Characterization of pyridoxal 5'-phosphate disappearance from in vivo perfused segments of rat jejunum.
The disappearance of pyridoxal 5'-phosphate (PLP) from the lumen of in vivo perfused segments of rat jejunum was evaluated utilizing a single-pass technique. Net water flux was monitored by [14C] dextran, a nonabsorbable volume marker. Unlabeled PLP was measured by the tyrosine apodecarboxylase assay. PLP disappearance was linear with respect to PLP concentration at concentrations below 300 micrometers but was saturable at high concentrations (3 micrometers). PLP disappearance was significantly inhibited by 1 micrometer pyridoxamine 5'-phosphate and 5 micrometers and 10 micrometers l-phenylalanine but not by 1 micrometer pyridoxamine. Both in vivo disappearance (during perfusion) and in vitro PLP decay (exiting perfusate used as medium) correlated with the measured alkaline phosphatase activity of exiting perfusate under low-phosphate (1.1 micrometer) conditions. In contrast, in vivo PLP disappearance was not correlated with perfusate alkaline phosphatase activity under high-phosphate (80 micrometer) conditions. When exiting perfusate was ultracentrifuged at 105,000 x g for 1 hour, only 35% of the initial alkaline phosphatase activity remained in the supernatant. Conclusions were: 1) PLP disappearance from the lumen of an in vivo perfused segment of rat jejunum is saturable and inhibited by l-phenylalanine; 2) PLP disappearance appears in part to be a function of intraluminal alkaline phosphatase; and 3) A major portion of the alkaline phosphatase activity measured in the exiting perfusate represents membrane-bound enzyme.
D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D004305:Dose-Response Relationship, Drug; D007583:Jejunum / Q000378:metabolism*; D008297:Male; D009943:Organophosphorus Compounds / Q000494:pharmacology; D010477:Perfusion; D011732:Pyridoxal Phosphate / Q000378:metabolism*; D011733:Pyridoxamine / Q000031:analogs & derivatives / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D014461:Ultracentrifugation
D009943:Organophosphorus Compounds; D011732:Pyridoxal Phosphate; D011733:Pyridoxamine; D000469:Alkaline Phosphatase; C010627:pyridoxamine phosphate