1G 2Wl (Canada),,: (Received 7 November 1983; accepted for publication 25 April 1985)

n of ~1"

tube.' ABSTRACT

~hen Peer, D.J. and Leeson, S., 1985. Nutrient and trypsin inhibitor content of hydroponical. :

'al ly sprou ted soya beans. Anim. Feed Sci. Techno/., 13: 203-214. " ,ue "~I tb Soya bean seeds were sprouted hydroponically in the light at 21°C for 1-7 days.

39~ Samples were freeze-dried, ground through a I-mm screen and analyzed for proximate. nutrients, amino acids, minerals and trypsin-inhibitor content. During sprouting, quanti-

t. ties of energy, dry matter (DM), total lipids (TL) and starch (NFE) decreased significant-I Ion Iy (P < 0.05). Protein weight did not change during sprouting. Weight of ash increased

slightly and weight of fibre increased markedly during sprouting- Among the amino39' acids, weight of glutamic acid decreased whilst weights of aspartic acid and leucine in-

,. creased. There was a slight gain in weights of Cu, Na and Zn, these being related to the ,, d mineral content of the water source. Weight of trypsin inhibitor (TI) decreased cubically -

rae . h . d t . t.i Wit Increase sprou mg Ime.

f ::, , Although nutrients are lost to a lesser degree during the sprouting of soya beans as~~o~- '1; , compared with barley, the substantial reduction in TL may mean that the energy value i i:.

,- is more adversely affected. It is not known whether the decrease in TI is great enough J;.. ii. '.,' to render the sprouts safe for direct consumption by farm animals. ~"i I

lDlm. -:;r :,.1

I Ed ';-;!:J ',~: 1;:. ;,i INTRODUCTION : !;tibil- Iy for , ~e~r and Lees~n (1985a,b) have recently described the nutrient co~- ~

l~ position and feeding value of barley grown for 1-7 days under hydropomc I

Ither-' conditions. Although there was a significant loss in most major nutrientswith sprouting, the process may have application to soya beans as an on-

It IInd farm method of processing.

r Hill Owing to the recent interest in soya bean sprouts for human consump-, tion, several researchers have investigated their nutrient concentration

Bull. j Fordham et al. 1975; Ky1en and McCready, 1975; Bates et al., 1977). In. most of these studies soya beans have been sprouted for only 3- 5 days in

;en~ng the dark and no work has been done with beans germinated hydroponi-urine cally in the light in modern growth chambers.~ffect Raw soya beans contain protease inhibitors which affect animal perfor-ion of

,;;~j~),t 0377-8401/85/$03.30 Ig) 1985 Elsevier Science Publishers B. V.

,

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I ~ 204

!

mance (Bowman, 1944; Ham and Sandstedt, 1944; Carver et al., 1946::1 I Gertler et al., ]967; Hull et al., 1968; Latshaw et al., 1974; McNaughton et) al., 1980, 1981). If germination of soya beans reduces the amount of I;) inhibitor, raw sprouts could be given directly to farm animals. Studies on t~ trypsin-inhibitor concentration of germinating soya beans have proved in-, conclusive. Freed and Ryan (1978) al1d Tanimura et al. (1980) reported a! decrease in trypsin inhibitor activity, while Collins and Sanders (19761

and Bates et al. (1977) found little or no difference between sprouted andnon-sprouted soya beans. It was decided to determine the nutrient com.

Il I position and the trypsin inhibitor activity of soya beans sprouted hydro-:. ponically in the light for 1-7 days. Although it is known that vitamin: concentration of grains change during germination (Wai et al., 1974; Vander.~ stoep, 1981) these nutrients were not assayed in this study., I; I. !',' i MATERIALS AND METHODS

I

;t Hydroponic unit"

~ I Soya bean seeds were germinated in a hydroponic sprouting chamberc ~ using the equipment previously described by Peer and Leeson (1985a). The- "~ seed was sprayed with water for 15 min every 4 h and the temperature

maintained at 21 DC. A nutrient solution was not added to the spray water.

