Comparison of methanol and acetonitrile eluents for the quantitation of chelators specific to soft-metal ions by HPLC

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Analytical technologies in the biomedical and life sciences, 2012-11, Vol.909, p.34-36
• Main Authors: Ogawa, Shinya, Yoshimura, Etsuro
• Format: Article
• Language: English
• Subjects: acetonitrile; Acetonitriles - chemistry; Aminoacyltransferases; Bathocuproine disulfonate; chelating agents; Chelating Agents - analysis; Chromatography, High Pressure Liquid - methods; copper; Copper - chemistry; Dequenching; Glutathione; Glutathione - analysis; heavy metals; high performance liquid chromatography; ions; methanol; Methanol - chemistry; Phenanthrolines - chemistry; Phytochelatin; Phytochelatins - chemistry; Plant Extracts - chemistry; Reproducibility of Results; Rhodophyta; Rhodophyta - chemistry
• ISSN: 1570-0232; 1873-376X
• DOI: 10.1016/j.jchromb.2012.10.005

► HPLC detection system was established to quantify chelators specific to soft-metal ions. ► The system employed dequenching of Cu(I)–BCS complexes as a detection system. ► GSH and PC2–PC4 were successfully determined. ► Methanol was superior over acetonitrile as an eluent. HPLC eluent systems employing acetonitrile and methanol were evaluated for the quantitation of glutathione (GSH) and phytochelatin (PCn), 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 PCn, 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 PC4 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.