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Simultaneous Determination of All-trans-and 13-cis-Retinoic Acid in Human Serum by HPLC

Yu Zicheng Yao Shuai Yu Renhai
(Institute of Clinical Pharmacology, Ruijin Hospital, Shanghai Second Medical University, Shanghai 200025)


Objective: To develop an isocratic HPLC for the simultaneous determination of all-trans-and 13-cis-retinoic acid (ATRA,13-cRA) in human serum.

Methods: The serum sample (1 mL) was extracted with 5 mL of ether for twice. The extract was analyzed on an μBondapak C18 (3.9 mm×300 mm) column with a mobile phase consisting of 85 Vols. of methanol and 15 Vols. of NH4Ac buffer (pH 6.0) at 22 .The flow rate was 0.8 mL.min-1. The retinoids were detected at 340 nm and quantitated by internal standard method, DMAB was used as an internal standard.

Results: The linearity of method was in the range of 0.8 to 1 120 μg.L-1 for ATRA and 0.82 to 1 312 μg.L-1 for 13-cRA.The limit of determination was 0.6 μg.L-1 in serum for both ATRA and 13-cRA.Average recoveries were 98.86%~105.2% for ATRA,101.6%~101.8% for 13-cRA. Within-day and between-day precision of determination were 0.84%~5.5% for ATRA and 1.4%~5.7% for 13-cRA.

Conclusion: This method is sensitive, accurate and good enough to be utilized for the investigation of the clinical pharmacokinetics of ATRA.
Key words: all-trans-retinoic acid, 13-cis-retinoic acid, serum drug concentration, HPLC

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