Department of Applied Biological Chemistry, The University of Tokyo, Yayoi, Bunkyo, Tokyo 113-8657, Japan.
J Chromatogr B Analyt Technol Biomed Life Sci. 2012 Nov 15;909:34-6. doi: 10.1016/j.jchromb.2012.10.005. Epub 2012 Oct 8.
HPLC eluent systems employing acetonitrile and methanol were evaluated for the quantitation of glutathione (GSH) and phytochelatin (PC(n)), a family of peptides implicated in heavy-metal detoxification in higher plants. The detection system is based on the dequenching of copper(I)-bathocuproine disulfonate and is specific for soft-metal chelators. Although both elution systems yielded comparable analytical performance for each PC(n), the acetonitrile system had a lower sensitivity for GSH and a steadily increasing baseline. The inferior properties of the acetonitrile system may be due to complex formation between acetonitrile and Cu(I) ions. Both methods were applied to measure peptide levels in the primitive red alga Cyanidioschyzon merolae. Coefficients of variation (CVs) were less than 5%, except for GSH and PC(4) determinations in the acetonitrile system, in cases when CV values were found to be 8.8% and 6.3%, respectively. Recoveries were greater than 96%, except for GSH determination in the acetonitrile system, with a recovery of 84.4%; however, the concentration measured in the acetonitrile system did not differ from that measured in the methanol system at a significance level of 0.05.
采用乙腈和甲醇的 HPLC 洗脱系统用于定量测定谷胱甘肽 (GSH) 和植物螯合肽 (PC(n))，后者是一类在高等植物中参与重金属解毒的肽。检测系统基于铜 (I)-二硫代二苯并喹啉的去猝灭，并且对软金属螯合剂具有特异性。尽管这两种洗脱系统对于每个 PC(n) 的分析性能都相当，但乙腈系统对于 GSH 的灵敏度较低，并且基线不断增加。乙腈系统的较差性能可能是由于乙腈与 Cu(I) 离子之间形成了配合物。这两种方法都被应用于测量原始红藻 Cyanidioschyzon merolae 中的肽水平。除了在乙腈系统中测定 GSH 和 PC(4) 的情况外，变异系数 (CV) 小于 5%，在这两种情况下，CV 值分别为 8.8%和 6.3%。除了在乙腈系统中测定 GSH 的回收率大于 96%，回收率为 84.4%；然而，在乙腈系统中测量的浓度与在甲醇系统中测量的浓度在 0.05 的显著水平上没有差异。
