基于红外光谱分析巴氏杀菌乳贮藏期间蛋白质二级结构的变化
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摘要
巴氏杀菌乳因最大限度地保留了乳的营养物质和风味,深受国内外消费者的普遍青睐,然而,因其杀菌温度低、货架期很短,大大限制了大规模生产、远距离销售。其中,在贮运期间发生的蛋白质腐败性变质,是导致巴氏杀菌乳在贮运期间品质劣变的主要原因之一。在前期研究的基础上,结合蛋白质含量与氨基酸含量的分析,利用傅里叶变换红外光谱技术(FT-IR)分别对采集的不同贮藏温度和时间条件下的巴氏杀菌乳样品的蛋白质二级结构进行测定,并对不同巴氏杀菌乳样品的蛋白及其酰胺Ⅰ带及酰胺Ⅲ带进行了拟合,对不同贮藏温度和时间条件下的巴氏杀菌乳的蛋白质二级结构的变化规律进行进一步研究。结果表明:贮藏温度和时间对巴氏杀菌乳中蛋白质二级结构具有显著的影响,其中,在0℃和4℃贮藏条件下,巴氏杀菌乳乳清蛋白中的α-螺旋结构所占比例显著降低,无规卷曲结构所占比例显著上升,β-折叠、β-转角结构所占比例趋于稳定。而在15℃和25℃贮藏条件下α-螺旋结构的比例先下降后上升,总体来看,乳清蛋白二级结构随着贮藏时间的增加,由有序向无序转变。该结果表明了在不同贮藏条件下巴氏杀菌乳中蛋白质二级结构由有序向无序变化,二级结构的变化降低了巴氏杀菌乳在贮藏期间的营养价值。从蛋白质二级结构的角度来看,4℃的贮藏条件能更好保持巴氏杀菌乳的营养品质。
Pasteurized milk is widely favored by consumers at home and abroad because it retains the nutrients and flavor of milk to the utmost extent. However, due to its low sterilization temperature and short shelf life, it has greatly restricted largescale production and long-distance sales. Among them, the deterioration of protein spoilage during storage and transportation is one of the main reasons leading to the deterioration of quality of pasteurized milk during storage and transportation. Based on the previous research, combined with the analysis of protein content and amino acid content, Fourier transform infrared spectroscopy(FT-IR) was used to separate the proteins of pasteurized milk samples under different storage temperatures and time conditions. Meanwhile the proteins of different pasteurized milk samples were fitted with their amide I and amide III bands.Under different storage temperatures and time conditions, the changes of protein secondary structure of pasteurized milk were studied. The results showed that storage temperature and time had a significant effect on the secondary structure of protein in pasteurized milk. Among them, under the storage conditions of 0 ° C and 4 ° C, the proportion of α-helical structure in pasteurized milk whey protein decreased significantly, and the proportion of random coil structure increased significantly. β-sheet and β-turn structure The proportion tended to be stable. Under the storage conditions of 15 °C and 25 °C, the proportions ofα-helical structure decreased first and then increased. Overall, the secondary structure of whey protein changed from ordered to disorder with the increase of storage time. The results indicated that the secondary structure of protein in pasteurized milkchanged from ordered to disorder under different storage conditions. Changes in secondary structure reduced the nutritional value of pasteurized milk during storage. From the perspective of protein secondary structure, storage conditions at 4 ° C can better maintain the nutritional quality of pasteurized milk.
