Table of Content

01 August 2022, Volume 45 Issue 4

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Basic Research

Computer Simulation of Molecular Docking of α-Lactalbumin with Saponins

HU Jialun, JIANG Zhanmei

2022, 45(4):  1-6.  DOI: 10.7506/rykxyjs1671-5187-20220505-025

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Small molecule saponins are highly physiologically active and exert their biological effects only when interacting with biomolecules in the target cells in the human body. α-Lactalbumin (α-LA) is a globular protein that binds to hydrophobic ligands and has been used as a functional ingredient in the food industry. In this study, the interaction between two types of saponins (dammarane-type and oleanane-type) and α-LA was investigated using computer simulations, and their interaction patterns were imaged. The molecular docking results showed that each type of saponins binds to amino acid residues within the active pocket of α-LA via hydrophobic interactions and hydrogen bonding. However, dammarane-type saponins are more complex in structure, providing more binding sites, and they form hydrogen bonds via greater force.

Effects of Tea Polyphenols on Conformation and Antigenicity of Bovine Milk αs1-Casein Allergen

GUO Honglei, ZHOU Shengyun, CONG Yanjun, YAN Wenjie

2022, 45(4):  7-13.  DOI: 10.7506/rykxyjs1671-5187-20220508-026

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In order to investigate the effects of tea polyphenols on the conformation and antigenicity of αs1-casein, epigallocatechin gallate (EGCG) and epigallocatechin (EGC), tea polyphenols with high bioactivity, were used for grafting onto αs1-casein. The conformational changes of αs1-casein were studied by fluorescence, synchronous fluorescence and circular dichroism (CD) spectroscopy, and the changes in the antigenicity of αs1-casein were analyzed by indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and Western blot assay. The results showed that EGC and EGCG both quenched the fluorescence of αs1-casein, and affected the Tyr and Trp residues of αs1-casein. The α-helix and β-sheet contents of αs1-casein increased slightly, and the random coil content decreased slightly. The conformational change of αs1-casein resulted in a significant decrease in the antigenicity of αs1-casein. The effect of EGCG on the conformation and antigenicity of αs1-casein was stronger than that of EGC.

Effects of Enzymatic Hydrolysis on Protein Antigenicity and Sensory Properties of Skim Milk

LI Min, LIU Aicheng, ZHU Qing, CHEN Xinping, LIU Wei, LIANG Xiaona, ZHENG Yan, YUE Xiqing

2022, 45(4):  14-21.  DOI: 10.7506/rykxyjs1671-5187-20220614-037

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The effects of Alcalase (AT), Protamex (PT) or Flavorzyme (FT) treatment on the antigenicity and molecular mass distribution of proteins, flavor and color of skim milk were investigated by using various techniques such as indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and Tricine-sodium dodecyl sulphate polyacrylamide gel electrophoresis (Tricine-SDS-PAGE). The results showed that the effect of AT treatment on reducing the antigenicity of the major milk allergens was significantly better than that of PT and FT treatment (P < 0.05). The antigenicity of α-lactalbumin (α-LA), β-lactoglobulin (β-LG) and casein (CN) was inhibited by 64.01%, 76.00% and 69.10% by AT treatment for 20 min at an enzyme/substrate ratio of 500 U/g, respectively. The amount of low molecular mass peptides in skim milk increased significantly after treatment with each of the three enzymes. The bitterness, astringency, aftertaste of bitterness (aftertaste-B) and aftertaste of astringency (aftertaste-A) increased with increasing enzymatic treatment time and with increasing enzyme/substrate ratio. The taste of FT-treated milk was better than that of AT- or PT-treated milk. The brightness value of milk decreased significantly, while the redness value increased significantly after enzymatic treatment (P < 0.05). The transmittance increased as well. The color of skim milk treated by AT was more similar to that of whole milk.

