Factors influencing the protein binding of YH-439 using an equilibrium dialysis technique. A new hepatoprotective agent.
Various factors influencing the plasma protein binding of YH-439 to 4% human serum albumin (HSA) were evaluated using the equilibrium dialysis method at the initial YH-439 concentration of 2 micrograms mL-1. It took approximately 12 h of incubation to reach an equilibrium between 4% HSA and isotonic phosphate buffer of pH 7.4 containing 3% of dextran ('the buffer') using a Spectra/Por 2 membrane (molecular weight cut-off, 12,000-14,000) in a water bath shaker kept at 37 degrees C and at a rate of 50 oscillations min-1. YH-439 was fairly stable both in 4% HSA and in the 'buffer' for up to 24 h incubation. The binding of YH-439 to 4% HSA was constant (97.4 +/- 0.55%) at YH-439 concentrations ranging from 0.5 to 10 micrograms mL-1. However, the extent of binding was dependent on HSA concentrations: the values were 90.7, 94.7, 96.7, 97.0, 97.0, 97.1, and 97.5% at HSA concentrations of 0.5, 1, 2, 3, 4, 5, and 6%, respectively. The plasma protein binding decreased with increasing incubation temperature: the binding values were 98.2, 97.6, 97.2, and 96.8% when incubated at 10, 21, 26, and 37 degrees C, respectively. The binding of YH-439 was also influenced by the chloride concentration in the buffer: the binding values were 94.5, 97.0, and 96.8% for the chloride concentrations of 0, 0.249, and 0.546%, respectively. The binding of YH-439 was also dependent on the buffer pH: the percentages of free fraction were 6.0, 4.1, 3.8, 2.8, 2.7 and 2.8% for the buffer pHs of 5.0, 6.0, 6.5, 7.0, 7.4, and 8.0, respectively. The free fraction of YH-439 was slightly increased by the addition of heparin (up to 40 U mL-1), sodium azide (NaN3, up to 0.5%), and its metabolites. The protein binding of YH-439 was influenced neither by AAG, acetylsalicylic acid, or sulphisoxazole, nor by the addition of citrate or EDTA. The free fractions of YH-439 in rabbit (4.2%) and dog (4.7%) plasma seemed to be higher than in rats (2.9%) and humans (3.1%).
Lee
W I
WI
College of Pharmacy, Seoul National University, Korea.
Yoon
W H
WH
Park
J H
JH
Lee
J W
JW
Shim
C K
CK
Lee
M G
MG
D000818:Animals; D000925:Anticoagulants / Q000494:pharmacology; D001386:Azides / Q000494:pharmacology; D002021:Buffers; D002851:Chromatography, High Pressure Liquid; D003956:Dialysis; D004285:Dogs; D004492:Edetic Acid / Q000737:chemistry; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents / Q000494:pharmacology; D008567:Membranes, Artificial; D008970:Molecular Weight; D009961:Orosomucoid / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D011817:Rabbits; D051381:Rats; D012709:Serum Albumin / Q000378:metabolism*; D019810:Sodium Azide; D013045:Species Specificity; D013844:Thiazoles / Q000378:metabolism*
D000925:Anticoagulants; D001386:Azides; D002021:Buffers; D007202:Indicators and Reagents; D008567:Membranes, Artificial; D009961:Orosomucoid; D012709:Serum Albumin; D013844:Thiazoles; C098009:YH 439; D006493:Heparin; D019810:Sodium Azide; D004492:Edetic Acid