Seed preparation

Soya beans (800 g) were pre-wetted with an equal volume of water, thpndistributed on 91.9 X 30.5-cm white plastic trays within the hydroponic

I unit. Growth period was from 1 to 7 days. Samples were collected each day.I freeze-dried, ground through a 1-mm mesh screen and stored in tightly

sealed glass jars at room temperature. Three samples of dry soya beans wereprepared similarly to serve as controls. All nutrient and trypsin-inhibitoranalyses were conducted on the dry, ground samples.

rI

Nutrient analysis 1

Dry matter (DM), crude protein (CP), crude fibre (CF) and mineral can.tents of sprouted and non-sprouted soya beans were determined accordin~to Association of Official Analytical Chemists (1975). Nitrpgen-free extra('t

I (NFE) was calculated by subtracting the summation of CP,. GF, fat and ash!' from 100. Cystine and methionine were determined by the performic at'idi procedure reported by Hirs (1967). The remaining amino acids were ana-I Iyzed according to the instruction manual for the Beckman Amino Acidi Analyzer, Model1]3B (1975).

Dry-matter loss (DML) was determined by sprouting three trays of 300 ~soya bean seed and drying the entire contents of the trays. Three, 300-g

;1;t,,

.

205

~; samples of dry seed were freeze-dried and their average dry-matter weight~t "'~' i served as a control. The DML was assumed to be the difference betweenIf ;,~! the DM weight of the control and test samples.n ,:. !t- Trypsin-inhibitor analysis !

\ 'a . C

i) I, Trypsin-inhibitor (TI) analysis was conducted on dry samples according id to the procedure developed by Hamerstrand et al. (1981). This method in- ,\1- .. valved reaction of sample alkali homogenates with trypsin and benzolyl- ; ;',>- -', DL-arginine-p-nitroanilide in Tris (hydroxymethyl) aminomethane buffer. in" The reaction was terminated after 10 min by the addition of 30% acetic ~

r. acid. Absorbance of solutions was determined at 410 nm (pye Unicam .' i;"; Sp6-5050 ). . 'K: t.. . 1.,

~ '

: ~ ... \

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i Statistical analysis ': ji')~", ! , '.. ',. ,',

A completely randomized design with three replications of eight treat- 'J, 1\ ~1"

ments of growing period was employed. The relationship between growth ;~ ,:;j',:' ,

er period of 1-7 days and nutrient and trypsin-inhibitor concentrations and :1 ~ I'

le weights was ~nalyzed .using orthogonal pol.ynomials (Steel and Torrie, ',,' !J!;

re 1980). RegressIon equatIons were also detennmed. All data from the analy- it! ;';'~isis are expressed on a dry-matter basis, unless otherwise stipulated. Day 0 :ir "t~;equals unsprouted soya beans in all tables. I"' j~~!; :, ~. ,

RESULTS AND DISCUSSION ;::: ;;i~ [fen I. Ii:' J

Ilic Germination of the soya beans was about 96%. The white tip of the ;l :~iflY, protruding radicle is visible after 24 h in the hydroponic unit. It is difficult j i: I Ii;ly to dist.ingui.sh between radicle and hypocotyl until after 3 days of sprouting, ~ ;,; I ~

!re by WhICh tIme the hypocotyl has turned green. Afte~ 5 days, the seed coat I t;~ ~

.or has broken off and the cotyledons have separated slIghtly. The cotyledons " ;!, tare open and the first primary leaf is barely visible after 7 days. \ i~, "

- Proximate anal ysis i! ~

(.'

