A simple liquid-chromatographic method for measuring vitamin B6 compounds in plasma.

This relatively simple high-performance liquid chromatographic (HPLC) method for measuring all seven known forms of vitamin B6 in plasma from individuals supplemented with pyridoxine hydrochloride shows good analytical recovery (85-98%) and precision. Within-run and between-run CVs for plasmas supplemented with standards were 4% and 7%, respectively. The major forms of B6 found in unsupplemented plasma from normal subjects were pyridoxal phosphate and 4-pyridoxic acid, with pyridoxal just detectable. The HPLC procedure correlated well (r = 0.94) with a modification of an enzymatic method involving apotryptophanase (Anal Biochem 1972;45:567-76) for measuring plasma pyridoxal phosphate, and also (r = 0.94) with a routine method for determining 4-pyridoxic acid in urine (Clin Chem 1964;10:479-89). Elimination of pyridoxine from the plasma of both normal and hyperoxaluric individuals was shown to be very rapid, with half-lives (t1/2) of 45 and 40 min, respectively. Finally, we present evidence for the existence of two other forms of B6 and discuss the possibility of a new metabolic pathway in vitamin B6 metabolism.