大肠杆菌DNA和RNA中核苷的高效液相色谱分析

Analysis of Nucleosides in DNA and RNA of E.coli by RP-HPLC

中文关键词:  高效液相色谱法  大肠杆菌  脱氧核糖核酸  核糖核酸  核苷

英文关键词:RP-HPLC  E.coli  DNA  RNA  nucleosides

中文摘要:

      建立了用反相高效液相色谱法(RPHPLC)测定大肠杆菌(E.coli)脱氧核糖核酸(DNA)和核糖核酸(RNA)水解产物中核苷的方法.色谱柱为SupelcoInc.C₁₈反相柱,254nm和280nm紫外吸收波长同时检测,分析条件经选择在流速为1mL/min,流动相A(2.5%甲醇+0.01mol/LKH₂PO₄,pH4.6)28min,流动相B(8.0%甲醇+0.01mol/LKH₂PO₄,pH4.6)37min,柱温为20℃条件下65min可实现核苷的完全分离.回收率为80.19%～99.31%,相对标准偏差RSD为0.86%～2.62%.该方法具有高灵敏度、高选择性的特点,得到的结果也十分满意.

英文摘要:

      A highly sensitive and selective reversed phase high performance liquid chromatography (RP-HPLC) method for the analysis of nucleosides in DNA and RNA of E.coli is described. The analytical column was Supelco Inc.C₁₈ (250 mm×4.5 mm) column(at 20℃). The separation of the nucleosides was achieved in 65 min using a two buffer step gradient system. Buffer A (2.5% v/v methanol, 0.01 mol/L KH₂PO₄, pH 4.6) was pumped for the first 28 min of the run, followed by 37 min of Buffer B (8.0% v/v methanol, 0.01 mol/L KH₂PO₄, pH 4.6). Absorption was measured simultaneously at 254 nm and 280 nm with sensitivity of 0.01 aufs and the flow rate was 1.0 mL/min. The relative standard deviation (RSD) were from 0.86% to 2.62% and the recoveries ranged between 80.20%～99.31%.