In- :~ By Day 7, there was an increase in concentrations of ash, crude fibreng.' ICF) and crude protein (CP) by 18.6%, 44.1%, and 7.8%, respectively,iCt .: relative to un sprouted beans. Nitrogen-free extract (NFE) and total lipidsISh ~ \TL) decreased by 4.2% and 20.4%, respectively, by 7 days of sprouting.::id .'~ Similar changes in nutrient profile were observed by Fordham et al. (1975)

,na- ~ and Kylen aJ1d McCready (1975).[:id ; The actual change in nutrient weights during sprouting is not clear until

l dT}'-matter loss has been considered. By Day 7, only 93.1% of the DM,0 g t 78.-1% of the NFE and 14.1% TL remained (Table I).O-g I Oil is a major energy source for the developing soya bean embryo during

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207

germination. The catabolism of oil produces glycerol and fatty acids, which

are used to provide carbohydrates for respiration and cell wall synthesis

(Hsu et al., 1973). Storage carbohydrates are used to a lesser degree. The

21.6% and 25.9% reduction in weights of NFE and TL by Day 7 reflects

the extent to which these energy reserves are utilized by the developing

seedling.

The amount of dry matter lost during sprouting will depend on the

energy source utilized. In barley, the major component by weight is starch

(Peer and Leeson, 1985a). The 34% decrease in amount of starch, therefore,

accounted for the large decrease in dry matter after 7 days of sprouting.

In soya beans, the major energy reserve is oil, which is a more concentrated

source of energy (Hamilton and Vanderstoep, 1979). The 6.9% dry-matter

loss in soya beans, as compared to 18% loss in barley, after 7 days of sprout-

ing reflects this situation.

Unlike barley, soya beans sprouted for 7 days did not change significant-

ly in weight of protein (Table I). Hurst and Sudia (1973) reported no change c;

in nitrogen weight of soya beans germinated in the dark, while soya beans ?,

given a nutrient solution and germinated in the light were able to increase t:,

their nitrogen weight by assimilating the nitrogen from the culture medium. 1~

Fibre weight increased with seedling growth owing to the increase in cell ('

size and numbers, (James, 1940; Mayer and Poljakoff-Mayber, 1975). "!.,11'"

4mino acids :~~. r;IJI';

Table II indicates the weights of amino acids present after sprouting 300 :!

g of soya beans. There was a significant linear decrease in weights of ala- ~l

nine, arginine, threonine, glycine, lysine, proline and serine, while weights ~

of aspartic acid and leucine increased linearly. Weights of histidine, iso- ;1

leucin~, methionine, phenylalanine and tyrosine did not change during ~

sproutmg (Table II). :,;Since the total amount of protein did not change during sprouting, the 11

change .in ~i~o acid weight~ was merely ~ shift in ami.no aci.d profile. I!!

GlutamIc acId IS used as a major source of nitrogen for ammo acId synthe- l'sis (Folkes and Yemm, 1958). Aspartic acid is formed from the decarboxy- ;{

lation of glutamic acid (Bidwell, 1974), which explains the increase in as- :t.:

! partic acid seen here. l'

. .f1 ,\1 in era Is ." .

~~ Table III illustrates the changes in mineral wei~hts which ?ccur whe~ 300

;~ g of soya beans are sprouted for up to 7 days. WeIght of Cu Increased lInear-

~ Iy with increased sprouting time. while weights of Na and Zn increased

~ cubically. The i.ncrease in weights of these minerals could be due to the

;1 amount of Na m the water supply used (Peer and Leeson, 1985a) and

~.. leaching of Cu and Zn from the metallic racks and nozzles of the hydro-

;'~

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It. 208

! TABLE II

.1!; 0 1 23 4 66 7

II Alanine 4.66 4.66 4.79 4.37 4.03 3.26 3.04 3.12i S.E. %0.340 %1.693 %0.072 %0.160 %0.270 %0.286 %0.459 %0.283, !I Arginine 7.21 7.09 7.20 6.64 6.93 6.06 4.60 4.44

S.E. %0.494 %2.460 %0.161 %0.234 to.266 to.218 %0.656 %0.339

, Aspartici acid 12.3 12.2 9.05 13.2 14..2 14.0 17.6 21.9I S.E. to.78 %4.21 t3.96 :to.286 t1.11 t1.07 t2.67 t2.11

, Cystine 0.77 1.09 0.94 0.88 0.83 0.73 0.80 0.81: S.E. 10.094 :to.116 to.008 to.007 %0.019 to.036 %0.069 %0.261,

