Circadian changes of d-alanine and related compounds in rats and the effect of restricted feeding on their amounts

The circadian changes of d-alanine (d-Ala), an intrinsic d-amino acid found in mammals, were investigated in rats with diurnal and nocturnal habits, and the profiles were compared to those of l-Ala, other d-amino acids and several hormones. Determination of d-Ala in the rat plasma, pancreas and anterior pituitary gland was carried out using a sensitive and selective two-dimensional HPLC system combining a micro-ODS column and an enantioselective column after fluorescence derivatization with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F). The amount of d-Ala was high during the sleeping period and low during the active period in rats with both diurnal and nocturnal habits, indicating for the first time that the d-Ala is closely related to the activity rhythm of animals. In contrast, l-Ala and other d-amino acids did not show any clear circadian changes. The circadian change of d-Ala inversely correlated with that of the plasma insulin level in rats with both diurnal and nocturnal habits. Considered together with our previous findings that d-Ala is localized in the insulin secreting beta-cells in the rat pancreas, it is strongly suggested that d-Ala has some functional relationships to insulin in mammals.