虾头、虾壳抗氧化肽的分离纯化及其对秀丽隐杆线虫的抗氧化作用
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摘要
采用超滤、凝胶过滤色谱和制备液相色谱对南美白对虾虾头、虾壳蛋白质的酶解液进行分离纯化,以1,1-二苯基-2-苦基肼自由基清除率、还原力、氧化自由基吸收能力为指标,制备纯度较高、抗氧化活性较强的抗氧化肽,同时,采用秀丽隐杆线虫(Caenorhabditis elegans)模型对其体内抗氧化活性进行评价。结果表明:酶解液经超滤分离后,分子质量为3～10 kDa组分的抗氧化活性最强;将该组分采用凝胶过滤色谱、制备液相色谱等手段分离出含有抗氧化肽的混合物(抗氧化肽纯度达到70%)。体内抗氧化活性结果显示,喂食该抗氧化肽的秀丽隐杆线虫的产卵量和热应激条件下的存活率均显著增长,寿命显著延长(P<0.05),且与VC相比效果无显著差异(P>0.05),说明本研究所获得的抗氧化肽对秀丽隐杆线虫具有和VC相当的抗氧化保护作用,具有潜在的应用价值。
Highly pure peptides with strong antioxidant activity in terms of 1,1-diphenyl-2-picrylhydrazyl(DPPH) radical scavenging capacity, reducing power and oxygen radical absorbance capacity(ORAC) were obtained from the enzymatic hydrolysates of heads and shells of Penaeus vannamei by sequential ultrafiltration, gel filtration chromatography and preparative liquid chromatography. Furthermore, Caenorhabditis elegans were used to determine the antioxidant activity in vivo of the obtained antioxidant peptides. The results showed that the ultrafiltration fraction with molecular mass of3–10 kDa exhibited the highest antioxidant activity. Further puri?cation by gel ?ltration chromatography and preparative liquid chromatography could obtain a mixture of antioxidant peptides with a purity of 70%. The investigation of antioxidant activity in vivo showed that Caenorhabditis elegans fed with the puri?ed antioxidant peptides laid more eggs and had higher survival rate under heat stress as well as significantly longer lifespan(P < 0.05), while the effect was not significantly different from that of VC(P > 0.05), indicating that the antioxidant peptides obtained in this study had protective effect against oxidative stress in Caenorhabditis elegans similar to that of VC and therefore could have potential application value.
Highly pure peptides with strong antioxidant activity in terms of 1,1-diphenyl-2-picrylhydrazyl(DPPH) radical scavenging capacity, reducing power and oxygen radical absorbance capacity(ORAC) were obtained from the enzymatic hydrolysates of heads and shells of Penaeus vannamei by sequential ultrafiltration, gel filtration chromatography and preparative liquid chromatography. Furthermore, Caenorhabditis elegans were used to determine the antioxidant activity in vivo of the obtained antioxidant peptides. The results showed that the ultrafiltration fraction with molecular mass of3–10 kDa exhibited the highest antioxidant activity. Further puri?cation by gel ?ltration chromatography and preparative liquid chromatography could obtain a mixture of antioxidant peptides with a purity of 70%. The investigation of antioxidant activity in vivo showed that Caenorhabditis elegans fed with the puri?ed antioxidant peptides laid more eggs and had higher survival rate under heat stress as well as significantly longer lifespan(P < 0.05), while the effect was not significantly different from that of VC(P > 0.05), indicating that the antioxidant peptides obtained in this study had protective effect against oxidative stress in Caenorhabditis elegans similar to that of VC and therefore could have potential application value.
引文
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[27]CASTEL V,ANDRICH O,NETTO F M,et al.Comparison between isoelectric precipitation and ultrafiltration processes to obtain Amaranth mantegazzianus protein concentrates at pilot plant scale[J].Journal of Food Engineering,2012,112(4):288-295.DOI:10.1016/j.jfoodeng.2012.05.010.
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[2]李二超.盐度对凡纳滨对虾的生理影响及其营养调节[D].上海:华东师范大学,2008:1-7.
[3]张伟权.世界重要养殖品种:南美白对虾生物学简介[J].海洋科学,1990,14(3):69-73.
[4]薛聪顺,戴海平,蔡惠凤,等.南美白对虾套养中华鳖试验[J].中国水产,2011(8):29-30.
[5]边晶晶,谢晶,陈舜胜.南美白对虾虾头中磷脂提取工艺的优化[J].食品科学,2011,32(24):11-15.
[6]刘洪亮,陈丽娇.对虾虾头、虾壳副产品综合利用的研究概况[J].福建水产,2011,33(2):65-69.DOI:10.3969/j.issn.1006-5601.2011.02.014.
[7]张祥刚,周爱梅,林晓霞,等.南美白对虾虾头、虾壳化学成分的对比研究[J].现代食品科技,2009,25(3):224-227.DOI:10.3969/j.issn.1673-9078.2009.03.002.
[8]张高静,韩丽萍,孙剑锋,等.南美白对虾营养成分分析与评价[J].中国食品学报,2013,13(8):254-260.
