An original HPLC assay was developed for pyrimethamine (PYR) in plasma, RBCs, and buffer for the purpose of studying its plasma protein binding and RBC partitioning.
Equilibrium dialysis (ED) was used to study protein binding. Isotonic phosphate buffer used in ED did not prevent small volume shifts. The pH of the plasma affected the protein binding of PYR although it was not significant for the comparison of binding at pH 7.4 vs. 8.0. PYR at 1000 ng/ml averaged 93.1% bound to plasma proteins. Binding to pure human albumin was 86.5% at lower levels of albumin and PYR (350 ng/ml). There was a significant difference (p<.03) in the plasma binding at two levels in the therapeutic range, with more free at higher levels. There was also concentration dependent binding at higher concentrations; the drug did not follow to law of mass action when binding increased at higher concentrations. This is a solubility phenomenon. Linear regression at the effect at albumin concentration on plasma binding yielded the equation percent free = -0.467(albumin g/L) + 23.5. The binding to pure albumin was only slightly above that predicted by this equation (83.1%). The first and second stoichiometric binding constants are K1 = 2.83 x 104 and K2 = 1.74 x 104 M-1 from nonlinear regression at data. There was no binding to normal levels at α1-acid glycoprotein.
PYR is preferentially bound to plasma proteins in comparison to RBCss. The mean RBC/plasma ratio was 0.42 (10.2% CV, n=5). when plasma was removed and pH 7.4 isotonic buffer substituted, mean RBC/buffer ratio was 5.2 (11.8% CV, n=2). Mean percent bound to hemolysate was 42.5% (19% CV, n=10). Binding to hemoglobin did not account for all the RBC uptake. Therefore, PYR binds to RBC membranes.
| Identifer | oai:union.ndltd.org:vcu.edu/oai:scholarscompass.vcu.edu:etd-6073 |
| Date | 01 January 1987 |
| Creators | Rudy, Anita C. |
| Publisher | VCU Scholars Compass |
| Source Sets | Virginia Commonwealth University |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Theses and Dissertations |
| Rights | © The Author |
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