Preparation and determination of zinc(II) chlorophylls by reversed-phase high-performance liquid chromatography

• Published in: Journal of Chromatography A 1993-08, Vol.645 (2), p.259-264
• Main Authors: Inoue, Hidenari, Imai, Miki, Naemura, Takashi, Furuya, Kenji, Shizuri, Yoshikazu
• Format: Article
• Language: English
• Subjects: Analytical, structural and metabolic biochemistry; Biological and medical sciences; chlorophyll; determination; Fundamental and applied biological sciences. Psychology; Heterocyclic compounds, pigments; high performance liquid chromatography; Miscellaneous; Other biological molecules; Spinacia oleracea; zinc
• ISSN: 0021-9673
• DOI: 10.1016/0021-9673(93)83385-6

A preparative method for zinc(II) chlorophyll a and b [Zn(II)-chl-a and -b] was developed on the basis of their purification by semi-preparative high-performance liquid chromatography. Zn(II)-chl-a and -b and the corresponding epimers were separated and determined with an ODS (C 18 chemically bonded silica gel) column using methanol—acetone (75:25, v/v) as mobile phase. Linear calibration graphs were obtained over the concentration range 0–50 μg cm −3 of each zinc(II) chlorophyll with photometric detection at 425 nm. The present HPLC determination provides an accurate and conventional method with a detection limit of 3.5 ng cm −3 for Zn(II)-chl-a, 2.5 ng cm −3 for Zn(II)-chl-a′ and 3.0 ng cm −3 for Zn(II)-chl-b with a relative standard deviation of less than 2.3% ( n = 10). The analytical values obtained for synthetic samples by a spectrophotometric method were confirmed to be high compared with those determined by the proposed HPLC method.