Evaluation method for degree of thermal denaturation of macromolecular collagen by using pepsin process

Abstract

The invention discloses an evaluation method for the degree of thermal denaturation of macromolecular collagen by using a pepsin process. The method is characterized by comprising the following steps: (1) selection of a raw material: a step of selecting leftovers of aquatic product processing-fishskin as the raw material; (2) immersion and dissolving: a step of adding acetic acid with a concentration of 0.01 to 1.0 mol/L into the raw material to obtain a collagen solution; (3) heat pretreatment: a step of heating the collagen solution, shaking the solution every five minutes during heating and carrying out rapid cooling after heating is finished; (4) enzymatic hydrolysis: a step of adding pepsin into the collagen solution, carrying out a heat reaction and adding phenylmethylsulfonyl fluoride after the heat reaction is finished so as to terminate the reaction; and (5) preparation of an electrophoresis sample and electrophoresis. The invention has the following advantages: used equipment is simple; more comprehensive and visual reflection of the denaturation state of protein is realized.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for evaluating thermal denaturation degree of macromolecular collagen by pepsin method. Background technique [0002] Macromolecular collagen refers to collagen with a complete triple helix structure and a molecular weight of more than 300,000. In the past, the methods used to study the degree of thermal denaturation of collagen mainly include viscosity method, DSC method, circular dichroism method (CD method) and so on. Viscosity method is based on the principle that under heating, collagen molecules uncoil and break, resulting in a decrease in solution viscosity; DSC method is based on the process of triple helix disintegration accompanied by heat absorption; CD method is based on the α-helix existing in collagen , there is a principle of positive or negative absorption at a specific wavelength. However, no matter which method requires corresponding equipment, especially...

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Problems solved by technology

However, no matter which method requires corresponding equipment, especially the CD method and DSC method have higher requirements for equipment, and the measurement results are affected by many factors.

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