Protease Screening for the Elimination of the Allergenicity of Casein

TAN Hongkai, CHENG Jianfeng, XIONG Ziyi, HU Wei, LI Xin, CHEN Hongbing

2022, 45(4):  22-28.  DOI: 10.7506/rykxyjs1671-5187-20220330-017

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In this study, casein was hydrolyzed with seven food-grade proteases, including flavourzyme, trypsin, chymotrypsin, alkaline protease, papain, bromelain, and neutral protease and the allergenicity of the resultant hydrolysates was assessed using a serum pool of patients with milk allergy. Moreover, the molecular mass distribution, degree of hydrolysis and residual epitope information were evaluated in order to select the most suitable protease for the production of casein products with reduced allergenicity. The results showed that under the same conditions, the binding ability of casein to specific antibodies decreased to the greatest extent after being treated with chymotrypsin or flavourzyme, so they could be used as a candidate protease for the preparation of subsequent hypoallergenic casein hydrolysates. The results of T-cell epitope prediction and analysis of casein hydrolysates showed that only one T-cell epitope peptide consisting of more than nine amino acids LHSMKEGIHAQQK remained after chymotrypsin hydrolysis of casein.

Effect of Thermal Processing on the Secondary Structure and Antigenicity of Bovine Milk αs1-Casein

WEI Jiaqi, ZHOU Shengyun, CONG Yanjun, YAN Wenjie

2022, 45(4):  29-35.  DOI: 10.7506/rykxyjs1671-5187-20220508-028

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To investigate the effect of thermal processing on the conformation and antigenicity of bovine milk αs1-casein, the changes in the immunogenicity of αs1-casein under different heating conditions were analyzed by indirect competitive enzyme immunosorbent assay (icELISA) and Western blotting (WB) methods, and then the secondary structure changes were analyzed by fluorescence spectroscopy using 8-anilino-1-naphthalenesulfonic acid (ANS) as a probe and circular dichroism spectroscopy to preliminarily reveal the regulatory mechanism of heat treatment on the antigenicity of αs1-casein. The results showed that the α-helix content of αs1-casein significantly decreased after treatment at 80 ℃ for 60 min, 90 ℃ for 10 min or 90 ℃ for 60 min compared with the untreated control, whereas the random coil content increased significantly after treatment at 70 ℃ for 20 min, 80 ℃ for 20 min or 90 ℃ for 20 min. The surface flourescence intensity was the strongest after heating treatment at 70-100 ℃ for 20 min, while the secondary structure changes were not significant under other heating conditions. The conformational changes of αs1-casein led to a significant decrease in the antigenicity of αs1-casein. The icELISA showed that the residual antigenicity of αs1-casein was the highest after heating treatment at 70-100 ℃ for 20 min, while the WB results showed that αs1-casein still had immunoreactive properties under all heating conditions. It is suggested to further reveal the regulatory mechanism of heat treatment on the antigenicity of αs1-casein through animal tests.

Effects of Dual-Enzyme Hydrolysis on Allergenicity of Skimmed Milk

QIAN Guanlin, SUN Jing, LIU Wei, CHENG Jiao, YUE Xiqing, ZHENG Yan

2022, 45(4):  36-44.  DOI: 10.7506/rykxyjs1671-5187-20220614-036

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In this study, the effect of dual-enzyme hydrolysis on the allergenicity of the major proteins in skimmed milk was evaluated based on the degree of hydrolysis, molecular mass distribution, immunoglobulin G (IgG) binding capacity and allergic patients’ serum IgE binding capacity of hydrolysates. The results showed that after one-step hydrolysis for 80 min at an enzyme/substrate ratio of 3 000 U/g and an Alcalase to Flavourzyme ratio of 3:1, the IgG binding capacity of casein, α-lactalbumin and β-lactoglobulinin in skimmed milk decreased significantly by 74.93%, 97.24% and 93.46%, respectively (P < 0.05).The advanced structure of proteins was damaged, leading to an increase in the α-helical and β-turn contents, and a decrease in the β-sheet content. In addition, the free sulfhydryl content increased significantly to 14.29 μmol/g (P < 0.05), the surface hydrophobicity also increased, and the structure became loose. The particles in skimmed milk were aggregated, causing the average particle size to increase significantly to 346.90 nm (P < 0.05) and reducing the stability of skimmed milk. The binding capacity to serum IgE from patients with milk allergy of the enzymatic hydrolysate decreased significantly by 23.53% (P < 0.05), and the overall allergenicity decreased significantly by 69.60% (P < 0.05). Therefore, dual-enzyme hydrolysis is an effective method to reduce the allergenicity of skimmed milk.

