高效液相色谱法同时测定发酵液中赤藓糖醇和L-赤藓酮糖的含量

建立了采用高效液相色谱(HPLC)同时测定发酵液中底物赤藓糖醇和产物L-赤藓酮糖含量的方法。采用Lichrospher 5-NH2色谱柱(250 mm×4.6 mm),柱温30 ℃,以乙腈-水(体积比为9:1)为流动相,流速1.0 mL/min。用示差折光检测器检测赤藓糖醇,检测器温度为35 ℃。用紫外检测器在室温下检测L-赤藓酮糖,检测波长为277 nm。所得赤藓糖醇的线性范围为1.00～100.00 g/L,相关系数为0.9985,检出限为0.10 g/L,定量限为0.45 g/L;所得L-赤藓酮糖的线性范围为1.00～100.00 g/L,相关系数为0.9958,检出限为0.50 g/L,定量限为0.87 g/L;赤藓糖醇的日内和日间相对标准偏差(RSD)分别小于3.28%和5.30%, L-赤藓酮糖的日内和日间RSD分别小于2.16%和2.25%;回收率均大于99%。取不同时间的发酵液样品分别用上述方法测定,结果表明所建立的HPLC法不受发酵液中其他组分的影响,可同时测定底物赤藓糖醇和产物L-赤藓酮糖的含量。

Simultaneous determination of meso-erythritol and L-erythrulose in fermentation broth using high performance liquid chromatography

A high performance liquid chromatographic(HPLC) method has been developed for the simultaneous determination of meso-erythritol and L-erythrulose in fermentation broth.The chromatographic conditions were as follows: Lichrospher 5-NH2 column(250 mm×4.6 mm) with the temperature of 30 ℃,acetonitrile-water(90∶10,v/v) as mobile phase with the flow rate of 1.0 mL/min.meso-Erythritol was detected by refractive index(RI) detector at 35 ℃ and L-erythrulose was detected by ultraviolet(UV) detector at 277 nm at room temperature.The linear range for meso-erythritol was 1.00-100.00 g/L with a correlation coefficient of 0.9985.The limit of detection(LOD) and the limit of quantification(LOQ) for meso-erythritol were 0.10 g/L and 0.45 g/L,respectively.The linear range for L-erythrulose was 1.00-100.00 g/L with a correlation coefficient of 0.9958.The LOD and LOQ for L-erythrulose were 0.50 g/L and 0.87 g/L,respectively.The relative standard deviations(RSDs) of intraday and interday for meso-erythritol were less than 3.28% and 5.30%,respectively.The intraday and interday RSDs for L-erythrulose were less than 2.16% and 2.25%,respectively.The recoveries of meso-erythritol and L-erythrulose in fermentation broth were greater than 99%.The samples from fermentation broth were detected at different time points.The measurement by the novel HPLC method was not affected by the other components in the fermentation broth.Furthermore,the HPLC method can be used for the determination of the substrate meso-erythritol and the product L-erythrulose simultaneously.