Measurement of the turnover of glycogen phosphorylase by GC/MS using stable isotope derivatives of pyridoxine (vitamin B-6)

The majority of vitamin B-6 in the body is in skeletal muscle, bound as the cofactor pyridoxal 5'-phosphate to one abundant protein, glycogen phosphorylase. Previous work has established that radiolabelled vitamin B-6 can be used as a turnover label for glycogen phosphorylase. In this study, a stable isotope derivative of pyridoxine (dideuterated pyridoxine; 3-hydroxy-4-(hydroxymethyl)-5-[hydroxymethyl-H-2(2)]-2-methylpyridine) ([H-2(2)]PN) has been used as a metabolic tracer to study the kinetics of labelling of the body pools of vitamin B-6 in mice. A non-invasive method was developed in which the isotope abundance of the urinary excretory product of vitamin B-6 metabolism, 4-pyridoxic acid, was analysed by GC/MS. The change in isotope abundance of urinary 4-pyridoxic acid following administration of [H-2(2)]PN reflects the kinetics of labelling of the body pools of vitamin B-6, and yields, non-invasively, the rate of degradation of glycogen phosphorylase.