Studies of the reversible binding of biotin to human plasma.
In this study we sought to determine the extent of the reversible binding of [3H]biotin to macromolecules (presumably proteins) of human plasma under physiologic conditions during physiologic time intervals ranging from minutes to days. Fresh, heparinized human plasma and [3H]biotin were incubated in 5% CO2, 95% N2 at 37 degrees C. After reaching equilibrium, free biotin was separated from bound biotin by centrifugal ultrafiltration (1500 x g for 60 min) using membranes with a molecular weight cutoff of 30,000. In a sample from a single individual, we detected a mean retentate:ultrafiltrate ratio of 1.20 +/- 0.03 (mean +/- SD, n = 14 determinations). In a group of 10 healthy adults, the mean retentate:ultrafiltrate ratio was 1.17 +/- 0.02; thus, approximately 8% of the biotin in the combined pool of free and reversibly bound biotin is bound. No consistent change in retentate:ultrafiltrate ratio was detected for total biotin concentrations ranging between 500 fmol/mL and 2 nmol/mL, but the ratio decreased to 1.1 at 400 nmol of biotin per mL. Thus, the biotin-binding system of human plasma appears to be a low affinity, high capacity system. A similar biotin-binding system was detected in experiments with purified human serum albumin. These results provide evidence that the majority of added [3H]biotin does not bind to plasma protein under physiologic conditions. To the extent that added [3H]biotin can be assumed to reflect reversible binding of endogenous unlabeled biotin, we infer that endogenous, reversibly bound biotin can account for no more than one-tenth of the increase in biotin detected after acid hydrolysis of plasma.
Mock
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DM
Department of Pediatrics, University of Iowa, Iowa City 52242.
Lankford
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D000328:Adult; D001665:Binding Sites / Q000187:drug effects; D001710:Biotin / Q000032:analysis* / Q000493:pharmacokinetics; D001798:Blood Proteins / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D005227:Fatty Acids / Q000494:pharmacology; D005260:Female; D006801:Humans; D008297:Male; D010949:Plasma / Q000032:analysis*; D014462:Ultrafiltration
D001798:Blood Proteins; D005227:Fatty Acids; D001710:Biotin