A reversed-phase HPLC method using acetonitrile-methanol-1 M
perchloric acid-water (30:9:1:95, v/v) at a flow-rate of 1.5 ml/min on a mu-Bondapak C18 column with UV detection at 254 nm was developed for the separation of
primaquine, its major metabolite
carboxyprimaquine and other metabolites such as N-acetylprimaquine, 4-hydroxyprimaquine,
5-hydroxyprimaquine, 5-hydroxy-6-methoxyprimaquine, demethylprimaquine and 6-methoxyprimaquine, and also other
antimalarials. The calibration graphs were linear in the range 0.025-100 micrograms/ml for
primaquine and 4-1000 micrograms/ml for
carboxyprimaquine. The within-day and day-to-day coefficients of variation averaged 3.65 and 6.95%, respectively, for
primaquine and 3.0 and 7.52%, respectively for
carboxyprimaquine in plasma. The extraction recoveries for
primaquine and
carboxyprimaquine were 89 and 83%, respectively. The mean
carboxyprimaquine concentration was much higher in plasma and blood cells of Plasmodium vivax patients than that in plasma from healthy subjects. The
carboxyprimaquine level was also higher in blood cells than plasma whereas the
primaquine concentration was the same in both cases.