title: Development of a high-performance liquid chromatography-based assay for carotenoids in human red blood cells: application to clinical studies.
abstract: Peroxidized phospholipid-mediated cytotoxicity is involved in the pathophysiology of many diseases; for example, there is an abnormal increase of phospholipid hydroperoxides in red blood cells (RBCs) of dementia patients. Dietary carotenoids have gained attention as potent inhibitors of RBC phospholipid hydroperoxidation, thereby making them plausible candidates for preventing disease. However, the occurrence of carotenoids in human RBCs is still unclear. This is in contradistinction to plasma carotenoids, which have been investigated thoroughly for analytical methods as well as biological significance. In this study, we developed a method to analyze RBC carotenoids using high-performance liquid chromatography (HPLC) coupled with ultraviolet (UV) diode array detection (DAD) and atmospheric pressure chemical ionization (APCI) mass spectrometry (MS). Under optimized conditions that included extraction, separation, and detection procedures, six carotenoids (lutein, zeaxanthin, beta-cryptoxanthin, alpha-carotene, beta-carotene, and lycopene) were separated, detected by DAD, and concurrently identified based on APCI/MS and UV spectra profiles when an extract from human RBCs was subjected to HPLC-DAD-APCI/MS. The amounts of carotenoids varied markedly (1.3-70.2 nmol/L packed cells), and polar oxygenated carotenoids (xanthophylls) were predominant in RBCs. The HPLC-DAD-APCI/MS method would be a useful tool for clinical studies for evaluating the bioavailability of RBC carotenoids.
authors.affiliation: Food and Biodynamic Chemistry Laboratory, Graduate School of Agricultural Science, Tohoku University, Sendai 981-8555, Japan.
authors.forename: Kiyotaka
authors.initials: K
authors.lastname: Nakagawa
authors.forename: Takehiro
authors.initials: T
authors.lastname: Kiko
authors.forename: Keijiro
authors.initials: K
authors.lastname: Hatade
authors.forename: Akira
authors.initials: A
authors.lastname: Asai
authors.forename: Fumiko
authors.initials: F
authors.lastname: Kimura
authors.forename: Phumon
authors.initials: P
authors.lastname: Sookwong
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Tsuduki
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Arai
authors.forename: Teruo
authors.initials: T
authors.lastname: Miyazawa
mesh_terms: D001681:Biological Assay / Q000379:methods*; D002338:Carotenoids / Q000097:blood* / Q000737:chemistry / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004912:Erythrocytes / Q000737:chemistry* / Q000187:drug effects; D005260:Female; D006801:Humans; D006878:Hydroxides / Q000494:pharmacology; D008297:Male; D013058:Mass Spectrometry; D017680:Potassium Compounds / Q000494:pharmacology; D011748:Pyrogallol / Q000494:pharmacology; D012015:Reference Standards; D013056:Spectrophotometry, Ultraviolet
chemical_list: D006878:Hydroxides; D017680:Potassium Compounds; D011748:Pyrogallol; D002338:Carotenoids; C029943:potassium hydroxide