Protein hydrolysates bitterness

Enzymatic hydrolysates of various proteins have a bitter taste, which may be one of the main drawbacks to their use in food. Arai el al. [90] showed that the bitterness of peptides from soybean protein hydrolysates was reduced by treatment of Aspergillus acid carboxypeptidase from A. saitoi. Significant amounts of free leucine and phenylalanine were liberated by Aspergillus carboxypeptidase from the tetracosapeptide of the peptic hydrolysate of soybean as a compound having a bitter taste. Furthermore, the bitter peptide fractions obtained from peptic hydrolysates of casein, fish protein, and soybean protein were treated with wheat carboxypeptidase W [91], The bitterness of the peptides lessened with an increase in free amino acids. Carboxypeptidase W can eliminate bitter tastes in enzymatic proteins and is commercially available for food processing. 

It seems that the negative charges can also be on a peptide chain. Fujimaki describes the bitter masking action of peptides rich in glutamyl residues (29 ) and the isolation and identification of acidic oligopeptides from a flavour-intensifying fraction from fish protein hydrolysate (30). 

From soybean protein hydrolysates several series of bitter peptides have been isolated. As an example Table XIV shows bitter peptides isolated by Fujimaki (69., 70) As before the high Q-values are evident. 

Bitter peptides from peptic soya protein hydrolysates... 

Fujimaki (77,78) condensed bitter soybean protein hydrolysates in a Plastein-Reaction (79) and obtained non-bitter protein-like products, unfortunately without determination of molecular weights. 

Above this molecular weight, also peptides with a Q-value above 1400 will no longer exhibit bitter taste. It is clear therefore, that 2 ways exist to come to non-bitter protein hydrolysates. As demonstrated in Figure 4... 

Ney, K.H. and RezzlafF, G. 1986. A computer program predicting the bitterness of peptides, especially in protein hydrolysates, based on amino acid composition and chain length (computer Q) In Shelf Life of Foods and Beverages , Proceedings of the 4th International Flavour Conference (G. Charalambous ed.), pp. 543-550. Elsevier, The Netherlands. 

Kukman, I.L., Zelenik-Blatnik, M., and Abram, V. 1995. Isolation of low-molecular-mass hydrophobic bitter peptides in soybean protein hydrolysates by reversed-phase high-performance liquid chromatography. J. Chromatogr. 704, 113-120. 

Stevenson, D.E., Ofman, D.J., Morgan, K.R., and Stanley, R.A. 1998. Protease-catalyzed condensation of peptides as a potential means to reduce the bitter taste of hydrophobic peptides found in protein hydrolysates. Enzyme Microb. Technol. 22, 100-110. 

The extent of hydrolysis of protein hydrolysates is measured by the ratio of the amount of amino nitrogen to the total amount of nitrogen present in the raw material (AN/ TN ratio). Highly hydrolyzed materials have AN/TN ratios of 0.50 to 0.60. To obtain the desired level of hydrolysis in a protein, a combination of proteases is selected. Serine protease prepared from Bacillus lichenifor-mis has broad specificity and some preference for terminal hydrophobic amino acids. Peptides containing terminal hydrophobic amino acids cause bitterness. Usually a mixture of different proteases is employed. The hydrolysis reaction is terminated by adjust-... 

Not only analytical or preparative separations can be performed on cyclodextrin polymer columns, but also undesired components can be removed from aqueous solutions, bitter tasting substances (narin-gin, limonin) can be removed or at least their concentration can be strongly reduced after treatment of citrus juice with cyclodextrin polymers in batch or column process (65,66). Phenylalanine can be eliminated from dietetic protein hydrolysates (67), water-soluble organic substances (e.g. polychlorinated biphenyls (68), 2-naph-talenecarboxylate or phenol can be removed from aqueous solutions (e.g. from pharmaceutical wastewater) by polystyrene-cyclodextrin derivatives (69), by 8-cyclodextrin immobilized on cellulose (70) or by 6-cyclodextrin-polyurethane polymer (71). 

Taste Evaluation. The four hydrolysates produced in pilot plant were evaluated for bitter taste by the laboratory s taste panel. Tasting took place in the taste panel room which is equipped with separate booths, and the panel has been selected and trained specially for discrimination of bitterness. The panel was instructed to rank two samples and four bitter-tasting standards, containing 20, 40, 80, and 160 ppm quinine hydrochloride dissolved in non-bitter iso-electric soluble soy protein hydrolysate (5, 6). 20 ppm quinine hydrochloride in this solvent had in previous experiments been established as the panel s threshold value. The protein (N x 6.25) concentration in the samples and standards was 4.0% and pH was adjusted to 6.5 with 4 N NaOH or 6 N HC1. 

Table I. Results from the Organoleptic Evaluation of the Bitterness of Four Soy Protein Hydrolysates...

Pure proteins are generally tasteless, though the predominant taste of protein hydrolysates is bitter. 

