高效液相色谱法测定唾液乳杆菌脱基因毒性作用

采用高效液相色谱法(HPLC)测定唾液乳杆菌对4-硝基喹啉-N-氧化物(4-NQO)基因毒性的抑制作用。Lact.salivariusFDB89与4-NQO(终浓度为20mg/L)共培养后,利用SOS-显色反应法检测唾液乳杆菌的脱基因毒性作用,同时采用高效液相色谱分析共培养前后的4-NQO含量变化。色谱分析条件为:C18反相色谱柱(kromasil100-5C18),以乙腈、水为流动相进行梯度洗脱,流速1mL/min;检测波长365nm。结果表明,唾液乳杆菌转化4-NQO,降低了4-NQO的基因毒性;HPLC测定的4-NQO峰面积与SOS-显色反应测定的基因毒性值呈正相关,相关系数r2为0.991。HPLC方法测定脱基因毒性检出限为1.2ng,菌体浓度在106~109cfu/mL范围内回收率为85%~98%。HPLC法测定唾液乳杆菌的脱基因毒性操作简单,其重复性和精确度均优于SOS-显色反应,可以采用HPLC代替SOS-显色反应。本研究在应用仪器分析方法代替生化分析方法测定乳杆菌脱基因毒性方面提供了一条新思路。

Analysis of Antigenotoxicity of Lactobacillus Salivarius by High Performance Liquid Chromatography

High performance liquid chromatography was used to determine the antigenotoxic activity of Lactobacillus salivarius(Lact.salivarius)against 4-nitroquinolin-N-oxide(4-NQO).After co-incubation of Lact.salivarius FDB89 with 4-NQO(20 mg/L)for 3 hours,SOS-chromotest assay was carried out to test the antigenotoxicity of Lact.salivarius FDB89,and simultaneously HPLC assay was used to determine the quantitative change of 4-NQO before and after co-incubation with Lact.salivarius FDB89.HPLC assay was performed on a reversed phase C18 column with UV detection at 365 nm and the flow rate was 1.0 mL/min.Results showed that 4-NQO was converted to less toxic compound by Lact.salivarius FDB89 and so its genotoxicity was inhibited.A positive correlation between genotoxicity measured in SOS-chromotest and 4-NQO area integral in HPLC was observed with a square correlation coefficient(r2)being 0.991.Under the above conditions,instrument detection limits was 1.2 ng.The recovery of 4-NQO was 85%-98% when the cell densities was in the range of 106-109cfu/mL.The results suggested that the HPLC method can be a usefulalternative to SOS-chromotest in detecting the antigenotoxicity of lactobacillus as being more sensitive and accurate than SOS-chromotest.