Liquid chromatography-mass spectrometry technology-based protein-polysaccharide complex digestibility analyzing method

Active Publication Date: 2019-03-08
DALIAN POLYTECHNIC UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, research in this field has not been fully carried out, and a few studies have used a relatively simple analysis method, mainly using in vitro simulated digestion combined with SDS-PAGE electrophoresis to analyze the effect of added polysaccharides on the digestibility of protein-polysaccharide beverages
The limitations of the research techniques us

Method used

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  • Liquid chromatography-mass spectrometry technology-based protein-polysaccharide complex digestibility analyzing method
  • Liquid chromatography-mass spectrometry technology-based protein-polysaccharide complex digestibility analyzing method
  • Liquid chromatography-mass spectrometry technology-based protein-polysaccharide complex digestibility analyzing method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] The preparation of embodiment 1 protein solution and protein-polysaccharide complex solution:

[0074] The preparation steps of the whey protein isolate solution and the whey protein isolate-Tremella polysaccharide complex solution are as follows: dissolve the whey protein isolate in water, stir at 200rpm for 4 hours at room temperature, wherein the mass ratio of protein to water is 20%, and then stir at 5 Overnight at ℃ to fully hydrate the protein to prepare a whey protein isolate solution for later use; dissolve tremella polysaccharide in water, wherein the mass ratio of polysaccharide to water is 2%, then fully hydrate the polysaccharide at 5°C overnight to prepare a tremella polysaccharide solution, spare.

[0075] Whey protein isolate-Tremella polysaccharide complex solution preparation steps: Tremella polysaccharide solution is added to the previously prepared whey protein isolate solution, the volume ratio is 1:1, the appropriate amount of water is controlled, t...

Embodiment 2

[0076]Example 2 In vitro simulated gastric juice digestion whey protein isolate solution and whey protein isolate-Tremella polysaccharide solution:

[0077] Centrifuge tubes containing 25mL of simulated gastric juice (containing 0.05M HCl and 0.5μg / L pepsin) were placed in a constant temperature shaker, shaken for 5 minutes in a water bath at 37°C and 95r / min, divided into two groups, and added Whey protein isolate solution and 10 mL of whey protein isolate-Tremella polysaccharide complex solution were placed in a water bath at 37°C and shaken at 95 r / min for 120 minutes. After digestion, the pH of the reaction solution was adjusted to 7 with 1M NaOH solution. The two groups of solutions after the digestion treatment are respectively referred to as the whey protein solution group and the whey protein-Tremella polysaccharide complex solution group below.

Embodiment 3

[0078] Example 3 Denaturation of whey protein isolate solution and whey protein isolate-Tremella polysaccharide solution and the second step of enzymatic hydrolysis, the specific steps are as follows:

[0079] The above-mentioned whey protein isolate solution group and whey protein isolate-Tremella polysaccharide solution group degraded by simulated gastric juice were added with protein denaturant (the formula of denaturant was 6M guanidine hydrochloride, 100mM tris(hydroxymethylaminomethane), 10mM tris(2- Carboxyethyl)phosphine and 40mM chloroacetamide), then heated at 95°C for 5 minutes, and cooled on ice. The denatured protein fragments were fractionated by centrifugation using an ultrafiltration tube with a membrane pore size of 10 kDa, and the centrifugation condition was 4000 g for 40 minutes. Ammonium bicarbonate solution was added to the large fragment polypeptide remaining in the ultrafiltration tube for centrifugation washing, and the centrifugation condition was 400...

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Abstract

The invention discloses a liquid chromatography-mass spectrometry technology-based protein-polysaccharide complex digestibility analyzing method. The liquid chromatography-mass spectrometry technology-based protein-polysaccharide complex digestibility analyzing method comprises in vitro digesting protein solution and protein-polysaccharide complex solution in simulative gastric juice; performing denaturation treatment on digested proteins and digested protein-polysaccharide complexes, removing low-molecular-weight peptide fragments through an ultrafiltration pipe, and further completely hydrolyzing large-fragment polypeptides retained inside the ultrafiltration pipe through a second endo protease with clear enzyme-digesting points except pepsase to obtain a number of polypeptide fragments;detecting the composition and abundance of the polypeptide fragments through the liquid chromatography-mass spectrometry technology; according to the abundance of the polypeptides digested by the second endo protease and the pepsase, determining the digestibility of the pepsase; further, according to the high-abundance polypeptides in the two types of peptide fragments in the protein-polysaccharide complex group, deducting the binding sites of proteins and polysaccharides.

Description

technical field [0001] The invention belongs to the technical field of analysis methods, and in particular relates to a method for analyzing the digestibility of protein-polysaccharide complexes based on liquid chromatography-mass spectrometry. Background technique [0002] Protein beverages are a large category of beverage products stipulated in my country's national standard GB / T 10789-2015 Beverage General Rules. Refers to the use of milk or dairy products, or other edible proteins derived from animals, or the fruits, seeds or kernels of plants with a certain protein content as raw materials, with or without the addition of other raw and auxiliary materials and (or) food additives, Liquid beverages made through processing and live fermentation. [0003] Protein drinks have the characteristics of low calories and rich in protein. In recent years, they have become a health drink of public concern and a leading role in the emerging beverage market. Taking whey protein drin...

Claims

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Application Information

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IPC IPC(8): G01N30/88
CPCG01N30/88
Inventor 纪超凡赵铁峰胡静税东宁李胜杰祁立波林心萍梁会朋董秀萍
Owner DALIAN POLYTECHNIC UNIVERSITY
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