Sodium acetate is used as a buffering agent in various intramuscular, intravenous, topical, ophthalmic, nasal, oral, otic, and subcutaneous formulations. It may be used to reduce the bitterness of oral pharmaceuticals. It can be used to enhance the antimicrobial properties of formulations it has been shown to inhibit the growth of S. aureus and E. coli, but not C. albicans in protein hydrolysate solutions. It is widely used in the food industry as a preservative. Sodium acetate has also been used therapeutically for the treatment of metabolic acidosis in premature infants, and in hemodialysis solutions. ... 

Pancreatin Peptidase Neutral Proteinase Exopeptidase Animal Pancreas Aspergillus ssp. Bitter-free Protein Hydrolysates... 

Exopeptidases are attractive as means of de-bittering protein hydrolysate made with endo-peptidases and a considerable research is invested in these types of enzymes for flavour generation purposes. 

An industrial process has been developed for production of isoelectric soluble soy protein hydrolysate with no bitterness and a bland taste (13). The raw material may be acid washed soy white flakes, soy protein concentrate or soy protein isolate. The raw material is hydrolyzed by the alkaline protease ALCALASE to a specified degree of hydrolysis using the pH-stat at pH 8.0... 

Solubilization of Protein. Fish protein concentrate has high nutritional quality as determined both from its essential amino acid composition and from animal feeding experiments. Unfortunately, the concentrate is quite insoluble in water because of its denaturation by the solvent extraction method used in processing thus it contributes no functional properties to a food and must be used in bakery products primarily. A potentially useful method of solubilizing the protein is by proteolysis (9-12). As is the case with protein hydrolysates of casein and soybean protein, bitter peptides are formed during the hydrolysis. Papain and ficin produce more of these bitter peptides than does Pronase, for example (12). Pronase was found to produce a more brothy taste (13). A possible method of removing the bitter peptides is to convert the concentrated protein hydrolysate to plastein by further proteolytic enzyme action (14) to remove the bitter peptides. 

The resulting protein hydrolysates, though free of nonprotein impurities, are accompanied by another problem in most cases—a problem caused by a bitterness resulting from protease action on proteins. As discussed below, the resynthesis reaction leading to plastein formation... 

Enzymes Unsaturated soybean phospholipid Encapsulation in the inner aqueous core Alleviate bitterness in protein hydrolysates [164]... 

Bitter Peptides Identified in Protein Hydrolysates and Cheese Products... 

Shipe, W.F., G.F. Senyk, R.A. Ledford, D.K. Handler, E.T. Wolff, Elavor and chemical evaluations of fresh and aged market milk, J. Dairy Sci., 63(Suppl. 1), p. 43, 1980. Chandran, R.C., K.M. Shahani, Milk lipases a review, J. Dairy Sci., 47, p. 471, 1964. Kwak, H., I.J. Jeon, S.K. Pemg, Statistical patterns of lipase activities on the release of short-chain fatty acids in Cheddar cheese slurries, J. Food Sci., 54, p. 1559, 1989. Murry, T.K., B.E. Baker, Studies on protein hydrolysis I — preliminary observations on the taste of enzymic protein hydrolysates, J. Sci. Food Agric., 3, p. 470, 1952. Fujimaki, M., M. Yamashita, Y. Okazawa, S. Aral, Diffusible bitter peptides in peptic hydrolyzate of soybean protein, Agric. Biol. Chem., 32, p. 794, 1968. 

Arai, S., M. Yamashita, H. Kato, M. Fujimaki, Applying proteolytic enzymes on soybean Part V. A nondialzable bitter peptide hydrolyzate of soybean protein and its bitterness in relation to the chemical structure, Agric. Biol. Chem., 34, p. 729, 1970. Arai, S., The bitter flavor due to peptides or protein hydrolysates and its control by bitterness — masking with acidic oligopeptides, in The Analysis and Control of Less Desirable Flavors in Foods and Beverages, G. Charalambous, Ed., Acad. Press, New York. p. 1, 1980. 

Manley, C.H., Havor Interactions Workshop, IFT sponsored, Orlando, FL, 2003. Blake, A., The world of meat flavours. Food Manuf., 57, 9, p. 65, 1982. Prendergast, K., Protein hydrolysis — a review. Food Trade Rev., 44, p. 14, 1974. Roozen, J.P., The bitterness of protein hydrolysates, in Progress in Flavour Research, J. Adda, Ed., Applied Sci. Publ., London, 1979, p. 321. 

Elimination of the bitter taste from a protein hydrolysate is also possible without incorporation of hydrophihc amino acids. Bitter-tasting peptides, such as Leu-Phe, which are released by partial hydrolysis of protein, react preferentially in the subsequent plastein reaction and are incorporated into higher molecular weight peptides with a neutral taste. 

Methods for eliminating bitter peptides in partial protein hydrolysates are known, but they cause a significant loss of essential amino acids. These procedures usually include additional enzymatic hydrolysis under controlled conditions (a shorter time for the hydrolysis leads to higher peptides that are not bitter) and a selection of suitable proteases, such as aminopeptidases, carboxypeptidases and some other proteases. Enzymes of plant and microbial origin have been successfully used for this purpose. For example, the intracellular peptidases from Lactococcus lactis ssp. cremoris and Brevibacterium linens, which have high proteolytic activity, successfully hydrolyse bitter peptides in cheeses. 

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