Gastro-intestinal absorption of fatty acid esters of pyridoxine was studied in human by the oral administration of 243.3 μmoles. It was found that the urinary excretion of pyridoxine 5-octanoate and 3,4-dioctanoate in 48 houre was as large as pyridoxine, but those of pyridoxine 3,4-dilaurate and -dipalmitate were small. By the study of thin-layer chromatography, the main metabolite of the esters was 4-pyridoxic acid, i.e., the same metabolite of pyridoxine. From the present study, it was found that the esters were hydrolyzed to pyridoxine in the human body. There was a good correlationship between the intestinal absorption of the esters in human and their penetration through everted sac of the rat intestine. The differences of absorption and penetration between the esters were closely correlated with the differences of degree of hydrolysis by homogenates of the mouse intestine.