Pasteurized milk is widely favored by consumers at home and abroad because it retains the nutrients and flavor of milk to the utmost extent. However, due to its low sterilization temperature and short shelf life, it has greatly restricted largescale production and long-distance sales. Among them, the deterioration of protein spoilage during storage and transportation is one of the main reasons leading to the deterioration of quality of pasteurized milk during storage and transportation. Based on the previous research, combined with the analysis of protein content and amino acid content, Fourier transform infrared spectroscopy(FT-IR) was used to separate the proteins of pasteurized milk samples under different storage temperatures and time conditions. Meanwhile the proteins of different pasteurized milk samples were fitted with their amide I and amide III bands.Under different storage temperatures and time conditions, the changes of protein secondary structure of pasteurized milk were studied. The results showed that storage temperature and time had a significant effect on the secondary structure of protein in pasteurized milk. Among them, under the storage conditions of 0 ° C and 4 ° C, the proportion of α-helical structure in pasteurized milk whey protein decreased significantly, and the proportion of random coil structure increased significantly. β-sheet and β-turn structure The proportion tended to be stable. Under the storage conditions of 15 °C and 25 °C, the proportions ofα-helical structure decreased first and then increased. Overall, the secondary structure of whey protein changed from ordered to disorder with the increase of storage time. The results indicated that the secondary structure of protein in pasteurized milkchanged from ordered to disorder under different storage conditions. Changes in secondary structure reduced the nutritional value of pasteurized milk during storage. From the perspective of protein secondary structure, storage conditions at 4 ° C can better maintain the nutritional quality of pasteurized milk.
引文
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[22] KHER A,UDABAGE P,MCKINNON I,et al.FTIR investigation of spray-dried milk protein concentrate powders[J].Vibrational Spectroscopy,2007,44(2):375-381.
[23] MORAND M,GUYOMARC'H F,PEZENNEC S,et al.On howκ-casein affects the interactions between the heat-induced whey protein/κ-casein complexes and the casein micelles during the acid gelation of skim milk[J].International Dairy Journal,2011,21(9):670-678.
[24]石燕,刘凡,葛辉,等.微胶囊形成过程中蛋白质二级结构变化的红外光谱分析[J].光谱学与光谱分析,2012,32(7):1815-1819.
[25]阎微.高压和热处理对蛋黄体系中蛋白质的影响[D].无锡:江南大学,2009.
[26]顾小红,孟旭,汤坚.豆浆凝固过程中大豆蛋白质二级结构的研究[J].分析科学学报,2006,22(6):675-678.
[27]李顺,张聚华,李秀艳,等.FT-IR自去卷积法对丝织物蛋白酰胺Ⅲ带二级结构研究初探[J].北京服装学院学报(自然科学版),2011,31(2):68-73.
[28] LONG G H,JI Y,PAN H B,et al.Characterization of thermal denaturation structure and morphology of soy glycinin by FTIR and SEM[J].International Journal of Food Properties,2015,18(4):763-774.
[29] GUYOMARC’H F,NONO M,NICOLAI T,et al.Heat-induced aggregation of whey proteins in the presence ofκ-casein or sodium caseinate[J].Food Hydrocolloids,2009,23(4):1103-1110.
[30] WIJAYANTI H B,BANSAL N,DEETH H C.Stability of whey proteins during thermal processing:a review[J].Comprehensive Reviews in Food Science and Food Safety,2014,13(6):1235-1251.
[31]张波波,王丹,马越,等.FT-IR分析超高压处理对调配草莓汁蛋白二级结构的影响[J].中国食品学报,2014,14(11):200-206.
[32] DANIEL J P,MARK R P.Alpha-helix mimetics:progress toward effective modulation of protein-protein complexes[J].Frontiers in Drug Design&Discovery,2012,3:5-44.
[33] YANG J,MA Y Q,PAGE R C,et al.Structure of an integrin alphaIIb beta3 transmembrane-cytoplasmic heterocomplex provides insight into integrin activation[J].Proceedings of the National Academy of Sciences of the United States of America,2009,106(42):17729-17734.
[34] ZHU J,LUO B H,BARTH P,et al.The structure of a receptor with two associating transmembrane domains on the cell surface:integrin alphaIIb beta3[J].Mol Cell,2009,34:234-249.
[35]王庆文.有机化学中的氢键问题[M].天津:天津大学出版社,1993:236-258.
[36] PORCELLATO D,ASPHOLM M,SKEIE S B,et al.Microbial diversity of consumption milk during processing and storage[J].International Journal of Food Microbiology,2018,266:21-30.
[37]宋慧敏.热处理对牛乳风味及保藏品质的影响[D].哈尔滨:东北农业大学,2015.
