SIMULTANEOUS DETERMINATION OF ENDOGENOUS RETINOIC ACID ISOMERS AND RETINOL IN HUMAN PLASMA BY ISOCRATIC NORMAL-PHASE HPLC WITH ULTRAVIOLET DETECTION

We describe a rapid and simple procedure for the simultaneous quantita tion of endogenous 13-cis-retinoic acid, all-trans-retinoic acid, and retinol by isocratic normal-phase HPLC with ultraviolet detection, in 0.5 mL of human plasma. A silica adsorption column was eluted with n-h exane:2-propanol:acetic acid (200:0.7:0.135 by vol) at 0.9 mL/min, and the effluent monitored at 350 nm. The arotinoid ethylsulfonic acid Ro 15-1570 was used as the internal standard. High sensitivity, allowing quantitation of physiological concentrations, was achieved, particula rly for the retinoic acid isomers. The detection limits were 0.5 mu g/ L in plasma for both 13-cis and trans-retinoic acid, and 10 mu g/L for retinol. The CVs for between-day determinations of the lowest quality -control concentration (n = 12) were 4.8% for 13-cis-retinoic acid, 3. 4% for trans-retinoic acid, and 3.0% for retinol. The mean (+/- SD) co ncentrations of 13-cis-retinoic acid (1.79 +/- 0.56 mu g/L), trans-ret inoic acid (1.35 +/- 0.42 mu g/L), and retinol (533 +/- 58 mu g/L) mea sured in plasma from 22 healthy volunteers agreed well with those prev iously reported.