Bitterness of Peptides: Amino Acid Composition and Chain Length

6 Bitterness of Peptides: Amino Acid Composition and

Downloaded by UNIV OF CALIFORNIA SANTA BARBARA on September 24, 2013 | http://pubs.acs.org Publication Date: December 14, 1979 | doi: 10.1021/bk-1979-0115.ch006

Chain Length KARL HEINZ NEY Unilever Forschungsgesellschaft mbH Behringstrasse 154, D-2000 Hamburg 50, West Germany

During our work on taste of foods we synthesized a series of peptides and soon came to the opinion, that the bitterness of peptides is caused by the hydrophobic action of amino acid side chains. Here I think some remarks on hydrophobic interactions (1) would be appropriate. It is generally accepted now, that hydrophobic interactions are a contributing factor to protein behaviour and esp. to the formation of the secondary structure, e.g. helix. This means, that as shown in Figure 1 hydrophobic residues of the amino acids in a peptide are driven together by clusters of water molecules and so the secondary structure of a peptide or protein is formed. For the transfer from the helical to the stretched form, Tanford (2) found that the transfer free energy of the total protein results from the sum of the contributions of the single amino acid residues.

Δ =ΣΔ F

f

The Δf values of the single amino acids given in Table I were determined by Tanford (2) from solubility data and they represent a measure of the hydrophobicity of an amino a c i d residue* Please note, that the values are r e l a t i v e to the methyl groups of glycine which i s taken to be 0* In Table II the taste of some "isomeric -dipeptide s i s described* A l l the dipeptides are composed of the natural 1amino acids, as are a l l the examples, that w i l l follow l a t e r * It i s i n t e r e s t i n g to note, that the p o s i t i o n of the amino acid has no influence on b i t t e r n e s s ( 3.) · The value Q given represents the average hydrophobicity of a peptide and i s obtained by summing t h e ^ f-values of the amino a c i d residues of a peptide and d i v i d i n g by the number of the amino a c i d residues* 11

«.

Σ

Δ

'

η 0-8412-0526-4/79/47-115-149$06.25/0 © 1979 American Chemical Society In Food Taste Chemistry; Boudreau, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1979.


150

FOOD TASTE CHEMISTRY

Figure 1. Hydrophobic interactions

In Food Taste Chemistry; Boudreau, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1979.

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151

Bitterness of Peptides Table I

^ f - v a l u e s of the side chains of amino acids, repre senting t h e i r hydrophobicity, according to Tanford /Vy f-value


Amino a c i d Glycine Serine Threonine Histidine Aspartic acid Glutamic a c i d Arginine Alanine Methionine Lysine Valine Leucine Proline Phenylalanine Tyrosine Isoleucine Tryptophan

cal/mol 0

40 440 500 540

550 730 730

13ΟΟ 15ΟΟ 1690

2420 262Ο

265Ο 287Ο 297Ο

3OOO

Table II Taste and Q-value of "Isomeric Peptide Gly-Ala Ala-Gly Glu-Ala Ala-Glu Met-Ala Ala-Met Leu-Met Met-Leu Ala-Phe Phe-Ala

bitter

11

dipeptides

non-bitter

Q

X X

365 365

X X

640

X X

IO15

640 1015

X

i860

X X

i860

X

1690 1690

You w i l l have noticed i n Table I I , that the Q-values are much higher i n the case of b i t t e r dipeptides compared with the non-bitter dipeptides. Table III shows a s e r i e s of non-bitter dipeptides. It should be noted here that the Q-values are a l l below 1 3 Ο Ο . We can compare t h i s with values of the following Table IV,


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which l i s t s a s e r i e s of b i t t e r dipeptides with Q-values above 1400. Table I I I Q-values of further non-bitter dipeptides


