Peptides, bitter, bitterness elimination

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. 

Elimination of Bitterness of Bitter Peptides by Squid Liver Carboxypeptidase... 

Molecular and enzymatic properties of serine carboxypeptidase (EC.3.4.1.6.1, CPase Top), isolated and refined from the common squid (Todarodes pacificus) liver, were studied. It was found that this enzyme reacts well at the C-terminal position of peptides having hydrophobic amino acids. Because of this property, it was anticipated that this enzyme would have the effect of eliminating bitterness of some peptides. This enzyme was used on bitter peptides prepared by hydrolysis of proteins with pepsin and trypsin. It was found that this CPase Top can eliminate bitter peptides prepared from soy protein and com gluten. 

Because it is quite useful in the food industry, we studied how to produce it inexpensively. We found that this enzyme occurs in the liver of the common squid Toardes pacificus) and determined its characteristic features. We found this enzyme to be useful in eliminating the bitterness of bitter peptides, which is described in this paper. 

Hydrolysis of Bitter Peptides from Soy Protein by CPase Top. Three bitter peptides were incubated with cmde CPase Top at 30 C, pH 4.0, with the bitterness and the amount of liberated amino acids being evaluated throughout incubation. When a 1 % bitter peptide solution from soy protein was incubated with crude CPase Top (enzyme/substrate ratio = 1 /tkat/g), the bitterness diminished as reaction time increased. The bitterness was almost completely eliminated after 15 hr (Fig. 1). The amount of amino acids liberated increased with time of incubation. 

Hydrolysis of Bitter Peptides from Casein by CPase Top. When a 0.5% solution of bitter peptides from casein was incubated with crude CPase Top (enzyme/substrate ratio = 2 /xkat/g), most of the bitterness was eliminated after 2 hr, but some bitterness remained even after 15 hr (Fig. 2). The liberated amino acids increased with time similarly as with the soy bitter peptides. 

It has been demonstrated that CPase Top enzyme is useful for eliminating bitterness originating from soy protein and com gluten. With respect to the bitterness from casein, although the bitterness was not completely eliminated, it was effective in reducing most of the bitterness. It is postulated that the bitterness of peptides is due to the hydrophobic amino acids in the C-terminus position (4). This assumption is necessary for the elimination of bitterness by liberating a hydrophobic amino acid at the C-terminus by the substrate specificity of CPase Top. 

Because of the high effectiveness of CPase Top to eliminate bitterness, and because it is obtained from the edible parts of squids (Japanese traditonal food, Shiokara), the safety of this enzyme is assured. It is hoped that the development of this method for eliminating the bitterness of bitter peptides will be used widely in the food industry. 

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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