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Preparation Of α-glucosidase Inhibitory Peptides By Enzymatic Hydrolysis From Chickpea Protein And Its Stability Research

Posted on:2017-03-30Degree:MasterType:Thesis
Country:ChinaCandidate:Y F LiFull Text:PDF
GTID:2180330503489545Subject:Agricultural Extension
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Chickpea protein has a lower sensitivity, high bioavailability, high digestion and absorption rate, etc.As a high-quality plant protein resource,which contains the human body needs a variety of amino acids, proteins, vitamins, etc..It also has reduced high blood pressure, high blood sugar; high blood fat and cholesterol, especially for the ―three high‖ people eat. Current research of chickpea mainly focused on protein modification, physical and chemical properties and initial processing technology, mainly used for cooking food, dry products and other traditional products, lacking of deep-processing technology, resource utilization rate is low. Therefore, it is of great significance to increase the strength of the development of chickpea.In recent years, the pressure of life and irregular diet leads to more and more people suffer from diabetes, China is the largest number of diabetic patients in the world’s countries, prevention and treatment of diabetes has become essential. Alpha-glucosidase inhibitors as novel oral hypoglycemic agents, have been widely used in patients with type Ⅱpostprandial glycemic control, but alpha-glucosidase inhibitor drugs sold on market, although has good curative effect, but it much expensive and has high toxicity. Therefore, the development of high efficiency and low toxicity and low price of natural alpha-glucosidase from animal and plant has become a hot spot in the treatment of diabetes drugs and functional foods.In this paper, the enzymatic hydrolysis process of chickpea protein and the isolation, purification and stability of alpha-glucosidase inhibitory peptides were preliminary inquiry; the main conclusions are as follows:(1) Taking alpha-glucosidase inhibition rate as the main investigation index, the degree of hydrolysis as reference index, using the alkaline protease, neutral protease, papain and flavor protease, four protease to hydrolyze degreasing treatment chickpea protein. The optimal enzymatic hydrolysis enzyme was alkaline protease, which produced the alpha-glucosidase inhibitory peptide has the highest inhibition rate reached 23.09%. Based on single factor experiments, response surface test of alkaline protease enzyme solution of chickpea protein enzyme hydrolysis process was optimized by, the optimum enzyme solution process parameters as follows: hydrolysis time 5.1h, hydrolysis temperature 57℃, substrate concentration 5.2%, pH10.0, enzyme concentration 4000U/g. Under these conditions, enzyme solution prepared alpha-glucosidase inhibitory peptide inhibition rate up to 31.86%.(2) By the ultrafiltration of chickpea protein hydrolysates were crude separated and obtain two components: less than 3kDa and greater than 3kDa, the alpha-glucosidase inhibition rate is 47.53% and 27.87%,respectively shows that less than 3kDa group inhibitory activity is better. Then less than 3kDa group divided by Sephadex G-25 and Sephadex G-10 to further separation and purification, and finally got high inhibitory activity of peak 2 components, the α-glucosidase inhibition rate was 59.01%,compared with the crude hydrolyzate activity it increased by 1.85 times. High resolution mass spectrometry analysis was carried out on the peak 2 component, and the molecular weight of the group was found to be mainly concentrated in about 200~300Da. Analysis amino acid of two components after ultrafiltration, found that less than 3kDa part’s cystine, leucine, phenylalanine and proline were significantly higher than more than 3kDa part, leucine, proline and phenylalanine are hydrophobic amino acids, speculate hypoglycemic peptide activity may be related to the hydrophobic amino acid.(3) Using ultrafiltration obtained less than 3kDa chickpea polypeptides as research object; explore the effects of temperature, pH, food ingredients, antiseptic and metal ions on the inhibition of α-glucosidase activity. The results showed that: Chickpea polypeptides have good heat resistance, heat treatment at 100℃ after 5h activity has declined, but it still kept the inhibitory activity of 72.09%. Chickpea polypeptides with strong acid and alkali resistance, and alkaline environment may contribute to inhibit the ability of the peptide. The addition of glucose and sucrose in food ingredients could not directly show the effect of the chickpea polypeptides, and the addition of lactose, citric acid and NaCl had no significant effect on chickpea polypeptides α-glucosidase inhibitory activity. Sodium benzoate to inhibit enzyme activity of α-glucosidase on chickpea polypeptides is not significant. Different metal ions have different effects on the inhibition ability of α-glucosidase, its influence ability from strong to weak in order of Cu2+>Zn2+>K+>Mg2+. In general, the stability of chickpea polypeptides obtained from the enzymatic hydrolysis was better.
Keywords/Search Tags:α-glucosidase inhibitory peptide, Chickpea protein hydrolysis, separation and purification, stability
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