Peptide Glu-Val Glu-Lys Gly-Gly Gly-Asp Ala-Asp Ser-Asp Ser-Glu Val-Asp Val-Glu Ala-Ala Asp-Asp Glu-Asp Glu-Gly Gly-Ser Gly-Thr Val-Gly Lys-Glu

non-bitter X X X X X X X X X X X

Q 1120 1025 0 270 635 290 295 1115 1120 730 540

X X

545 225 20 220

X X

1025

X X

845

Table IV Q-values of f u r t h e r b i t t e r dipeptides Peptide Leu-Tyr Leu-Leu Arg-Pro Asp-Phe Asp-Tyr Val-Leu Gly-Ile Gly-Phe Gly-Try Val-Val Glu-Phe Gly-Tyr Ala-Leu

bitter X X X X X X X X X X X X X

Q 2645 2420 1665 1595 1705 2055 1485 1325 1500 1690 1600 1435 1575

On Table V a s e r i e s of b i t t e r d i - and t r i p e p t i d e s synthesized by S h i r a i s h i (60) i s given* It follows therefore, that i n the case of peptides from the natural 1-amino acids no b i t t e r n e s s occurs when Q i s


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Bitterness of Peptides

153 only when the value Q

below 13°°i b i t t e r n e s s occuring exceeds 1400 ( 3 ) · Table V

Q-values of further b i t t e r d i - and t r i p e p t i d e s


Peptide

Q 1665 1665 2600 I69O 2145 2145 2510 2510 2785 2785 2735 2735 1665 2050 2625 2625 1750 1750

Pro-Ala Ala-Pro Pro-Pro Val-Val Val-Pro Pro-Val Leu-Pro Pro-Leu lie-Pro Pro-Ile Tyr-Pro Pro-Tyr Arg-Pro Lys-Pro Pro-Phe Phe-Pro Gly-Phe-Pro Phe-Pro-Gly

If the Q-values l i e between 13ΟΟ and 1400 no p r e d i c t i o n can be made of the peptides b i t t e r n e s s * It was i n t e r e s t i n g to see i f our method can also be applied to i n d i v i d u a l 1-amino acids* This means, that η s 1 and consequently i n

Χ

Δ ί

Q equals f. As can be seen from Table VI, the i n d i v i d u a l 1-amino acids also follow the r u l e * The only exceptions are l y s i n e and p r o l i n e , which have too high Q-values f o r non-bitter amino acids* However, a s l i g h t b i t t e r note i s detectable i n the otherwise sweetish taste of l y s i n e and p r o l i n e * In t h i s context i t i s worth taking a b r i e f look at the question of flavour enhancing q u a l i t i e s of glutamate, generally substances of the UMAMI-type as described by Shizuko Yamaguchi i n her c o n t r i b u t i o n to t h i s symposium* Kuninaka (4) proposed the following s t r u c t u r a l element f o r flavour i n t e n s i f i e r s : H -C-C-O-C-COOH Ά '

ι A—


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but he pointed out, that the element i s not absolute, as otherwise glutaraine would have been a f l a v o u r enhancer. Table VI Q-values and taste of i n d i v i d u a l 1-amino acids


1-Amino-Acid

non-bitter

bitter

Glycine (opt* non a c t i v e ) Serine Threonine Histidine Aspartic acid Glutamic a c i d Arginine Alanine Methionine Lysine Valine Leucine Proline Phenylalanine Isoleucine Tryptophan

Q

X

0

X X

40 440

X X X X X X X X X X X X X

500 540 550 730 730 1300 1500 1690 2420 2620 2650 2970 3000

Based on a s e r i e s of examples from p u b l i c a t i o n s and patents, I would l i k e to discuss, however, the hypothesis, that i n order to achieve f l a v o u r enhancing, glutamatel i k e e f f e c t , a compound must have two negative charges* These should be located 3 to 9, p r e f e r a b l y 4 to 6 C-atoms from one another* Instead of a C-atom, a S-atom can also occur* The presence of an α-amino group i n 1-configuration has a d d i t i o n a l f l a v o u r enhancing e f f e c t (5)I Θ

00C[-0-3 C00 © n

n

=

1-7

The f a c t s , on which our assumption i s based, are given i n Table VII. An extension of our hypothesis to f l a v o u r - a c t i v e nucleotides seems to be p o s s i b l e because these compounds also have negative charges at two d i f f e r e n t points of the molecule: i n a d d i t i o n to the a c i d i c phosphate group, they also possess a phenolic hydrogen. I t seems that the negative charges can a l s o be on a peptide chain. Fujimaki describes the b i t t e r masking a c t i o n of peptides r i c h i n glutamyl residues (29) and the i s o l a t i o n and i d e n t i f i c a t i o n of a c i d i c oligopeptides from a f l a v o u r i n t e n s i f y i n g f r a c t i o n from f i s h protein hydrolysate (32)·


