亚硫酸钠辅助酶解蛋壳膜制备抗氧化多肽的工艺
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摘要
禽蛋壳膜蛋白含量丰富,蛋白质约占蛋壳膜干重的90%,然而蛋壳膜在食品工业中通常被作为废弃物丢弃,造成了资源浪费和环境污染,因此高效地利用蛋壳膜资源是极为必要的。本研究使用亚硫酸钠辅助酶解法制备禽蛋壳膜抗氧化肽,研究了酶种类、酶添加量、亚硫酸钠浓度和酶解时间对禽蛋壳膜多肽抗氧化性和水解度的影响。结果表明,亚硫酸钠辅助下碱性蛋白酶酶解禽蛋壳膜,效果优于其他四种酶;亚硫酸钠辅助碱性蛋白酶酶解禽蛋壳膜的最佳条件是:碱性蛋白酶添加量12 U/mg,亚硫酸钠浓度40 mM,时间4 h。在最优工艺条件下,禽蛋壳膜多肽的总抗氧化能力为245.88μg/m L,水解度为8.58%,表明禽蛋壳膜的亚硫酸钠辅助碱性蛋白酶酶解产物具有很好的体外抗氧化活性。SDS-PAGE表明禽蛋壳膜多肽的分子量主要在3.3~14.4 ku之间。
The eggshell membrane has abundant protein, which acounts for about 90% of the dry weight of the eggshell membrane. However, eggshell membranes are often discarded as waste in food industry, resulting in waste of resources and environmental pollution, thus, the efficient use of eggshell membranes is extremely necessary. In this study, the antioxidant peptides were prepared by sodium sulfite-assisted enzymatic hydrolysis. Effect of protease species, enzyme dosage, sodium sulfite concentration and hydrolysis time on antioxidant activity and hydrolysis degree of eggshell membrane peptides was researched. The results showed that alkaline protease assisted by sodium sulfite was better than the other four proteases. At 12 U/mg alkaline protease, sodium sulfite concentration 40 mM, hydrolysis time 4 h, the degree of hydrolysis and total antioxidant activity of the hydrolysates were 8.58% and 245.88 μg/m L, respectively, indicating that the hydrolysates have good antioxidant activity in vitro. The results of SDS-PAGE and Tricine-SDS-PAGE showed that the molecular weight of the eggshell membrane polypeptide was mainly in the range of 3.3~14.4 ku.
The eggshell membrane has abundant protein, which acounts for about 90% of the dry weight of the eggshell membrane. However, eggshell membranes are often discarded as waste in food industry, resulting in waste of resources and environmental pollution, thus, the efficient use of eggshell membranes is extremely necessary. In this study, the antioxidant peptides were prepared by sodium sulfite-assisted enzymatic hydrolysis. Effect of protease species, enzyme dosage, sodium sulfite concentration and hydrolysis time on antioxidant activity and hydrolysis degree of eggshell membrane peptides was researched. The results showed that alkaline protease assisted by sodium sulfite was better than the other four proteases. At 12 U/mg alkaline protease, sodium sulfite concentration 40 mM, hydrolysis time 4 h, the degree of hydrolysis and total antioxidant activity of the hydrolysates were 8.58% and 245.88 μg/m L, respectively, indicating that the hydrolysates have good antioxidant activity in vitro. The results of SDS-PAGE and Tricine-SDS-PAGE showed that the molecular weight of the eggshell membrane polypeptide was mainly in the range of 3.3~14.4 ku.
引文
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[27]Nasri R, Jridi M, Lassoued I, et al. The influence of the extent ofenzymatichydrolysisonantioxidativepropertiesand ace-inhibitoryactivitiesofproteinhydrolysatesfromgoby(Zosterisessorophiocephalus)muscle[J].ApplBiochem Biotechnol, 2014, 173(5):1121-1134
[2]QianZ-J,JungW-K,KimS-K.Freeradicalscavenging activityofanovelantioxidativepeptidepurifiedfrom hydrolysateofbullfrogskin,ranacatesbeianashaw[J].Bioresource Technology, 2008, 99(6):1690-1698
[3]Moure A, Domínguez H, ParajóJ C. Antioxidant properties ofultrafiltration-recoveredsoyproteinfractionsfrom industrialeffluentsandtheirhydrolysates[J].Process Biochemistry, 2006, 41(2):447-456
[4]Bougatef A, Nedjar-Arroume N, Manni L, et al. Purification andidentificationofnovelantioxidantpeptidesfrom enzymatichydrolysatesofsardinelle(Sardinellaaurita)by-products proteins[J]. Food Chemistry, 2010, 118(3):559-565
[5]KatalinicV,MilosM,KulisicT,etal.Screeningof70medicinalplantextractsforantioxidantcapacityandtotal phenols[J]. Food Chemistry, 2006, 94(4):550-557
[6]GibbsBF,ZougmanA,MasseR,etal.Productionand characterizationofbioactivepeptidesfromsoyhydrolysate andsoy-fermentedfood[J].FoodResearchInternational,2004, 37(2):123-131
