A simple and sensitive radioenzymatic assay for pyridoxal 5′ - Phosphate in human plasma

Pyridoxal 5′-phosphate (PLP) is the biologically active form of vitamin B6. Low concentrations of plasma PLP have been reported to be associated with hyperhomocysteinemia - an independent risk factor for cardiovascular disease. A simple, rapid and inexpensive radioenzymatic assay for PLP has been developed on the basis of the principle that it is a coenzyme for tyrosine decarboxylase apoenzyme, therefore, the activity of the apoenzyme to convert [³H]-tyrosine to [³H]-tyramine would be dependent upon the concentration of PLP. The assay system has a lower limit of detection of 0.16 nmol/l and is linear upto 5 nmol/l. The recovery of PLP was 92% and the mean values of intra-assay and inter-assay coefficient of variation were found to be 8.3% and 12.9%, respectively. The procedure developed neither requires extraction of PLP from plasma nor deproteinization. As little as 5μl of plasma can be used in the reaction with reproducible results. Structurally similar vitamin B6-related compounds, such as pyridoxal, pyridoxamine and pyridoxine had no interference in the assay. The assay developed was used to monitor the PLP status of a small population of patients with acute myocardial infarction and normal healthy subjects.