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Table VII Facts on which our hypothesis i s based No. 1)


2) 3) 4) 5) 6)

7) 8)

9) 10)

11) 12)

13)

Fact Acc. to J . Solms only the d i s s o c i a t e d form of 1-glutamic a c i d i s flavouractive 1-Cystein-S-sulfonic a c i d has a s i m i l a r e f f e c t to that of MSG 1-Homocysteic a c i d has a s i m i l a r e f f e c t to that of MSG 1-Aspartic a c i d has a s i m i l a r e f f e c t to that of MSG l-oc-Aminoadipic a c i d has a s i m i l a r e f f e c t to that of MSG Adipic a c i d makes the b i t t e r a f t e r taste of sweetners Succinic a c i d i s comparable i n i t s e f f e c t with that of MSG The flavour enhancing properties of the f r u i t acids - v i z . malic a c i d , t a r t a r i c a c i d and c i t r i c a c i d - are known Lemon j u i c e i n t e n s i f i e s the flavour of strawberries The tastes of leguminose products are improved by t r e a t i n g with solutions of more than two of the following acids: malic a c i d , l a c t i c a c i d , t a r t a r i c acid, c i t r i c acid The odour of g a r l i c can be reduced by adding fumaric a c i d or maleic a c i d Glutathione (γ-glutamylcysteinylglycine) i s reported to contribute towards the flavour of meat as an enhancer The diammonium s a l t s of the dicarbo x y l i c acids from malonic to sebacic a c i d are used as table s a l t - s u b s t i t u t e s

Lit. (6,92)

(7,8) (9,l£,JLl) (12)

(11) (12,14,15) (11,16)

(I7il8,19»2£)

(21)

(22)

(23)

(24)

(25)

Furthermore, i t may be, that the well known action of polyphosphates i n increasing the taste of chicken meat (31) or processed cheese (3.2) can be traced back on the negative charges of the polyphosphates. Asparagine, unlike a s p a r t i c a c i d i s completely lacking any flavour i n t e n s i f y i n g property, because one of the a c i d i c groups was eliminated. Also the f i n d i n g s on the d e r i v a t i v e s of glutamic (33) acid are very i n t e r e s t i n g ! i f the glutamic a c i d i s



156


e s t e r i f i e d or amidified, the flavour i n t e n s i f y i n g properties are l o s t . I would l i k e now to return to the topic of the Q-values dimensions. Since Tanford gave h i s A, f-values i n c a l o r i e s , the dimension of the Q-value i s c a l r e s " · A l l the Q-values mentioned i n t h i s paper are given i n these dimensions. Up to t h i s point only amino acids, d i - and t r i p e p t i d e s had been considered. However, we wanted to see i f the Qconcept could be extended to higher peptides as w e l l . Stepwise we synthesized a heptapeptide and followed the change of the taste. The following Table VIII shows t h i s synthesis (5).

Table VIII Peptide Glu-Lys Met-Glu-Lys Ala-Met-Glu-Lys Ile-Ala-Met-Glu-Lys Asp-Ile-Ala-Met-Glu-Lys Glu-Asp-Ile-Ala-Met-Glu-Lys

