柠檬皮中柠檬苦素对根霉的抑菌活性和机理
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摘要
通过测定柠檬皮中柠檬苦素对根霉的最小抑菌浓度(MIC)、最小杀菌浓度(MBC)、孢子萌发抑制率、菌丝萌发抑制率确定其抑菌活性,根据测定各因素对抑菌活性的影响确定抑菌活性的稳定性;并通过扫描电镜观察和测定根霉细胞外碱性磷酸酶(AKP)含量和蛋白质含量,菌丝体的总糖含量,分析柠檬皮中柠檬苦素对根霉的抑菌机理。结果表明:柠檬皮中柠檬苦素对根霉的最低抑菌浓度(MIC)和最小杀菌浓度(MBC)分别为1250、5000μg/mL;对根霉孢子萌发的抑制效果更强,对孢子萌发和菌丝生长抑制的EC_(50)值分别为918.34和1707.31μg/mL。柠檬苦素对根霉的抑菌活性不受温度和明胶的影响;pH为4时,抑菌活性最强;Fe~(3+)、Fe~(2+)均能显著(p<0.05)提高柠檬苦素的抑菌活性。柠檬苦素能使根霉的糖、蛋白质、AKP等胞内物质渗出;扫描电镜观察也发现经柠檬苦素处理的根霉孢子凹陷和皱缩严重,且溶出物明显;菌丝出现断裂扭曲、变形、变细。因此,柠檬苦素能破坏根霉的细胞壁和细胞膜的完整性,使胞内物质渗出,菌丝体和孢子变形,从而使根霉生长代谢受到影响。
The antimicrobial activity of limonoids from lemon peel on Rhizopus was investigated by determining the minimal inhibitory concentration(MIC),the minimal bactericidal concentration(MBC),spore germination inhibition rate and mycelium germination inhibition rate.The stability of the antimicrobial activity was determined according to the influence of various factors.The antibacterial mechanism of limonoidsfrom lemon peel on Rhizopus was analyzed by observing scanning electron microscopy(SEM)and determining the contents of extracellular alkaline phosphatase(AKP),protein and total sugar in Rhizopus mycelium.The results showed that the minimum inhibitory concentration(MIC)and minimum bactericidal concentration(MBC)of limonoids on Rhizopus were 1250 and 5000 μg/mL respectively,and the EC_(50) values of limonoids on spore germination and mycelium growth were 918.34 and 1707.31 μg/mL respectively. Antimicrobial activity was strongest when pH was 4. The antibacterial activity of limonoids on Rhizopus was not influenced by temperature and gelatin,significantly(p<0.05)improved by Fe~(3+) and Fe~(2+). Limonoids could exudate intracellular substances in Rhizopus such as sugar,protein and AKP.The observation of scanning electron microscope(SEM)showed that the spores of Rhizopus treated with limonoids were depressed and shrunk seriously,and the dissolution was obvious.The mycelium of Rhizopus was distorted,deformed and thinned. Therefore,limonoids could destroy the integrity of the cell wall and membrane of Rhizopus,and the substance exudate out of the cell,anddeform mycelium and spore,thus affecting the growth and metabolism of Rhizopus.
The antimicrobial activity of limonoids from lemon peel on Rhizopus was investigated by determining the minimal inhibitory concentration(MIC),the minimal bactericidal concentration(MBC),spore germination inhibition rate and mycelium germination inhibition rate.The stability of the antimicrobial activity was determined according to the influence of various factors.The antibacterial mechanism of limonoidsfrom lemon peel on Rhizopus was analyzed by observing scanning electron microscopy(SEM)and determining the contents of extracellular alkaline phosphatase(AKP),protein and total sugar in Rhizopus mycelium.The results showed that the minimum inhibitory concentration(MIC)and minimum bactericidal concentration(MBC)of limonoids on Rhizopus were 1250 and 5000 μg/mL respectively,and the EC_(50) values of limonoids on spore germination and mycelium growth were 918.34 and 1707.31 μg/mL respectively. Antimicrobial activity was strongest when pH was 4. The antibacterial activity of limonoids on Rhizopus was not influenced by temperature and gelatin,significantly(p<0.05)improved by Fe~(3+) and Fe~(2+). Limonoids could exudate intracellular substances in Rhizopus such as sugar,protein and AKP.The observation of scanning electron microscope(SEM)showed that the spores of Rhizopus treated with limonoids were depressed and shrunk seriously,and the dissolution was obvious.The mycelium of Rhizopus was distorted,deformed and thinned. Therefore,limonoids could destroy the integrity of the cell wall and membrane of Rhizopus,and the substance exudate out of the cell,anddeform mycelium and spore,thus affecting the growth and metabolism of Rhizopus.
