The stable o‐phthalaldehyde‐ferrocene/6‐ferrocenyl‐1‐hexanethiol pre‐column derivatization high‐performance liquid chromatography of dimethylarginine by novel dual fluorescence and electrochemical detector |
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Authors: | Katsuhiro Suzuki Masatoki Katayama Kiyoshi Takamatsu Satoru Kaneko Hiromichi Ishikawa Takahiro Tsukimura Tadayasu Togawa |
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Institution: | 1. Drug Metabolism Research Laboratories, Drug Discovery Research, Astellas Pharma Inc., , Osaka, 532‐8514 Japan;2. Department of Functional Bioanalysis, Meiji Pharmaceutical University, , Tokyo, 204‐8588 Japan;3. Department of Obstetrics and Gynecology, Ichikawa General Hospital, Tokyo Dental College, , Ichikawa, Chiba, 272‐0824 Japan;4. Department of Urology, School of Medicine, Ichikawa General Hospital, Tokyo Dental College, , Ichikawa, Chiba, 272‐0824 Japan |
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Abstract: | Monomethylarginine, asymmetric dimethylarginine and symmetric dimethylarginine were separated on a Wakopak Combi ODS with an acetonitrile–100 mm potassium phosphate buffer (pH 7.0; 1:1, v/v). Dimethylarginines were derived from o‐phthalaldehyde for the fluorescence detector and from 6‐ferrocenyl‐1‐hexanethiol for the electrochemical detector. The detection limits of the dimethylarginines in spiked plasma were 0.3–0.5 pmol by electrochemical detection and 1–2 pmol by fluorescence detection. The detection limits were improved over 30 times by electrochemical detection and 10 times by fluorescence detection compared with previous reports. In previous derivatization liquid chromatography, the reaction solutions, o‐phthalaldehyde, 2‐mercaptethanol and dimethylarginines were unstable and required quick derivatization at 4°C. By our proposed pre‐column methods, the dimethylarginines were derivatized at room temperature and the fluorescent products were stable for 6 h. The manipulation performance was greatly advanced compared with previous LC reports. This is the first report on stable and sensitive dimethylarginines by dual detection. The selectivity was also improved by dual detection. The proposed method was applied to preliminary monitoring of dimethylargines in plasma and urine. Copyright © 2012 John Wiley & Sons, Ltd. |
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Keywords: | dual detection fluorescence detection pre‐column derivatization HPLC electrochemical detection asymmetric dimethylarginine |
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