bitter

non-bitter

Q

X

1025 1116 1020 1410 1265

X

II63

X X X X

As you can see, the d i - , t r i - and tetrapeptides have Q-values below 1300 and are not b i t t e r . In the step leading to the pentapeptide the introduction of the strong hydrophobic i s o l e u c i n e with i t s high ^ \ f - v a l u e of 297° confers a b i t t e r n e s s and correspondingly a Q-value of l 4 l 0 . When a s p a r t i c acid with i t s l o w / \ f - v a l u e of 540 i s added, i n the next step, the hexapeptide again becomes non-bitter with a Q-value of 1265· Glutamic acid - with a low/\ f-value of 550 - added i n the f i n a l step gives a non-bitter heptapeptide with a Q-value of 1163. This example shows the influence of the amino a c i d residues as a polypeptide i s synthesized and i t gives a good demonstration of the p o s s i b i l i t i e s of the method and we regarded i t as a c r u c i a l experiment. Whereas i n t h i s example the b i t t e r taste during the synthesis of peptides was followed, Table IX gives according to Minamiura (34) the degradation of a b i t t e r peptide obtained from the action of B a c i l l u s s u b t i l i s on c a s e i n . Table IX Degradation of a b i t t e r peptide obtained from the a c t i o n of B a c i l l u s s u b t i l i s on Casein Peptide Arg-Gly-Pro-Pro-Phe-Ile-Val Gly-Pro-Pro-Phe-Ile-Val Arg-Gly-Pro-Pro-Phe Gly-Pro-Pro-Phe

Q 1891

2085 1716 1963


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NEY

Ve now wanted to extend the range to peptides of longer chain length* As you see from Table X, the Q-method works well up to eikosapeptides. Table X Q-values and taste of t r i - to eikosapeptides bitter


Peptide Val-Val-Val Ala-Ser-Phe Val-Val-Glu Pro-Gly-Gly-Glu Ser-Pro-Pro-Pro-Gly Gly-Pro-Phe-Pro-Val-Ile Val-Ser-Glu-Glu-Glu-AspIle-Ala-Met-Glu-Lys Lys-Asp-Glu-Glu-Glu-GluVal-Glu-Ser-Gly-Pro-AspAla-Pro-Leu-Pro-Ala-Glu Phe-Phe-Val-Ala-Pro-PhePro-Glu-Val-Phe-Glu-LysPhe-Ala-Leu-Pro-Glu-TyrLeu-Lys

non-bitter

Q 1690

X

1140

X X X

X

1310 787 1508 2085 815

X

1121

X X

1912

X

Kauffmann and Kossel (35) Isolated a s e r i e s of o l i g o peptides from spinach and these are shown i n Table XI· Table XI Q-values of non-bitter oligopeptides from spinach Peptide

Q

Glu-Gly Glu-(Gly,Ser) Gly-(Glu,Ser) Ala-(Glu,Gly-Ser) Glu-(Gly,Gly,Ala) Asp-(Glu,Gly,Ser,Ser) Ser-(Gly,Gly,Thr) Ala-(Glu,Glu,Gly,Ser)

225 196 196 330 320 234 120 374

As you see, the Q-values are extremely low and therefore the peptides non b i t t e r * As given i n Table XII the Q-raethod was also successf u l l y applied i n the case of b i t t e r peptides from the rennet-sensitive sequence of K-casein ( 3 6 ) ·

Ve published the Q-hypothesis i n 1971 (2)

a n d

t n u s

established f o r the f i r s t time a q u a n t i t a t i v e r e l a t i o n s h i p between the amino a c i d composition of a peptide and i t s b i t t e r n e s s , as we introduced the Tanford values and so


158


opened the way

f o r a c a l c u l a t i o n of b i t t e r n e s s . Table XII

B i t t e r peptides synthesized acc. to the rennets e n s i t i v e sequence of K-Casein


Peptide

Q

Ser-Leu-Phe-Met-Ala Lys-Hi s-Pro-Pro-Hi s-LeuSer-Phe Lys-Hi s-Pro-Pro-Hi s-LeuSer-Phe-Met-Ala-Ile-ProPro-Lys-Lys

1428 1726 2001

In Table XIII we have c o l l a t e d other former postulates for b i t t e r n e s s of peptides. The r e s u l t s are i n agreement with the Q-rule, f o r example the sequence Gly-Pro-Pro-Phe postulated by Minamiura (34) to be the core of the b i t t e r n e s s has a high Q-value of 1963. Table XIII Former postulated requirements

f o r b i t t e r n e s s of peptides

Amino acid or sequence inducing b i t t e r n e s s

Lit.

