高效阴离子交换色谱-脉冲安培检测法同时测定生物转化样品中的海藻糖、葡萄糖和麦芽糖

摘要

An analytical method for the determination of trehalose,maltose,and glucose in biotransformation samples was developed by using high performance anion exchange chroma-tography coupled with pulsed ampere detection( HPAEC-PAD). The analysis was performed on a CarboPacTM 10 column( 250 mm × 2 mm ) with the gradient elution of NaOH-NaAc as the mobile phase. The column temperature was set at 30 ℃,the flow rate was 0. 30 mL/min. The results showed that trehalose,maltose,and glucose in biotransformation system were com-pletely separated and determined in 15 min. The linear ranges and the working curves were determined by using standard samples. The correlation coefficients of three kinds of carbohy-drates were over 0. 999 8 . The detection limits( LODs)were 0. 010-0. 100 mg/L. Under the optimized separation conditions,the recoveries of saccharides in the transformation system at three different spiked levels ranged from 89. 4% to 103. 2%. In biotransformation system,50 IU trehalose synthase were added into 200 g/L maltose for reaction of 8 h at 37 ℃,pH 8. 0. Under the above conditions,the concentration of trehalose in biotransformation sample was 101. 084 g/L,and the conversion rate of trehalose reached 50. 5%. The method can be applied to deter-mine the composition in the transformation system with the advantages of simplicity and con-venience.%建立了高效阴离子交换色谱-脉冲安培电化学检测法同时测定生物转化样品中海藻糖、葡萄糖和麦芽糖的分析方法。选用 CarboPacTM10色谱柱（250 mm×2 mm）对分离条件进行优化，使用标准样品测定了线性范围和工作曲线，柱温为30℃，流速为0.30 mL／min，以氢氧化钠溶液和醋酸钠溶液为流动相进行梯度洗脱，脉冲安培法进行检测。研究结果表明，该方法可在15 min内实现海藻糖生物转化液中3种糖的快速定量分析。海藻糖、葡萄糖和麦芽糖峰面积与质量浓度的线性关系良好，检出限为0.010~0.100 mg／L。将此方法用于酶法制备海藻糖的检测，加标回收率为89.40％~103.2％。在生物转化样品中检测到海藻糖浓度为101.084 g／L，转化率达到了50.5％。该方法灵敏度高，简便快速，可应用于海藻糖制备样品中各种成分的分离和定量检测。