A B S T R A C T In studies with pyridoxine and other B6 compounds in blood, the active forms pyridoxal and pyridoxal phosphate were measured by differential assays using Lactobacillus casei. Red cell uptake of tritiated pyridoxine was also measured. A new metabolic pathway for conversion of pyridoxine to active forms was demonstrated in red cells.In vivo studies in normal subjects suggested that pyridoxine was taken up by red cells where it was converted to pyridoxal phosphate and then pyridoxal, followed by gradual release of a proportion of pyridoxal into plasma. In vitro incubation of pyridoxine with blood confirmed this observation.Increasing amounts of pyridoxine were taken up and converted as the amount added to blood was increased, and only very small numbers of red cells were needed to convert appreciable amounts. Conversion was markedly inhibited at temperatures lower than 370C, and stopped altogether at -20'C.Release of pyridoxal into plasma was always directly proportional to the amount of pyridoxal formed and to the volume of plasma present. That pyridoxal phosphate was not released into plasma was demonstrated in stored blood, for pyridoxine was converted mainly only as far as pyridoxal phosphate, probably due to inactivation of the phosphatase. Pyridoxal phosphate remained in the red cells.Pyridoxine was converted when incubated with washed red cells in saline or phosphate buffer suspension (0.08 M). In saline suspension, pyridoxal formed but was not released in the absence of plasma. In phosphate buffer suspension, pyridoxal phosphate was formed but was not changed to pyridoxal, probably due to inactivation of phosphatase by excess phosphate.This work was presented in part at the 8th International

SYNOPSIS A new method has been developed for the assay of serum pyridoxal using L. casei. Bound pyridoxal phosphate in serum is converted by acid hydrolysis to pyridoxal for which this organism is specific. This method proved to be considerably more sensitive than other methods so far reported in the literature.Serum pyridoxal concentrations were measured in 151 control subjects aged 17 to 80 years. The range of concentrations found was 1l5 to 13'5 ng/ml which compared well with values obtained by most workers measuring pyridoxal phosphate by enzymatic methods. A marked fall with age was confirmed, and levels in women of childbearing age were lower than in men of comparable age. Subnormal serum pyridoxal concentrations were found in 62 % of patients with sideroblastic anaemia and in the majority of patients with rheumatoid arthritis, Crohn's disease, coeliac disease, and in pregnant women at term.