-Leucine40 -Try-Phe-Leu34 -Gly-Pro-Pro-Phe-2 neutral amino acids with large a l k y l groups C ^3 -1 neutral amino acid with a large a l k y l group 41 C ^ 3 with a short a l k y l group -1 neutral amino a c i d + 1 aromatic amino acid -1 neutral amino a c i d + 1 basic amino a c i d

Q 2420 2647 1963

high high high high open

The same holds f o r the sequence Tyr-Phe-Leu, postulated by Fujimaki (40) to be e s s e n t i a l f o r b i t t e r n e s s , here the Q-value i s 2647. Also leucine, postulated e a r l i e r by F u j i maki ( 3 7 i 3 8 , 3 9 ) to be e s s e n t i a l f o r b i t t e r n e s s , has a. /\ fvalue of 2420 and therefore contributes considerably to the Q-value of any peptide of which i t forms a p a r t . Also the postulates of Kirimura (kl) correspond to our theory. It follows that the Q-concept represents a general r u l e f o r p r e d i c t i n g b i t t e r n e s s under which the previously c i t e d postulates are s p e c i a l cases. The dipeptide glutamyl-tyrosine i s b i t t e r below pH 10,



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NEY

159


and not b i t t e r above pH 10· This coincides with the d i s s o c i a t i o n of the phenolic hydroxy1 group of t y r o s i n e . The corresponding dipeptide glutamyl-phenylalanine has no phenolic group, and i s b i t t e r over the whole pH-range. Q of t h i s compound i s 1660 (42). The Q-concept has been assessed and accepted by the s c i e n t i s t s ( 4 3 - 6 l ) working i n t h i s f i e l d . Series of b i t t e r peptides have been i s o l a t e d from enzymatic hydrolysates of proteins, esp. c a s e i n and soybean protein. Figure 2 gives the sequence (6l.i^2i63) of tt ~ c a s e i n which represents about 40 % of c a s e i n - and shows the b i t t e r peptides, that have been i s o l a t e d . According to Mercier (63) the polypeptide chain of a ^ - c a s e i n contains 3 hydrophobic regions, v i z . 1-44, 90-113 and 132-199· I t i s very i n t e r esting that a l l b i t t e r peptides derived from α - c a s e i n and i s o l a t e d by the groups of Mercier (63,), Matoba 16j5 ), B e l i t z ( 6 6 ) , Solms ( 4 7 ) , H i l l (67) are located i n these hydrophobic regions and have Q-values above 1400. Figure 3 gives the sequence of β-casein - which r e presents 30 % of c a s e i n - and the b i t t e r peptides derived from i t and i s o l a t e d by the groups of Clegg ( 4 9 ) , K l o s t e r meyer (46), Gordon (64). Here also the Q-values of the b i t t e r peptides are above 1400. Please note, that no s p e c i a l single amino a c i d or sequence i s needed to impart the b i t t e r t a s t e . From soybean protein hydrolysates several s e r i e s of b i t t e r peptides have been i s o l a t e d . As an example Table XIV shows b i t t e r peptides i s o l a t e d by Fujimaki (69., 7 £ ) . As before the high Q-values are evident. sl

Table XIV B i t t e r peptides from peptic soya p r o t e i n hydrolysates Peptide Leu-Phe Leu-Lys Arg-Leu Arg-Leu-Leu Phe-Ile-Ile-Glu-Gly-Val

Q 2535 I960 1575 1856 1766

From peptic Zein hydrolysates, Wieser and B e l i t z (71) i s o l a t e d b i t t e r peptides which are given i n Table XV to gether with the corresponding high Q-values. Regarding the whole p i c t u r e of enzymatic hydrolysates we came to the conclusion, that c e r t a i n proteins are more prone to y i e l d b i t t e r peptides than others. Therefore we t r i e d to t r a n s f e r our method also to proteins as w e l l . This would enable a p r e d i c t i o n to be made as to whether i n the


160


Θ

®

Arg-Pro-Lys-His-Pro-lle-Lys-His-Gln-Gly-Leu-Pro-Gln-Glu-Val-Leu-Asn-Glu-Asn-Leu Û-- 1830, Matoba *Belitz *

Mercier *

®

© -Leu-Arg-Phe-Phe-Val-Ala-Pro-Phe-Pro-Gln-Val-Phe-Gty-Lys-Glu-Lys-Val-Asn-Gln-Leu65


-

66

J

\Q = 1933,Felissier"*\

- Ser-Lys -Asp-He-

1

.

