辣木叶蛋白质酶解产物的制备及其抗菌活性研究
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摘要
为了研究辣木叶蛋白及其酶解产物的抗菌活性,利用碱提酸沉法提取辣木叶中的蛋白质,并分析其氨基酸组成;分别采用6种单酶和3种复合酶对辣木叶蛋白进行酶解,考察辣木叶蛋白及其酶解产物的抑菌活性,确定最佳蛋白酶。通过二倍稀释法,研究有效抑菌组分对不同菌的最小抑菌浓度(minimum inhibitory concentration,MIC)。结果显示:辣木叶蛋白提取率为32.67%±0.45%,必需氨基酸占氨基酸总量的39.27%。抑菌活性研究表明辣木叶蛋白仅对金黄色葡萄球菌显示抑菌活性,抑菌圈大小为(7.67±0.24) mm;胃蛋白酶结合胰蛋白酶分步酶解蛋白所得酶解产物抑菌能力均高于其他几种蛋白酶,酶解产物对化脓性链球菌的抑菌活性最强,抑菌圈大小为(10.83±0.62) mm,且其对金黄色葡萄球菌、单增李斯特菌及化脓性链球菌的MIC分别为5、10、2.5 mg/mL。说明辣木叶蛋白及其酶解产物均有抗菌活性,双酶酶解法制备的酶解产物具有较强的抑菌性及广抑菌谱。
In order to study the antimicrobial activity of horseradish leaf protein and its enzymatic hydrolysate,the protein in Moringa oleifera leaf was extracted by alkali extraction and acid precipitation,and its amino acid composition was analyzed. Using six single enzymes and three complex enzymes to enzymatic hydrolysis protein,the antimicrobial activity of Moringa oleifera leaves protein and its enzymatic hydrolysates was investigated to determine the best protease. The minimal inhibitory concentration(MIC)of effective antibacterial components on different bacteria was studied by double broth dilution method. The results showed that the extraction rate of protein was 32.67%±0.45%,and essential amino acids account for 39.27% of total amino acids. The research of antibacterial activity showed that the protein of Moringa oleifera leaves showed antibacterial activity only against Staphylococcus aureus and the antimicrobial zone diameter was(7.67±0.24) mm. The antimicrobial activity of the enzymatic hydrolysate obtained by sequential hydrolysis with pepsin followed by trypsin was higher than that of other proteases. The enzymatic hydrolysate had the strongest antibacterial activity for Streptococcus pyogenes and the antimicrobial zone diameter was(10.83±0.62) mm. The minimum inhibitory concentration(MIC)were 5,10 and 2.5 mg/mL,respectively. The results showed that both the protein and its enzymatic hydrolysate of Moringa oleifera leaf had antimicrobial activity,and the enzymatic hydrolysate prepared by double enzymatic hydrolysis had strong antimicrobial activity and wide antimicrobial spectrum.
In order to study the antimicrobial activity of horseradish leaf protein and its enzymatic hydrolysate,the protein in Moringa oleifera leaf was extracted by alkali extraction and acid precipitation,and its amino acid composition was analyzed. Using six single enzymes and three complex enzymes to enzymatic hydrolysis protein,the antimicrobial activity of Moringa oleifera leaves protein and its enzymatic hydrolysates was investigated to determine the best protease. The minimal inhibitory concentration(MIC)of effective antibacterial components on different bacteria was studied by double broth dilution method. The results showed that the extraction rate of protein was 32.67%±0.45%,and essential amino acids account for 39.27% of total amino acids. The research of antibacterial activity showed that the protein of Moringa oleifera leaves showed antibacterial activity only against Staphylococcus aureus and the antimicrobial zone diameter was(7.67±0.24) mm. The antimicrobial activity of the enzymatic hydrolysate obtained by sequential hydrolysis with pepsin followed by trypsin was higher than that of other proteases. The enzymatic hydrolysate had the strongest antibacterial activity for Streptococcus pyogenes and the antimicrobial zone diameter was(10.83±0.62) mm. The minimum inhibitory concentration(MIC)were 5,10 and 2.5 mg/mL,respectively. The results showed that both the protein and its enzymatic hydrolysate of Moringa oleifera leaf had antimicrobial activity,and the enzymatic hydrolysate prepared by double enzymatic hydrolysis had strong antimicrobial activity and wide antimicrobial spectrum.
引文
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[2]彭勃,冯孔龙,苗建银,等. 副干酪乳杆菌FX-6产抗菌肽粗提物对荔枝贮藏品质的影响[J]. 食品科学,2018,39(7):249-255.
[3]Lee Y,Phat C,Hong S C. Structural diversity of marine cyclic peptides and their molecular mechanisms for anticancer,antibacterial,antifungal and other clinical applications[J]. Peptides,2017,95:94.
[4]Saucedo S,Torres J A,Castro C,et al. Moringa plants:Bioactive compounds and promising applications in food products[J]. Food Research International,2018,2018(111):438-450.
