One-pot labeling-based capillary zone electrophoresis for separation of amino acid mixture and assay of biofluids

A fast, simple and cost-effective one-pot labeling strategy coupled with capillary zone electrophoresis was developed for the complete separation of amino acid mixture. The strategy includes two steps of reactions: Cyanuric chloride was made to react first with 7-amino-1,3-naphthalenedisulfonic acid monopotassium salt at 0 °C for 10 min, and then with amino acids at 55 °C for 6 min. The resulted products, after diluted with water, were injected into capillary zone electrophoresis system for separation. Using a running buffer of 20 mM sodium tetraborate decahydrate at pH 10.1, nineteen amino acids were efficiently separated in 25 min, with relative standard deviation of 0.36–1.6% and 0.96–2.1% (within and between days, respectively) for migration time and 0.030–1.6% and 0.22–2.4% (within and between days, respectively) for peak area. The proposed method has been successfully applied to the determination of free amino acids in biofluids, including human serum, urine, and saliva. The linearity of quantification was over two orders of magnitude for most amino acids, with a correlation coefficient larger than 0.999. The average recovery, determined by spiking a known amount of amino acid standards into real samples, was in a range from 91.6% to 105.9%. This method can be a noninvasive means since it could directly assay the urine and saliva samples.