[38] FITOUHI N,GMAR M,SBOUI N,et al.Correlations between quality,enzymatic activities and storage stability of UHT milk in Tunisia[J].Nature&Technology,2018(18):36-41.
[39]张熙桐,关博元,孔繁华,等.人乳、牛乳、羊乳中乳清部分氨基酸组成及乳清蛋白中蛋白质二级结构的对比[J].食品科学,2017,38(24):107-112.
[40] KIRSCHNER A,FRISHMAN D.Prediction ofβ-turns andβ-turn types by a novel bidirectional Elman-type recurrent neural network with multiple output layers[J].Gene,2008,422:22-29.
[2] MOTTA J,NAUDTS M.巴氏杀菌乳与超高温灭菌乳及瓶(罐)装灭菌乳的品质比较研究[J].中国乳业,2005(7):50-54.
[3]巫庆华,龚广予.巴氏杀菌牛乳和UHT牛乳的差别[J].乳业科学与技术,2003,26(4):149-153.
[4]卢蓉蓉,刘莹,张国农.屋顶奶贮藏期品质变化的研究[J].中国乳品工业,2006,34(10):29-33.
[5]杨怀谷,郑楠,王加启.巴氏杀菌乳和超高温灭菌乳营养价值及卫生安全对比研究[J].中国乳业,2016(7):62-67.
[6]孙雪姣.不同贮藏温度对巴氏杀菌乳品质和微生物的影响[D].沈阳:沈阳农业大学,2018.
[7] PORCELLATO D,ASPHOLM M,SKEIE S B,et al.Microbial diversity of consumption milk during processing and storage[J].International Journal of Food Microbiology,2018,266:21-30.
[8] MARKOSKA T,HUPPERTZ T,GREWAL M K,et al.Structural changes of milk proteins during heating of concentrated skim milk determined using FTIR[J].International Dairy Journal,2019,89:21-30.
[9] JAISWAL P,JHA S N,BORAH A,et al.Detection and quantification of soymilk in cow-buffalo milk using Attenuated Total Reflectance Fourier Transform Infrared spectroscopy(ATR-FTIR)[J].Food Chemistry,2015,168:41-47.
[10]谢孟峡,刘媛.红外光谱酰胺Ⅲ带用于蛋白质二级结构的测定研究[J].高等学校化学学报,2003(2):226-231.
[11] GREWAL M K,CHANDRAPALA J,DONKOR O,et al.Fourier transform infrared spectroscopy analysis of physicochemical changes in UHT milk during accelerated storage[J].International Dairy Journal,2017,66:99-107.
[12] YANG M,CAO X Y,WU R N,et al.Comparative proteomic exploration of whey proteins in human and bovine colostrum and mature milk using iTRAQ-coupled LC-MS/MS[J].International Journal of Food Sciences and Nutrition,2017,20:1-11.
[13]冯志强,周芳梅,黄永连,等.全自动氨基酸分析仪鉴别不同种类酱油中氨基酸的分析研究[J].中国食品添加剂,2013(5):198-205.
[14] YAN P M,WANG Q,YAN X Y,et al.Evaluation of amino acid and vitamin B 2 and E composition of genetically modified antidwarf mosaic maize by automatic amino acid analyzer and HPLC[J].Asian Journal of Chemistry,2013,25(6):3525-3526.
[15]张元福,徐怡庄,翁诗甫,等.傅立叶变换中红外光纤光谱技术及其应用[J].现代仪器,2009(5):27-29.
[16]陈庆华,王欣.冷藏温度及时间对巴氏杀菌乳品质的影响研究[J].食品科技,2009(1):84-87.
[17]陈平生.贮存温度对玻璃瓶装巴氏杀菌乳品质的影响研究[J].安徽农业科学,2014,18(34):12287-12289.
[18]吴达雄,李冬梅,李颂群,等.巴氏杀菌乳门店冰箱冷藏贮存售卖期间的温度及品质变化[J].现代食品科技,2017(12):99-103.
[19] KAUR K S,RAGHAVA G P.A neural network method for prediction of beta-turn types in proteins using evolutionary information[J].Bioinformatics,2004,16:2751-2758.
