Incorporation studies with 13C-, 15N-, and 2H-labeled substrates, followed by NMR analysis, show that the pyrimidine unit of thiamin (Vitamin B1) originates from a C5N fragment, derived from C-2', 2, N, C-6, 5, 5'of pyridoxol (Vitamin B6) and an NCN fragment derived from l-histidine. Urocanic acid serves as an intermediate on the route of the NCN fragment of histidine into the thiamin pyrimidine.