Improved HPLC assay for lipid peroxides in human plasma using the internal standard of hydroperoxide |
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Authors: | Email author" target="_blank">Shu-Ping?HuiEmail author Tsuyoshi?Murai Teruki?Yoshimura Hitoshi?Chiba Hironori?Nagasaka Takao?Kurosawa |
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Affiliation: | (1) Department of Health Sciences, Hokkaido University School of Medicine, 060-081 Sapporo, Japan;(2) Division of Metabolism, Chiba Children’s Hospital, 266-0007 Chiba, Japan;(3) Faculty of Pharmaceutical Sciences, Health Sciences University of Hokkaido, Ishikari-Tobetsu, 061-0293 Hokkaido, Japan |
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Abstract: | We have developed a sensitive reversed-phase chemiluminescence HPLC approach for simultaneous quantitative and qualitative
analyses of hydroperoxides of cholesteryl ester and TC in human plasma. Standard hydroperoxides of cholesteryl ester and TG
and a novel internal standard (1-tetradecanyl 3-octadecenoyloxy-5β-cholan-24-oate monohydroperoxide) (I.S.) were chemically
synthesized and the standard curves confirmed to be linear throughout the calibration range (1–1000 pmol). Within-day and
between-day CV were less than 7%, and the recoveries were within the range of 84–93%. With sample size minimized to 0.1 mL
of plasma for each run, plasma cholesteryl ester hydroperoxide levels were 189±87 nM (mean±SD) in healthy young (22–25 yr
old; n=15, male/female=6∶9) and 210 ±69 nM in healthy elderly (39–60 yr old; n=6, male/female= 3∶3). TG hydroperoxide was not detected in healthy subjects. In patients with advanced liver failure (36–67
yr old; n=4, male/female=2∶2), hydroperoxide levels of plasma cholesteryl ester and TG were 11,903±9,553 nM and 3,318±1,590 nM, respectively,
indicating an involvement of lipid oxidation. Sensitive and specific monitoring of plasma lipid peroxides using the present
chemiluminescence HPLC approach with the synthesized I.S. may help our understanding of chemical and pathophysiological aspects
of lipid peroxidation. |
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