[20] KIRSCHNER A,FRISHMAN D.Prediction ofβ-turns andβ-turn types by a novel bidirectional Elman-type recurrent neural network with multiple output layers[J].Gene,2008,422:22-29.
[21]王树青,刘红,杜奇石,等.依据氨基酸残基的相关性预测蛋白质的结构类型[J].物理化学学报,2004,20(5):498-502.
[22] KHER A,UDABAGE P,MCKINNON I,et al.FTIR investigation of spray-dried milk protein concentrate powders[J].Vibrational Spectroscopy,2007,44(2):375-381.
[23] MORAND M,GUYOMARC'H F,PEZENNEC S,et al.On howκ-casein affects the interactions between the heat-induced whey protein/κ-casein complexes and the casein micelles during the acid gelation of skim milk[J].International Dairy Journal,2011,21(9):670-678.
[24]石燕,刘凡,葛辉,等.微胶囊形成过程中蛋白质二级结构变化的红外光谱分析[J].光谱学与光谱分析,2012,32(7):1815-1819.
[25]阎微.高压和热处理对蛋黄体系中蛋白质的影响[D].无锡:江南大学,2009.
[26]顾小红,孟旭,汤坚.豆浆凝固过程中大豆蛋白质二级结构的研究[J].分析科学学报,2006,22(6):675-678.
[27]李顺,张聚华,李秀艳,等.FT-IR自去卷积法对丝织物蛋白酰胺Ⅲ带二级结构研究初探[J].北京服装学院学报(自然科学版),2011,31(2):68-73.
[28] LONG G H,JI Y,PAN H B,et al.Characterization of thermal denaturation structure and morphology of soy glycinin by FTIR and SEM[J].International Journal of Food Properties,2015,18(4):763-774.
[29] GUYOMARC’H F,NONO M,NICOLAI T,et al.Heat-induced aggregation of whey proteins in the presence ofκ-casein or sodium caseinate[J].Food Hydrocolloids,2009,23(4):1103-1110.
[30] WIJAYANTI H B,BANSAL N,DEETH H C.Stability of whey proteins during thermal processing:a review[J].Comprehensive Reviews in Food Science and Food Safety,2014,13(6):1235-1251.
[31]张波波,王丹,马越,等.FT-IR分析超高压处理对调配草莓汁蛋白二级结构的影响[J].中国食品学报,2014,14(11):200-206.
[32] DANIEL J P,MARK R P.Alpha-helix mimetics:progress toward effective modulation of protein-protein complexes[J].Frontiers in Drug Design&Discovery,2012,3:5-44.
[33] YANG J,MA Y Q,PAGE R C,et al.Structure of an integrin alphaIIb beta3 transmembrane-cytoplasmic heterocomplex provides insight into integrin activation[J].Proceedings of the National Academy of Sciences of the United States of America,2009,106(42):17729-17734.
[34] ZHU J,LUO B H,BARTH P,et al.The structure of a receptor with two associating transmembrane domains on the cell surface:integrin alphaIIb beta3[J].Mol Cell,2009,34:234-249.
[35]王庆文.有机化学中的氢键问题[M].天津:天津大学出版社,1993:236-258.
[36] PORCELLATO D,ASPHOLM M,SKEIE S B,et al.Microbial diversity of consumption milk during processing and storage[J].International Journal of Food Microbiology,2018,266:21-30.
[37]宋慧敏.热处理对牛乳风味及保藏品质的影响[D].哈尔滨:东北农业大学,2015.
[38] FITOUHI N,GMAR M,SBOUI N,et al.Correlations between quality,enzymatic activities and storage stability of UHT milk in Tunisia[J].Nature&Technology,2018(18):36-41.
[39]张熙桐,关博元,孔繁华,等.人乳、牛乳、羊乳中乳清部分氨基酸组成及乳清蛋白中蛋白质二级结构的对比[J].食品科学,2017,38(24):107-112.
[40] KIRSCHNER A,FRISHMAN D.Prediction ofβ-turns andβ-turn types by a novel bidirectional Elman-type recurrent neural network with multiple output layers[J].Gene,2008,422:22-29.