Studies on the establishment of the analytical method for, and the distribution and change of the total D-amino acids in mammalian body

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During the course of our studies on the determination of D-amino acids in mammals, we found that a large amount (mMorder) of D-aspartic acid (D-Asp) is present in the rat pineal gland. The method we usedisa chiral stationary phase-HPLC separation adn fluorometoric detection of NBD-labelled amino acids. In the present study, to increase the efficiency of the enantioneric separation, we developed 3 new chiral stationary phase containing the same chiral moieties as the previous phases with an extended long alkyl chain from the aminopropylsilica support. One of them, APS-11-OA-2500S was the best for the enantiomeric separation of NBD-amino acids, amino acids ester and beta-amino butyricacid. The interaction between the analyte and the chiral moiety was also studied with NMR,showing some interaction of proton of amino moiety of amino acids and carbonyl moiety of the chiral stationary phase.The whole body autoradiography of rat injected with ^<14>C labelled D-Asp showed the incorpration of radioactivity to the tissues where the endogenous D-Asp existed.The polyclonal antibody to D-Asp revealed the localization of the endogenous D-Asp in the cytoplasm of pinealocyte which locate in the distal region of the gland. The specifid location of D-Asp was also observed in the pituitary gland, suggesting the exsitence of some important physiological roles of D-Asp in the mammaliam body.