固相萃取-离子色谱-积分脉冲安培法测定人血清中游离氨基酸

Determination of Free Amino Acids in Human Serum Using Solid Phase Extraction Followed by Anion-exchange Chromatography with Integrated Pulsed Amperometric Detection

比较了磺基水杨酸和无水甲醇沉淀人血清中蛋白质的效果，优化了国产732型阳离子交换树脂进行固相萃取净化的条件和Dionex-600离子色谱分析系统的梯度淋洗条件。用该离子色谱仪检测了谷氨酰胺等14种氨基酸，结果表明，峰面积-浓度的校正曲线的相关系数为0．9900～0．9995，检出限为0.02～0.59mg／L；峰面积的相对标准偏差为1．9％～10．4％；回收率为80．6％～114．8％。

The efficiencies of precipitating proteins in serum by sulfosalicylic acid and by anhydrous methanol were compared and the conditions of solid phase extraction for purification with 732 cation-exchange resin and gradient conditions of Dionex-600 ion chromatograph （IC） were optimized. On adding 0.8 mL sulfosalicylic acid, 0.2 mL of human serum was centrifuged at 15 000 r/min for 15 rain. All supernatant was diluted to 10 mL, and 2.50 mol/L NaOH was added to adjust pH to 3.0, and then extracted on 732 cation-exchange resin. Finally, the amino acids was eluted by 5 mL ammonia（32：68, V/V）. After the ammonia being volatilized over in water bath at 80℃, the eluent was detected by the IC. The separation was performed at a column temperature of 30℃ and at a flow rate of 0.22 mL/min. Fourteen amino acids such as glutamine in human serum were detected by the IC. The calibration curve of peak area to concentration for amino acids was linear with a correlation coefficient of 0.9900 - 0.9995. The minimum detection concentrations, the RSDs of peak area and the recoveries of samples for amino acids were 0.02 - 0.59 mg/L, 1.9% - 10.4% , 80.6% - 114. 8%, respectively.