[5]Yang L,Xiao-Yue W,Xue-Min W,et al. Values,properties and utility of different parts of Moringa oleifera:An overview[J]. Chinese Herbal Medicines,2018,10:371-378.
[6]汪泰,顾文宏,何军,等辣木新资源食品研究进展[J]. 食品工业科技,2017,38(8):363-368.
[7]Zhou C L,Mi L,Hu X Y,et al. Evaluation of three pumpkin species:Correlation with physicochemical,antioxidant properties and classification using SPME-GC-MS and E-nose methods[J].Journal Food Science and Technology,2017,54(10):3118-3131.
[8]Tiloke C,Anand K M,Gengan R,et al. Moringa oleifera and their phytonanoparticles:Potential antiproliferative agents against cancer[J]. Biomedicine & Pharmacotherapy,2018,108:457-466.
[9]Bolaňos X,Vega R C,Camacho R R,et al. Moringa infusion(Moringa oleifera)rich in phenolic compounds and high antioxidant capacity attenuate nitric oxide pro-inflammatory mediator in vitro[J]. Industraial Crops and Products,2018,118:95-101.
[10]Jayawardana B C,Liyanage R,Lalantha N,et al. Antioxidant and antimicrobial activity of drumstick(Moringa oleifera)leaves in herbal chicken sausages[J]. LWT-Food Science and Technology,2015,64(2):1204-1208.
[11]Yang F,Liu X,Ren X,et al. Swirling cavitation improves the emulsifying properties of commercial soy protein isolate[J]. Ultrasonics Sonochemistry,2018,42:471-481.
[12]Falowo A B,Mukumbo F E,Idamokoro E M,et al. Multi-functional application of Moringa oleifera Lam. in nutrition and animal food products:A review[J]. Food Research International,2018,106:317.
[13]施娅楠,和丽菊,赵存朝,等. 辣木叶渣中非水溶性蛋白及酶解效果研究[J]. 食品研究与开发,2018(2):103-108.
[14]韩志萍,黄茂芳,静玮,等超滤分离纯化菠萝蛋白酶的研究[J]. 食品工业科技,2013,34(8):282-286.
[15]Paula P C,Sousa D O,Oliveira J T,et al. A Protein isolate from Moringa oleifera leaves has hypoglycemic and antioxidant effects in alloxan-induced diabetic mice[J]. Molecules,2017,22(2):271.
[16]Kaushik P,Dowling K,Mcknight S,et al. Preparation,characterization and functional properties of flax seed protein isolate[J]. Food Chemistry,2016,197(A):212-220.
[17]Jemil I,Abdelhedi O,Nasri R,et al. Novel bioactive peptides from enzymatic hydrolysate of Sardinella(Sardinella aurita)muscle proteins hydrolysed by Bacillus subtilis A26 proteases[J]. Food Research International,2017,100(1):121.
[18]Kobbi S,Balti R,Bougatef A,et al. Antibacterial activity of novel peptides isolated from protein hydrolysates of RuBisCO purified from green juice alfalfa[J]. Journal of Functional Foods,2015,18:703-713.
[19]Shakeri A,Khakdan F,Soheili V,et al. Chemical composition,antibacterial activity,and cytotoxicity of essential oil from Nepeta ucrainica L. spp. kopetdaghensis[J]. Industrial Crops & Products,2014,58(7):315-321.
[20]熊瑶. 辣木叶蛋白质提取及其饮品研制[D]. 福州:福建农林大学,2012.
[21]吕晓亚,白新鹏,伍曾利,等. 辣木叶水溶性蛋白的超声-微波萃取及其性质研究[J]. 食品工业科技,2016,37(5):212-216.
[22]陈英娇. 苦荞蛋白酶解物的制备及抗菌活性的研究[D]. 上海:上海师范大学,2015:30-40.
[23]Zhang Y,Wu Y T,Zheng W,et al. The antibacterial activity and antibacterial mechanism of a polysaccharide from Cordyceps cicadae[J]. Journal of Functional Foods,2017,38:273-279.
[24]Wald M,Schwarz K,Rehbein H,et al. Detection of antibacterial activity of an enzymatic hydrolysate generated by processing rainbow trout by-products with trout pepsin[J]. Food Chemistry,2016,205:221-228.
[25]Sikorska E,Stachurski O,Neubauer D,et al. Short arginine-rich lipopeptides:From self-assembly to antimicrobial activity[J].Biochimica et Biophysica Acta(BBA)-Biomembranes,2018,1860:2242-2251.
[26]施永清,王巧巧,吴丹丽,等. 响应面试验优化双酶酶解法制备鱼鳞抗菌肽工艺及其抑菌性能分析[J]. 食品科学,2018,39(6):155-161.
[27]宋风霞,酶解苦荞蛋白制备抗菌肽及其在牛肉保鲜的研究[D]. 昆明:昆明理工大学,2017.
[28]孙宜君. 螺旋藻抗菌肽的纯化鉴定及其抑菌机理的研究[D]. 北京林业大学,2016:37-47.