\Qr1309, Pelissier **\ 6

Q = 1984,

Θ

63

Gly

Hamilton * 67

$9t-ΆΛρ

.....

©

Q: 1670,

©

Hamilton * 67

Q= 1638, Mercier *]

®

63

^(f9-Clfli 62

-Leu-Lys

© -Pro-Val-Val-Val-Pro-Pro-Phe-Leu-Glu-Pro-Glu-Vol-Met-Gly-Vol-Ser-Lys-Vol-Lys-Glu@

@

\Qf>Q.™iSsier">\

@

-Ala-Met-Ala-Pro-Lys-His-Lys-Glu-Met-Pro-Phe-Pro-Lys-Tyr-Pro-Val-Gin-Pro-Phe-Thr® -Gln-Ser-Glu-Ser-Leu-Thr-Leu-Thr-Asp-Val-Glu-Asn-Leu-His-Leu-Pro-Pro-Leu-Leu-Leu-

Θ

- Gin -Ser - Tyr-Met-His

- Gin - Pro-His - Gin-Pro-Leu-Pro-Pro

®

@

- Thr - Val-Met-Phe-Pro-Pro-Glu

® -Ser-Val-Leu-Ser-Leu-Ser-Gln-Ser-Lys-Val-Leu-Pro-Val-Pro-Glu-Lys-Ala-Val-Pro-Tyr-

®

®

\Q--2497,Pelissier">\

-

-

@

Pro-Gin-Arg-Asp-Met-Pro-lle-Glu-Ala-Phe-Leu-Leu-Tyr-Gln-Gln-Pro-Val-Leu-Gly-ProQ = 1700, Gordon * 64

Q=2085,Pelissier > 62

-Val-Arg-Gly-Pro-Phe-Pro-lle-lle-Val I Q-- 1700,

@

Gordon * 64

Figure 3.

Bitter peptides from β-casein

Table XV B i t t e r peptides from peptic Zein hydrolysates Peptide Ala-Ile-Ala Ala-Ala-Leu Leu-Gin-Leu Leu-Glu-Leu Leu-Val-Leu Leu-Pro-Phe-Asn-Glu-Leu Leu-Pro-Phe-Ser-Glu-Leu

Q 1477

1293 1613 1797 2177 1682 1688


|

162


course of a h y d r o l y s i s of a protein, b i t t e r peptides would be formed (72). Generally pure proteins are considered to be without any taste* Secondary, t e r t i a r y and quaternary structures generally prevent a taste impression being ob tained* The following Table XVI gives the Q-values of some proteins* Table XVI Downloaded by UNIV OF CALIFORNIA SANTA BARBARA on September 24, 2013 | http://pubs.acs.org Publication Date: December 14, 1979 | doi: 10.1021/bk-1979-0115.ch006

Q-values of proteins and b i t t e r hydrolysates derived Protein Collagen Gelatin Bovine muscular tissue Wheat Gluten Zein Soybean p r o t e i n Potato p r o t e i n Casein

Q 1280 1280 1300

1420 1480 1540 1567 1600

B i t t e r hydrolysates known no no no yes yes yes yes yes

It i s i n t e r e s t i n g to see that proteins with high Qvalues above 1400 as e.g. soybean protein, c a s e i n wheat gluten, potato protein, Zein are the "parents" of b i t t e r peptides, whereas no b i t t e r peptides have been i s o l a t e d from hydrolysates prepared from c o l l a g e n or g e l a t i n , proteins with Q-values below 13ΟΟ. Petrischek (74) confirmed that the p r o t e i n and not the protease i s responsible f o r the occurence of b i t t e r pep t i d e s * However, when the "parent" proteins are not b i t t e r but the peptides derived from them are b i t t e r , the questions a r i s e as to why t h i s i s so and as to where we must place the molecular weight l i m i t s of peptides with Q > 1400 that are also not b i t t e r * An i n d i c a t i o n of the values to be expected can be obtained from the r e s u l t s of our synthesis of b i t t e r peptides with Q 1400 and molecular weights up to 2000 Dalton* Fujimaki (75) i s o l a t e d from the peptic hydrolysate of soybean p r o t e i n a non-dialysable b i t t e r peptide of a molecular weight of about 2800 Dalton* P i l n i k (76) found by the p r o t e o l y s i s of soybean p r o t e i n i n a membrane-filtration apparatus that no b i t t e r peptides e x i s t e d with molecular weights above 6000 Dalton* Clegg (49) obtained from digests of Casein with Papain a b i t t e r peptide having a molecular weight of about 3OOO Dalton* Fujimaki (77*78) condensed b i t t e r soybean p r o t e i n hydro l y s a t e s i n a Plastein-Reaction (79) and obtained non-bitter p r o t e i n - l i k e products, unfortunately without determination of molecular weights* We studied the influence of chain length on the b i t t e r ness of peptides by gel permeation chromatography of