Evaluation of Gastrointestinal Stability and Antigenicity of Whey Protein in Commercial Infant Milk Powder

ZHOU Qi, YANG Fan, FU Siqi, QIU Yu, LI Xin, CHEN Hongbing

2022, 45(4):  45-51.  DOI: 10.7506/rykxyjs1671-5187-20220614-034

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Cow’s milk is rich in protein, fat, and other high-quality nutrients, which is an important nutritional source for infants. On the other hand, milk protein is the main factor inducing food allergy in infants and young children. In this study, five kinds of commercially available infant milk powder were selected for static digestion in vitro. The digestive stability of proteins in the five kinds of milk powder was analyzed by electrophoresis and degree of hydrolysis measurement. An enzyme-linked immunosorbent assay (ELISA) was used to evaluate the antigenicity of whey protein in milk powder. The results showed that α-lactalbumin and β-lactoglobulin showed good digestive stability in samples 1, 2, and 4. After gastrointestinal digestion, the antigenicity of α-lactalbumin and β-lactoglobulin decreased in sample 1 but increased in sample 2. The antigenicity of α-lactalbumin increased, while that of β-lactoglobulin decreased in sample 4. Both allergens had lower digestion stability in sample 3 and 5. The antigenicity of α-lactalbumin and β-lactoglobulin increased in sample 5. The antigenicity of α-lactalbumin increased while that of β-lactoglobulin decreased in sample 3.

Analysis & Detection

Determination of Protein in Commercial Goat Milk Powder by Reversed Phase High Performance Liquid Chromatography

SONG Zhuoyan, QIAO Chunyan, ZHANG Xueru, HAO Guo, LIU Yongfeng

2022, 45(4):  52-56.  DOI: 10.7506/rykxyjs1671-5187-20220614-035

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To identify and test the quality of commercial goat milk powder sold in different areas, reversed-phase high performance liquid chromatography (RP-HPLC) was used to determine the protein composition of 35 samples of pure and formulated goat milk powder purchased in different regions (Shaanxi and non-Shaanxi) of China and in other countries, and the difference in protein composition among goat milk powder from different production areas and that between pure and formulated goat milk powder were analyzed. The results showed that the contents of αs2-casein and β-casein in formulated milk powder were 20% and 24% lower than those in pure milk powder, respectively, and the contents of κ-casein and whey proteins were 30% and 50% higher than those in pure milk powder, respectively. However, the protein composition and content of pure goat milk powder from different regions were similar. Therefore, RP-HPLC is useful for protein quality evaluation of commercial goat milk powder.

Reviews

Effect of Different Nutrients on Allergenicity of Milk Protein: A Review

LIU Yanchen, WANG Qi, XU Yunpeng, MU Guangqing, QIAN Fang, ZHU Xuemei

2022, 45(4):  57-62.  DOI: 10.7506/rykxyjs1671-5187-20220508-027

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Milk protein is an important source of dietary protein for humans. In the modern food industry, milk and milk protein have become essential components in various processed foods. Milk protein is one of the most common allergens, and the problem of food allergies caused by milk protein has become a topical issue in the field of food safety. With the development of the dairy industry, non-milk derived food ingredients have been introduced in various dairy products, which may interact with milk protein, altering the protein’s structure and affecting its antigenic epitopes and consequently its allergenicity. Herein, we discuss the effects of food components on milk protein allergenicity from macronutrients, micronutrients and functional activities contained in the food base, and other substances. We hope that this review can improve the scientific understanding of milk protein allergenicity in food components, guide food processing safety to reduce milk protein allergenicity, and provide a basis for food allergenicity risk assessment.

Progress in Research on Iron Supplements and Polypeptide-Iron Complexes

ZHONG Liwen, LU Yingrui, ZHANG Weibing, GUO Zhaobin, ZHANG Jinliang, WEN Pengcheng

2022, 45(4):  63-69.  DOI: 10.7506/rykxyjs1671-5187-20220605-033

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As the most common type of anemia, iron-deficiency anemia has attracted more and more attention. With the change of nutrition and health needs and the enhancement of safety awareness, there has been increasing interest in the research and development of oral iron supplements with good iron supplementation effect, easy absorption, small side effects and high safety. This paper reviews recent progress in the development and research of iron supplements. In this paper, the raw material source, structure, action mechanism, preparation, application, and digestion and absorption mechanism of iron protein succinate are reviewed, and future prospects of polypeptide-iron complexes and iron protein succinate are discussed. We anticipate that this review will provide a theoretical basis for the research and development of iron supplements.