[9]COWARD-KELLY G,AGBOGBO F K,HOLTZAPPLE M T.Lime treatment of shrimp head waste for the generation of highly digestible animal feed[J].Bioresource Technology,2006,97(13):1515-1520.DOI:10.1016/j.biortech.2005.06.014.
[10]周爱梅,张风,银凤,等.利用对虾加工下脚料制备抗氧化肽的研究[J].中国食品学报,2014,14(8):85-91.
[11]SHAZLY A B,HE Z Y,EL-AZIZ M A,et al.Fractionation and identification of novel antioxidant peptides from buffalo and bovine casein hydrolysates[J].Food Chemistry,2017,232:753-762.DOI:10.1016/j.foodchem.2017.04.071.
[12]范建凤,王泽南,杨柯,等.蟹抗氧化肽的分离纯化及活性研究[J].食品科学,2010,31(13):48-51.
[13]HAMEDY P A,O’KEEFE M B,FITZGERALD R J.Fractionation and identification of antioxidant peptides from an enzymatically hydrolysed Palmaria palmata protein isolate[J].Food Research International,2017,100(1):416-422.DOI:10.1016/j.foodres.2017.07.037.
[14]LIU J B,JIN Y,LIN S Y,et al.Purification and identification of novel antioxidant peptides from egg white protein and their antioxidant activities[J].Food Chemistry,2015,175:258-266.DOI:10.1016/j.foodchem.2014.11.142.
[15]OHATA M,UCHIDA S,ZHOU L X,et al.Antioxidant activity of fermented meat sauce and isolation of an associated antioxidant peptide[J].Food Chemistry,2016,194:1034-1039.DOI:10.1016/j.foodchem.2015.08.089.
[16]PETERSEN C,DIRKSEN P,SCHULENBURG H.Why we need more ecology for genetic models such as C.elegans[J].Trends in Genetics,2015,31(3):120-127.DOI:10.1016/j.tig.2014.12.001.
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[19]NAJAFIAN L,BABJI A S.Production of bioactive peptides using enzymatic hydrolysis and identification antioxidative peptides from patin(Pangasius sutchi)sarcoplasmic protein hydolysate[J].Journal of Functional Foods,2014,9:280-289.DOI:10.1016/j.jff.2014.05.003.
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[21]戴卉卿,陈承瑜,杨滨.22种黄酮、酚酸类化合物和9种中药清除AAPH活性考察[J].中国中药杂志,2010,35(17):2296-2302.
[22]陈运娇,李伟,陈洪璋,等.酪蛋白-葡萄糖-PGG纳米复合物的制备及其延缓线虫衰老能力的研究[J].现代食品科技,2016,32(8):21-27.
[23]CHEN Y,ONKEN B,CHEN H,et al.Mechanism of longevity extension of Caenorhabditis elegans induced by pentagalloyl glucose isolated from eucalyptus leaves[J].Journal of Agricultural and Food Chemistry,2014,62(15):3422-3431.DOI:10.1021/jf500210p.
[24]BRAIN O,MONICA D.Metformin induces a dietary restrictionlike state and the oxidative stress response to extend,C.elegans,healthspan via AMPK,LKB1,and SKN-1[J].PLoS ONE,2010,5(1):e8758.DOI:10.1371/journal.pone.0008758.
[25]卢伟伟,操晶晶,胡玉,等.氯化甲基汞对秀丽隐杆线虫运动和感觉功能的影响[J].环境与健康杂志,2015,32(7):565-568.
[26]郝绵霞.秀丽隐杆线虫AMPK蛋白AAK-2及线粒体腺苷酸转运蛋白ANT-1.1在热应激中的功能研究[D].厦门:厦门大学,2016:1-7.
[27]CASTEL V,ANDRICH O,NETTO F M,et al.Comparison between isoelectric precipitation and ultrafiltration processes to obtain Amaranth mantegazzianus protein concentrates at pilot plant scale[J].Journal of Food Engineering,2012,112(4):288-295.DOI:10.1016/j.jfoodeng.2012.05.010.
[28]殷健,李万芳,王爱平,等.氧自由基吸收能力分析法的发展和应用[J].中国食品卫生杂志,2013,25(1):97-101.
[29]ZHANG Longze,JIE Guoliang,ZHANG Junjing,et al.Significant longevity-extending effects of EGCG on Caenorhabditis elegans under stress[J].Free Radical Biology and Medicine,2009,46(3):414-421.DOI:10.1016/j.freeradbiomed.2008.10.041.
[30]ZHOU J B,ZHENG Y L,ZENG Y X,et al.Marine derived xyloketal derivatives exhibit anti-stress and anti-ageing effects through HSFpathway in Caenorhabditis elegans[J].European Journal of Medicinal Chemistry,2018,148:63-72.DOI:10.1016/j.ejmech.2018.02.028.
[31]MUNOZ M J,RIDDLE D L.Positive selection of Caenorhabditis elegans mutants with increased stress resistance and longevity[J].Genetics,2003,163(1):171-180.