粪肠球菌DU10产菌素的部分纯化、鉴定及其益生菌特性

Venkatesh Perumal, Ayyanna Repally, Ankaiah Dasari, Arul Venkatesan
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引用次数: 13

摘要

摘要从鸭分离的乳酸菌粪肠球菌DU10中分离出一种新的细菌素。该细菌素对重要食源性致病菌具有广谱的抗菌活性,经C-18柱的粒径排除层析和反相高效液相层析纯化得到。Tricine-SDS PAGE显示存在一个条带,估计分子质量为6.3 kDa。酶谱图清楚地将抗菌活性与该条带联系起来。质量辅助激光解吸电离飞行时间质谱进一步证实了这一结果,因为检测到一个对应于6.313 kDa的尖峰,并且傅里叶变换红外光谱显示了官能团。细菌素DU10活性受蛋白酶k和胃蛋白酶敏感,部分受胰蛋白酶和α-凝乳胰蛋白酶影响。细菌素DU10对30 ~ 90℃热处理30min具有部分抗性。在121°C下处理10分钟。甲基噻唑基二苯基溴化四氮唑法测定细菌素DU10的细胞毒性结果显示,在细菌素浓度为3200 AU/mL时,HT-29和HeLa细胞活力分别下降60±0.7%和43±4.8%。菌株能承受0.3% w/v的胆汁酸,pH值在培养2 ~ 4小时内影响细菌生长。用HT-29细胞株检测黏附特性,发现菌株粪肠球菌DU10的初始群体为69.85%,对该细胞株具有较强的黏附性。综上所述,细菌素DU10可作为一种潜在的生物保存剂,粪肠杆菌DU10可作为一种潜在的益生菌控制沙门氏菌感染。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Partial purification and characterization of bacteriocin produced by Enterococcus faecalis DU10 and its probiotic attributes
ABSTRACT A novel bacteriocin produced by avian duck isolated lactic acid bacterium Enterococcus faecalis DU10 was isolated. This bacteriocin showed a broad spectrum of antibacterial activity against important food-borne pathogens and was purified by size exclusion chromatography followed by reverse-phase high-performance liquid chromatography in a C-18 column. Tricine–SDS PAGE revealed the presence of a band with an estimated molecular mass of 6.3 kDa. The zymogram clearly linked the antimicrobial activity with this band. This result was further confirmed by mass-assisted laser desorption ionization time-of-flight mass spectrometry, since a sharp peak corresponding to 6.313 kDa was detected and the functional groups were revealed by Fourier transform infrared spectroscopy. Bacteriocin DU10 activity was found sensitive to proteinase-K and pepsin and partially affected by trypsin and α-chymotrypsin. The activity of bacteriocin DU10 was partially resistant to heat treatments ranging from 30 to 90°C for 30 min. It also withstood a treatment at 121°C for 10 min. Cytotoxicity of bacteriocin DU10 by methyl-thiazolyl-diphenyl-tetrazolium bromide assay showed that the viability of HT-29 and HeLa cells decreased 60 ± 0.7% and 43 ± 4.8%, respectively, in the presence of 3,200 AU/mL of bacteriocin. The strain withstood 0.3% w/v of bile oxgall and pH 2 affected the bacterial growth between 2 and 4 hr of incubation. Adhesion properties examined with HT-29 cell line showed 69.85% initial population of strain E. faecalis DU10, which was found to be strongly adhered to this cell line. These results conclude bacteriocin DU10 may be used as a potential biopreservative and E. faecalis DU10 may be used as a potential probiont to control Salmonella infections.
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