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enzymatic protein hydrolysates (80). Table XVII sums up the r e s u l t s of these experiments* We can conclude, that a l i m i t of about 6000 Dalton can be placed on the molecular weight* Table XVII


Molecular weights and tastes of enzymatic Parent Protein

Soybean p r o t e i n Soybean protein Casein Casein Wheat Gluten Potato protein Potato protein Gelatine

Q

1540 1540

16Ο5 1605 1420 1567 1567 1280

Molecular weight of hydrolysate i n Dalton

hydrolysates Taste b i t t e r nonbitter

4000

X

125ΟΟ 4000 8000

X

5000 400

X X X X

8000 3000

X X

Above t h i s molecular weight, also peptides with a Qvalue above 1400 w i l l no longer e x h i b i t b i t t e r taste* I t i s c l e a r therefore, that 2 ways e x i s t to come to non-bitter protein hydrolysates* As demonstrated i n Figure 4 a) choice of the s t a r t i n g material, t h i s means proteins with Q-values below 13ΟΟ b) choice of the working conditions, t h i s means, i f the Qvalue of the s t a r t i n g protein i s above 1400, c a r e f u l h y d r o l y s i s to obtain peptides with main molecular weights of above 6000 Dalton* It should be pointed out, that we were concerned with presence or absence of bitterness* B i t t e r n e s s i n terms of sensory threshold values or b i t t e r n e s s r a t i n g s was not assessed* What i s now the current state of a f f a i r s of the Q-ruleî As mentioned i t has been accepted and applied by the s c i e n t i s t s working i n t h i s f i e l d . The most comprehensive and c a r e f u l assessment of the Q-rule has been c a r r i e d out by Guigoz and Solms (54). They found that the r u l e can be applied to the majority of the b i t t e r peptides known and observed, that only peptides containing glycine sometime do not comply f u l l y with the rule* They therefore propose, that glycine should be l e f t out of the c a l c u l a t i o n s , which then gives Q-values higher than 1400 f o r a l l b i t t e r peptides* Guigoz and Solms conclude that the Q-values should be a usef u l assessment of the r e l a t i o n s h i p between amino acid composition and the b i t t e r taste of peptides* Wieser and B e l i t z (Ql) have suggested a very i n t e r e s t i n g extension of the r u l e * They obtained the b i t t e r n e s s threshold values of



164


a) Q < 1300 non bitter Mol. wt. Da I ton to*

ι to* Amino acid residues Protein

Amino acid

b) Q >1400 6000 Mol. wt. Dalton bitter

;

Amino acid

non bitter

W

4

to* Amino acid residues Protein

Figure 4. Molecular weight, average hydrophobicity Q, and bitter taste of peptides



6.

NEY


165

d i - and t r i p e p t i d e s by c a l c u l a t i n g the sum of the hydro phobicity of the "backbone" peptide c o n s i s t i n g only of glycine residues adding to i t the hydrophobicities of the side chains* In t h i s way an estimation of the threshold values of d i - and t r i p e p t i d e s was obtained* We now i n v e s t i g a t e d the hypothesis i f the b i t t e r n e s s of l i p i d s - and carbohydrates - could also be linked to hydro phobic i n t e r a c t i o n s (82,

Bitterness of Peptides: Amino Acid Composition and Chain Length

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