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Multi-phosphopeptide-enriched casein hydrolyzate and preparation method thereof

A casein hydrolyzate and proteolytic enzyme technology, applied in peptide sources, animal/human peptides, fermentation, etc., can solve problems such as sequence structure analysis, achieve high activity, good research samples, and good application prospects

Inactive Publication Date: 2011-01-19
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] At present, researchers have reported the separation and purification of casein phosphopeptides, but some have failed to analyze their sequence structures, and some have prepared various types of phosphopeptides

Method used

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  • Multi-phosphopeptide-enriched casein hydrolyzate and preparation method thereof
  • Multi-phosphopeptide-enriched casein hydrolyzate and preparation method thereof
  • Multi-phosphopeptide-enriched casein hydrolyzate and preparation method thereof

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Embodiment 1

[0024] Prepare 1 L of 20 g / L cow's milk casein (Sigma company) solution, adjust the pH value to 8.0, and place it in a reactor with constant stirring and a constant temperature of 37°C. 0.2 g of trypsin (Sigma Company) was added according to the mass ratio of enzyme to substrate of 1:100, and the timing was started, while NaOH was added dropwise to maintain the pH of the system at 8.0. Stop the reaction after reacting for 10 minutes, immediately inactivate the enzyme in a boiling water bath for 10 minutes, and the degree of hydrolysis at this time is 6%. The enzymolysis solution is filtered with a 5kDa hollow fiber membrane to obtain a filtrate (product), and then filtered with a 1kDa hollow fiber membrane to obtain a retentate with a molecular weight of 1k-5kDa. Add CaCl to the retentate 2 , to make the concentration 1.0%, let it stand for 60min, then add ethanol to make the ethanol concentration 60%, and form a precipitate, which is centrifuged (10000rpm) to get the precipi...

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Abstract

The invention relates to a multi-phosphopeptide-enriched casein hydrolyzate and a preparation method thereof. The method comprises the following steps of: with the casein as a raw material, suitably hydrolyzing by adopting a proteolytic enzyme, controlling a hydrolysis degree within 10 percent, heating with a boiling water bath for killing enzyme and separating out various polypeptide products from a reaction system by adopting an ultrafiltration membrane of 5 k Da; then, intercepting the product with a hyperfiltration membrane of 1 k Da and selectively precipitating phosphoeptide from the obtained intercepting liquid by adopting calcium-ethanol; dissolving the phosphoeptide with deionized water or a volatile salt solution, sampling on a strong cation exchange column and separating to obtain a plurality of phosphorylated peptides; acquiring a product by adopting spraying drying or freezing drying; and finally, carrying out the sequence analysis on the obtained phosphoeptide by adopting LC / MS (Coupled HPLC-Mass Spectrometry). The invention can selectively acquire the phosphoeptide with higher activity, has favorable application prospects in the field of food biotechnology and provides a favorable research sample for researching phosphoproteomics.

Description

technical field [0001] The invention relates to a casein hydrolyzate rich in polyphosphorylated polypeptides, and a preparation method thereof. Most of these phosphopeptides carry SerP-SerP-SerP-Glu-Glu sequence fragments, and belong to the field of food biotechnology. Background technique [0002] Casein accounts for about 80% of the total protein in mammalian milk and is composed of α S1 - casein, alpha S2 -A micellar macromolecule formed by the aggregation of four phosphorylated proteins such as casein, β-casein, and κ-casein. When newborns are fed breast milk, casein micelles act as calcium phosphate colloidal transporters, providing essential mineral elements for the child's bone formation and tooth growth. During the digestion of breast milk in the body, casein will be hydrolyzed into various polypeptides, but its metal-binding function will be retained or even strengthened by the hydrolyzate-casein phosphopeptides (CPPs). At present, a large number of experimental ...

Claims

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Application Information

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IPC IPC(8): C07K14/47C07K1/36C07K1/34C07K1/30C07K1/18C12P21/06
Inventor 苏荣欣齐崴何志敏
Owner TIANJIN UNIV
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