Glutamicacid 19.1 19.4 19.6 17.3 14.8 10.4 8.67 7.92S.E. t1.20 %6.69 %0.211 to.68 %1.01 t1.10 %1.39 %0.876

Glycine 4.69 4.67 4.76 4.37 4.02 3.11 2.83 2.91S.E. to.327 t1.698 to.036 to.146 %0.262 to.308 10.383 to.238

Hbtidine 2.91 3.07 3.03 2.89 2.87 2.35 2.49 2.66S.E. to.178 to.823 %0.021 to.076 to.194 to.134 10.319 10.090

Isoleucine 4.46 6.20 4.97 4.67 4.48 4.19 4.36 4.67S.E. to.162 t1.766 to.063 %0.144 to.276 %0.026 to.483 to.178

Leucine 4.46 6.20 4.97 4.67 7.97 7.02 6.86 7.08S.E. to.162 t 1.766 to.063 to.144 %0.432 to.340 to.821 to.409

Lysine 7.06 6.96 7.08 6.60 6.95 4.48 4.14 4.29S.E. to.504 t2.337 to.094 to.177 to.381 to.419 %0.677 %0.266

Methionine 1.64 1.86 1.69 1.61 1.42 1.29 1.36 1.48S.E. :to.164 to.206 to.027 %0.021 to.039 %0.073 to.078 to.443

Phenylalanine 6.61 6.36 6.50 6.14 4.94 4.34 4.31 4.37S.E. %0.444 t1.800 10.064 to.146 to.317 10.219 to.481 to.219

Proline 5.64 5.66 6.61 6.72 4.85 3.80 3.24 3.23S.E. %0.376 t1.843 %0.090 %0.262 %0.341 to.348 to.453 %0.219

Serine 4.98 6.10 6.30 4.95 4.51 3.61 3.46 3.61S.E. to.290 %1.687 to.061 to.167 to.254 to.309 to.490 %0.286

Threonine 4.31 4.23 4.33 3.96 3.62 2.97 2.90 3.00S.E. %0.303 t 1.484 to.110 to.097 to.173 to.288 10.416 %0.170

I Tyrosine 3.66 4.02 4.12 3.89 3.97 3.46 3.10 3.07S.E. to.201 t1.360 to.059 %0.136 to.300 to.208 to.423 %0.192

Valine 6.17 6.82 6.90 6.06 5.90 4.76 4.64 6.11S.E. %0.123 t1.684 %0.464 :to.206 to.442 %0.407 %0.609 %0.204 -

i; ponic chamber. Weight of Fe decreased linearly, while weight of P decreased~ cubically over time. Calcium and Mg weights did not change during sprout-

I ing (Table III). Since no nutrient solution was used, the increase in mineralweights must be due to the amount of minerals in the water supply.

i Table IV summarizes changes in the weights of major nutrients when 1

~ kg soya bean dry matter is germinated.

209

~

Regression Regression RSD R' .coefficient 1 S.E. oonstant P < j

- i: -0.28,x 10.086 4.98 0.960 0.33 .', 0.003 ,." :,: -0.47,x j; 0.127 7.63 1.420 0.38 \..: 0.001 I, ~ ' ~

". ," +1.30,x 10.375 9.74 4.209 0.35 f

0.002 ~,

-0.02,x j; 0.017 0.93 0.191 0.07 ~.0.219 1

-1.93,x 10.357 21.4 4.00 0.57 ~:'!0.0001 ;4, !.

1(. I'f c-0.32,x 10.085 5.04 0.956 0.39 ' : j0.001 .: .I-:

; ,. I!

-0.08,x 10.046 3.06 0.517 0.12 . ;.