[7]Parrado J, Miramontes E, Jover M, et al. Preparation of a rice bran enzymatic extract with potential use as functional food[J]. Food Chemistry, 2006, 98(4):742-748
[8]SakanakaS,TachibanaY,IshiharaN,etal.Antioxidant activityofegg-yolkproteinhydrolysatesinalinoleicacid oxidation system[J]. Food Chemistry, 2004, 86(1):99-103
[9]SaigaA,TanabeS,NishimuraT.Antioxidantactivityof peptidesobtainedfromporcinemyofibrillarproteinsby proteasetreatment[J].JournalofAgricultural&Food Chemistry, 2003, 51(12):3661-3667
[10]Zhong Y, MAK, Shahidi F. Compositional characteristics and antioxidantpropertiesoffreshandprocessedseacucumber(Cucumariafrondosa)[J].JournalofAgricultural&Food Chemistry, 2007, 55(4):1188
[11]张超.禽蛋生产消费市场供需基本平衡[J].农产品市场周刊,2017,16:34ZHANG Chao. Fundamental balance of supply and demand inpoultryeggproductionandconsumptionmarket[J].Market Weekly, 2017, 16:34-34
[12]李鑫,赵燕,李建科,等.蛋壳膜中美容活性物质的提取与应用[J].食品工业科技,2012,33(7):427-430LIXin,ZHAOYan,LIJian-ke,etal.Extractionand applicationofcosmeticactivesubstancesineggshell membranes[J].ScienceandTechnologyofFoodIndustry,2012, 33(7):427-430
[13]Nys,Gautron,McKee,etal.Biochemicalandfunctional characterisationofeggshellmatrixproteinsinhens[J].Worlds Poultry Science Journal, 2001, 57(4):401-413
[14]RuffKJ,DevoreDP.Reductionofpro-inflammatory cytokines in rats following 7-day oral supplementation with a proprietaryeggshellmembrane-derivedproduct[J].Modern Research in Inflammation, 2014, 3(1):19-25
[15]HuangX,ZhouY,MaM,etal.Chemiluminescence evaluationofantioxidantactivityandpreventionofDNA damageeffectofpeptidesisolatedfromsolubleeggshell membrane protein hydrolysate[J]. Journal of Agricultural&Food Chemistry, 2010, 58(23):12137-12142
[16]DevoreDP,LongFD,AdamsRG,etal.Therapeutic,nutraceuticalandcosmeticapplicationsforeggshell membraneandprocessedeggshellmembranepreparations:US, PCT/US2004/007832[P] 2004.09.23.
[17]周艳华.可溶性鸡蛋壳膜蛋白与多肽的制备及其抗氧化活性的研究[D].武汉:华中农业大学,2010ZHOU Yan-hua. Preparation of soluble egg shell membrane proteinandpolypeptideanditsantioxidantactivity[D].Wuhan:Huazhong Agricultural University, 2010
[18]HuangY,RuanG,QinZ,etal.Antioxidantactivity measurementandpotentialantioxidantpeptidesexploration fromhydrolysatesofnovelcontinuousmicrowave-assisted enzymolysisoftheScomberomorusniphoniusprotein[J].Food Chemistry, 2017, 223:89-95
[19]RuanG,ChenZ,WeiM,etal.Thestudyonmicrowave assisted enzymatic digestion of ginkgo protein[J]. Journal of Molecular Catalysis B:Enzymatic, 2013, 94:23-28
[20]尤娟.鲢鱼鱼肉蛋白抗氧化肽的制备及其糖基化产物功能特性的研究[D].北京:中国农业大学,2014YOUJuan.Preparationofantioxidantpeptidesofcarpfish proteinandfunctionalpropertiesofglycosylationproducts[D]. Beijing:China Agricultural University, 2014
[21]田旭静,段鹏慧,范三红,等.响应面法优化酶解藜麦糠蛋白制备抗氧化肽工艺[J].食品科学,2018,10:158-164TIANXu-jing,DUANPeng-hui,FANSan-hong,etal.Optimizationofantioxidantpeptidespreparedby enzymatic hydrolysisofbuckwheatproteinbyresponsesurface methodology[J]. Food Science, 2018, 10:158-164
[22]Jakoveti?S,Lukovi?N,Jugovi?B,etal.Productionof antioxidanteggwhitehydrolysatesinacontinuousstirred tank enzyme reactor coupled with membrane separation unit[J]. Food&Bioprocess Technology, 2015, 8(2):287-300
[23]Hamada JS. Use of proteases to enhance solubilization of rice bran proteins[J]. Journal of Food Biochemistry, 1999, 23(3):307-321
[24]Katoh K, Tanabe T, Yamauchi K. Novel approach to fabricate keratinspongescaffoldswithcontrolledporesizeand porosity[J]. Biomaterials, 2004, 25(18):4255-4262
[25]Ketnawa S, Benjakul S, Martínez-Alvarez O, et al. Fish skin gelatinhydrolysatesproducedbyvisceralpeptidaseand bovine trypsin:Bioactivity and stability[J]. Food Chemistry,2017, 215:383-390
[26]Klompong V, Benjakul S, Kantachote D, et al. Antioxidative activityandfunctionalpropertiesofproteinhydrolysateof yellow stripe trevally(Selaroides leptolepis)as influenced by thedegreeofhydrolysisandenzymetype[J].Food Chemistry, 2007, 102(4):1317-1327
[27]Nasri R, Jridi M, Lassoued I, et al. The influence of the extent ofenzymatichydrolysisonantioxidativepropertiesand ace-inhibitoryactivitiesofproteinhydrolysatesfromgoby(Zosterisessorophiocephalus)muscle[J].ApplBiochem Biotechnol, 2014, 173(5):1121-1134