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Antifreeze polypeptide prepared through enzymolysis of pigskin collagen protein by utilizing alkali protease

A technology of antifreeze peptides and amino acids, applied in the preparation method of peptides, peptides, organic chemistry, etc., can solve the problems of small quantities and limit the application prospects of large laws

Active Publication Date: 2012-07-25
FUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Repeated rise and fall of temperature causes ice crystals to grow continuously, freeze-thaw and recrystallize, which seriously damages the cell and tissue structure and loses the quality of the product.

Method used

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  • Antifreeze polypeptide prepared through enzymolysis of pigskin collagen protein by utilizing alkali protease
  • Antifreeze polypeptide prepared through enzymolysis of pigskin collagen protein by utilizing alkali protease
  • Antifreeze polypeptide prepared through enzymolysis of pigskin collagen protein by utilizing alkali protease

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preparation example Construction

[0031] The antifreeze activity detection system for preparing antifreeze polypeptide of the present invention adopts a combined cold-heat-stage system, multi-fold microscope and automatic camera system to construct an antifreeze activity inhibition ice crystal growth detection system platform. The cryogenic environment during operation was maintained with liquid nitrogen. Simulate temperature fluctuations of products during cryogenic storage. Through the system program setting, set the temperature fluctuation of the sample and repeat it continuously for 7 to 25 cycles, connect a multi-fold microscope and a real-time photographic recording system, and record the real-time ice crystal growth of the sample during the cold-hot cycle.

[0032] Sephadex G-50 molecular sieves, Sepharose SP C-25 ion exchange chromatography, RP-HPLC reversed-phase high-performance liquid chromatography, dialysis and other separation and purification methods are used to achieve efficient separation and ...

Embodiment 1

[0036] Weigh 1.65 g of pigskin collagen and dissolve it in 6 ml of Mili-Q water, then adjust its pH to 9.0 with 2 mol / L NaOH. First heat the solution to 45°C in a water bath, then add the corresponding amount of enzyme according to the enzyme-substrate ratio of 1:15, and the enzymatic hydrolysis time is 30 minutes. Then inactivate the enzyme in a boiling water bath for 10 minutes, then centrifuge at 14000 rpm for 10 minutes after cooling, and collect the supernatant for later use.

[0037] The supernatant was separated by Sephadex G-50 gel chromatography (length 100cm, diameter 2.6cm), the eluent was deionized water, the flow rate was 2mL / min, and the elution peak was measured at 225nm, see figure 1 . Antifreeze activity tests showed that Fraction2 had the best antifreeze activity, and statistical analysis showed that the average diameter of ice crystals was 5.67 μm, which was significantly smaller than Fraction1 (18.83 μm) and Fraction3 (9.02 μm), with a significant diff...

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Abstract

The invention provides a method for preparing antifreeze polypeptide through the enzymolysis of pigskin collagen protein by utilizing alkali protease. The pigskin collagen protein is adopted as a raw material and separated and purified after the enzymolysis of alkali protease to obtain purified special antifreeze polypeptide, and the complete sequence of amino acids is Gly-His-Arg-Gly-Phe-Ser-Gly-Leu-Asp-Gly-Ala-Lys. The invention breaks through traditional antifreeze protein (polypeptide) research ideas and methods existing at home and abroad, overcomes the quantity limitation of antifreeze protein purified from natural organisms and the safety worry of the international FDA (Food and Drug Administration) organization on the applications of the transgenic antifreeze protein to foods, obtains the high-efficiency antifreeze polypeptide based on food sources and lays a theoretical basis for developing the antifreeze polypeptide based on food sources and discovering the wide applicationsthereof to foods and medical science.

Description

technical field [0001] The invention relates to an antifreeze polypeptide, in particular to an antifreeze polypeptide prepared by hydrolyzing pigskin collagen with alkaline protease, and belongs to the field of biotechnology. Background technique [0002] The problem that food and pharmaceutical products repeatedly suffer from ice crystal growth and recrystallization due to fluctuations in ambient temperature during cryogenic storage and transportation has attracted increasing attention. The repeated rise and fall of temperature causes ice crystals to grow continuously, freeze-thaw and recrystallize, which seriously damages the cell and tissue structure and loses the quality of the product. Scientists around the world are facing serious challenges: how to control the growth and recrystallization of ice crystals, and realize the control of ice crystal growth on the low-temperature cold chain is the key to restricting the quality of many food and pharmaceutical products. [0...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/08C07K1/20C07K1/18C07K1/16C12P21/06
Inventor 汪少芸洪晶饶平凡
Owner FUZHOU UNIV
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