, .,0 099 ' I'. , ,,:-0.07,x 10.091 4.85 1.016 0.03 ,:;;~ I

0.450 _: ~+0.43,x 10.121 4.52 1.36 0.37 j'

0.002 , :

~,i.-0.50,x j; 0.125 7.55 1.401 0.42 , :0.0006 '~ i I,

:1, ~-0.05,x 10.027 1.67 0.302 0.12 ~.!' ~

I0.100 '. . t

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-0.20,x j; 0.093 5.64 1.045 0.18 i' r~ i0.042 i; .'~'I

-0.41,x 10.103 6.11 1.153 0.42 1:1':0.0007 i ,

j"'I-0.29,x j; 0.091 5.42 1.023 0.31 ,::;0.005 Ii' ~I

-0.24,x 10.078 4.50 0.873 0.30 ~I;ll~0.005 ~ 't [

,\ ~-0.13,x 10.073 4.10 0.824 0.12 !: ..,'!

0 101 j i ". ~, !' .-0.24,x 10.102 6.51 1.150 0.20 i;, .'

0.026 I ;

age Trypsin inhibitor

ouler There was a highly significant (P < 0.002) cubic relationship between

growth period and TI weight (Table V). The regression equation is y =~n 1 :' 58.6 - 4.46x + 2.12x2 - 0.23xJ, where y = TI (mg g-1 dry matter) and

~~ .t = time (days). Day 7 sprouts contained 12.7% less TI by weight than the. unsprouted beans.1 Several researchers have investigated the effect of germination on TI

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.Jec ~ E E~~ ~~ EI- ~ S., .,s ~.~.~...~.~.~.~.~.

«( .. .~ :s~ .,~ ~~ ~~ c:~ .~ ~~ c:~!-o ~ ~ u,,; ().,; fI.";~";~";~"; Z,,; ~,,; N";

;.. I

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211

TABLE IV

~utrient weights (g) during the sprouting of 1 kg of soya bean dry matter-Day DM Ash CF CP Lya NFE T.L

Mean S.E. Mean S.E. Mean S.E. Mean S.E. Mean S.E. Mean S.E. Mean S.E.-0 1000 fO.O 55.2 fO.43 52.1 fO.52 398 f 1.9 25.1 tl.80 272 f 5.6 222 14.71 98913.4 54.7 10.59 50.7 fO.52 391 f 1.7 24.8 f8.33 267 '2.5 226 f2.12 994 11.1 56.4.. 1.41 51.4 f 2.82 397 f 2.9 25.3 10.34 265 .. 5.8 223 f 4.43 967 f1.6 57.4 10.83 51.5 ..0.90 388 1 1.4 23.2 to.63 264 1 4.8 206 ..4.7~ 96511.7 57.9 11.52 578 ..0.67 391 1 4.2 21.2 11.37 255 1 5.5 204 14.05 94210.8 59.4 11.30 602 10.49 400 f11.8 14.3 fO.48 233 112.2 190 f2.86 929 t 2.6 60.8 f 1.02 67.0 f 4.91 397 t 3.4 14.8 f 2.06 242 110.6 163 t 3.2'; 930 f8.7 63.1 10.75 87.8 f4.32 401 f 6.1 15.3 fO.94 213 f 8.40 164 t5.13-DM z Dry matter; CF ~ crude fibre; CP - crude protein; Lya - lysine, NFE - nitrogen free extract;TL a ,totallipida; S.E. - standard error of the mean. ~

;t::T ABLE V r j,,. JTrypsin inhibitor (TI) weights of soya beans sprouted for up to 7 days 11, !- ,i::."Day TI (mg I-I dry matter) \1:'

"';,Mean S.E.;. - .~.,!

0 58.6:t 0.91 f;,

1 56.1:t 1.26 I i.

f;' ,2 55.9 t 2.36 ~fl

~3 58.3:t 0.84 :t','1.t 60.1:t 1.92 ~ ,I.Ii '5 59.1:t 1.45 lil~

6 58.7:t 1.90 }1 ~7 51.1:t 2.53 ;, ~ic(i~

Day 0 = unsprouted soya beans. Ij,' f~Regression equation: y = 58.6 - 4.47x + 2.12x' - 0.23.1:3, where y = TI (mg g-1 dry mat- (".f'ter) and x = time (days); P < 0.002, R' = 0.51; residua! standard deviation = t 2.777. 1:1 f

J' , 't.' t11 activity in soya beans. Often, there is a reduction in the activity of TI, rl!: .

with the decrease being dependent on the growth period and the method of ~i;.analysis employed. Collins and Sanders (1976), after germinating soya : ibeans for 3 days, reported a small decrease in the amount of TI activity, ;which was attributed, in part, to the leaching of TI during daily washings.