引文
[1]沈兆敏. 我国柠檬生产现状、优势机遇及发展对策意见[J]. 果农之友,2014(7):3-4,18.
[2]向羽. 柚类果实柠檬苦素类化合物组分鉴别、分离纯化及生物活性研究[D]. 杭州:浙江大学,2014.
[3]Lv M,Ping X,Yuan T,et al. Medicinal uses,phytochemistry and pharmacology of the genus Dictamnus(Ru-taceae)[J]. Journal of Ethnopharmacology,2015,17(3):48.
[4]Tundis R,Loizzo M R,Menchini F. An overview on chemical aspects and potential health benefits of limonoids and their derivatives[J]. Critical Reviews in Food Science and Nutrition,2014,54(2):225.
[5]刘影. 黄芩醇提物抑菌成分的初步确定及对黑根霉抑菌机理研究[D]. 南京:南京师范大学,2014.
[6]梁硕,杨志萍,费振鸿,等. 罗汉果抑菌性能的研究[J]. 食品工业,2016,37(7):207-209.
[7]薛桂新,黄世臣,宋益冬. 木醋液对桃黑根霉抑菌作用和对久保桃保鲜效果的研究[J]. 延边大学农学学报,2013,35(2):123-130.
[8]檀星,秦振华,田艺阁,等. 竹干黄酮的抑菌性能研究[J]. 化学与生物工程,2017,34(2):38-41,53.
[9]李彪,施蕊,熊智,等. 柚子皮中柠檬苦素提取工艺及其抑菌活性研究[J]. 食品工业科技,2012,33(5):198-200.
[10]李赤翎,俞建,蒋雪薇,等.柠檬苦素类似物糖苷的抗菌与抗氧化作用研究[J]. 食品科技,2008(5):154-156.
[11]张贝,白卫东,冯卫华,等. 柠檬皮中柠檬苦素的提取及其抑菌稳定性研究[J]. 安徽农业科学,2018,46(10):150-152.
[12]周梦娇,万春鹏,陈金印. 柑橘绿霉病中草药高效抑菌剂的筛选及抑菌机理研究[J]. 现代食品科技,2014,30(3):144-149,75.
[13]Qiu M,Wu C,Ren G,et al. Effect of chitosan and its derivatives as antifungal and preservative agents on postharvest green asparagus[J]. Food Chemistry,2014,155(15):105-111.
[14]Zhong Z M,Chen R,Xing R,et al. Synthesis and antifungal properties of sulfanilamide derivatives of chitosan[J]. Carbohydrate Research,2007,342(16):2390-2395.
[15]韩亚珊. 食品化学实验指导[M]. 北京:中国农业出版社,1996.
[16]任先伟,魏晓璐,黄鑫,等. 核桃青皮提取物抑菌活性及抑菌机理研究[J]. 食品工业科技,2015,36(18):93-98.
[17]蓝蔚青,车旭,谢晶,等.复合生物保鲜剂对荧光假单胞菌的抑菌活性及作用机理[J]. 中国食品学报,2016,16(8):159-165.
[18]张传军,刘超,姜晓坤.薤白乙醇提取物的抑菌特性[J]. 食品科学,2011,32(5):119-122.