'" Freed and Ryan (1978), using an immunoelectrophoretic TI assay, found\: a 13% and 50% reduction in the concentration of Kunitz TI on a dry and

\\'et basis, respectively, after sprouting soya beans in the dark for 9 days.Bates et al. (1977) reported a 33% reduction in TI on a fresh basis, after 4days of sprouting.

The decrease in the amount of TI on a dry-matter basis during the hydro-, ; panic sprouting of soya beans for 7 days seen here is comparable to the

I: 212ii~ results of Freed and Ryan (1978). If this decrease during sprouting reduces

; the weight of TI to a safe level, sprouted soya beans could be given to:: livestock as a protein supplement directly on the farm, since protein is not

: lost duru1g sprouting and the dry-matter loss is small. However, unlike thl'i situation seen w~th barley. (Pe:r and Leeson, 1985a), this loss of. DM repre-

sents a substantial reduction m TL and hence energy values W111 be morE'

I adversely affected.Limited research concerning the feeding value of sprouted soya beaJls

has been conducted in the past. Everson et al. (1944) compared growthrate of rats fed on raw immature and mature soya beans sprouted for 60 hat 26°C. Rats fed on raw sprouted soya beans gained more weight thanrats fed on raw mature soya beans, but less than those fed on raw immaturebeans. Autoclaving improved the digestibility of all three bean preparations-Mattingly and Bird (1945) gave 5-day sprouts to rats and chicks. The ratexperiment confirmed the results of Everson et al. (1944), although thE'chick trial resulted in no difference in growth rate for chicks fed on sproutedor mature beans. Desikacher and De (1950), using rats, found that thE'biological value of 3-day sprouted soya beans was higher than that of ra\\-soya beans, but observed no difference in TI levels of raw and sproutedbeans. These feeding trials suggest that sprouting of soya beans improvestheir feeding value; even if weight of TI is not affected.

The TI analysis of the sprouted soya beans, reported herein, was con-ducted according to Hamerstrand et al. (1981), so limited results are avail.able for direct comparison owing to the methodology of TI assay. KakadE'et al. (1974) obtained a value of 12.0 trypsin units inhibited (TIU) per m~sample for commercial soy protein concentrate. Zamora and Veum (1979)obtained values of 1.2 TIU mg-1 and 7.1 TIU mg-t of dry sample for heatedunfermented and fermented soya beans, respectively. When the resultsfrom the analyses reported herein are converted to TIU mg-1 dry matter.a value of 93.5 TIU mg-1 is obtained for Day 7 sprouts, representing thE'lowest level of TI of all sprouts analyzed. Hamerstrand et al. (1981) ob-tained a value of 28.2 mg TI g- I dry matter for a heat-treated sample ofsoya bean meal. The val~e for Day 7 sprouts, when expressed in the same

units, is 51.1 mg TI g-1 dry matter.From these comparisons, it would seem that the amount of TI in sproutS

is still too high for their direct consumption by livestock. Feeding trialswith different sprout ages and species of livestock need to be conductedto determine the digestibility of the sprouts and their effect on animalperformance. In view of the results obtained with regular soya beans andthe fact that sprouting has only a limited influence on amount of Tl, it

I does not seem advisable or economical to use raw sprouted soya beans a~

. an animal feed at this time.

I: ACKNOWLEDGEMENT

This work was supported by the Ontario Ministry of Agriculture and

Food.

, ;:\~ :!( 1:' il' \' ,~;

213 ,i I.IMI' t !t;

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