[19]刘安军,李海燕,王云霞,等.核桃种皮多糖抑制大肠杆菌的研究[J].现代食品科技,2011,27(1):29-31.
[20]Pramanik K,Ghosh P K,Ray S,et al. An in silico structural,functional and phylogenetic analysis with three dimensional protein modeling of alkaline phosphatase enzyme of Pseudomonas aeruginosa[J]. Journal of Genetic Engineering and Biotechnology,2017,15(2):527-537.
[2]向羽. 柚类果实柠檬苦素类化合物组分鉴别、分离纯化及生物活性研究[D]. 杭州:浙江大学,2014.
[3]Lv M,Ping X,Yuan T,et al. Medicinal uses,phytochemistry and pharmacology of the genus Dictamnus(Ru-taceae)[J]. Journal of Ethnopharmacology,2015,17(3):48.
[4]Tundis R,Loizzo M R,Menchini F. An overview on chemical aspects and potential health benefits of limonoids and their derivatives[J]. Critical Reviews in Food Science and Nutrition,2014,54(2):225.
[5]刘影. 黄芩醇提物抑菌成分的初步确定及对黑根霉抑菌机理研究[D]. 南京:南京师范大学,2014.
[6]梁硕,杨志萍,费振鸿,等. 罗汉果抑菌性能的研究[J]. 食品工业,2016,37(7):207-209.
[7]薛桂新,黄世臣,宋益冬. 木醋液对桃黑根霉抑菌作用和对久保桃保鲜效果的研究[J]. 延边大学农学学报,2013,35(2):123-130.
[8]檀星,秦振华,田艺阁,等. 竹干黄酮的抑菌性能研究[J]. 化学与生物工程,2017,34(2):38-41,53.
[9]李彪,施蕊,熊智,等. 柚子皮中柠檬苦素提取工艺及其抑菌活性研究[J]. 食品工业科技,2012,33(5):198-200.
[10]李赤翎,俞建,蒋雪薇,等.柠檬苦素类似物糖苷的抗菌与抗氧化作用研究[J]. 食品科技,2008(5):154-156.
[11]张贝,白卫东,冯卫华,等. 柠檬皮中柠檬苦素的提取及其抑菌稳定性研究[J]. 安徽农业科学,2018,46(10):150-152.
[12]周梦娇,万春鹏,陈金印. 柑橘绿霉病中草药高效抑菌剂的筛选及抑菌机理研究[J]. 现代食品科技,2014,30(3):144-149,75.
[13]Qiu M,Wu C,Ren G,et al. Effect of chitosan and its derivatives as antifungal and preservative agents on postharvest green asparagus[J]. Food Chemistry,2014,155(15):105-111.
[14]Zhong Z M,Chen R,Xing R,et al. Synthesis and antifungal properties of sulfanilamide derivatives of chitosan[J]. Carbohydrate Research,2007,342(16):2390-2395.
[15]韩亚珊. 食品化学实验指导[M]. 北京:中国农业出版社,1996.
[16]任先伟,魏晓璐,黄鑫,等. 核桃青皮提取物抑菌活性及抑菌机理研究[J]. 食品工业科技,2015,36(18):93-98.
[17]蓝蔚青,车旭,谢晶,等.复合生物保鲜剂对荧光假单胞菌的抑菌活性及作用机理[J]. 中国食品学报,2016,16(8):159-165.
[18]张传军,刘超,姜晓坤.薤白乙醇提取物的抑菌特性[J]. 食品科学,2011,32(5):119-122.
[19]刘安军,李海燕,王云霞,等.核桃种皮多糖抑制大肠杆菌的研究[J].现代食品科技,2011,27(1):29-31.
[20]Pramanik K,Ghosh P K,Ray S,et al. An in silico structural,functional and phylogenetic analysis with three dimensional protein modeling of alkaline phosphatase enzyme of Pseudomonas aeruginosa[J]. Journal of Genetic Engineering and Biotechnology,2017,15(2):527-537.