Maia M B, Saivin S, Chatelut E, Malmary M F, Houin G
Laboratoire Cinétique des Xénobiotiques, Faculté des Sciences Pharmaceutiques, Hôpital Rangueil, Toulouse, France.
Int J Clin Pharmacol Ther. 1996 Aug;34(8):335-41.
The purpose of the study was firstly to estimate methotrexate protein binding using microdialysis, and secondly to determine the influence of the protein content in the dialyzed medium on the dialysis membrane recovery. In vitro recovery was estimated by both water recovery method and concentration difference method. The relative recovery was independent of methotrexate concentration: 39.3% +/- 2.86% and 39.2 +/- 1.27% for 50 microM and 300 microM, respectively. A significant influence of the presence of proteins on the dialysis membrane recovery was observed: 55.9% +/- 2.7%, 42.3% +/- 7.5% and 45.5% +/- 0.1% for buffer, human serum albumin (HSA) (600 microM) and human plasma, respectively. Methotrexate binding to human and rat plasma proteins showed a nonsaturable phenomenon. The bound percentages and corresponding total binding capacities were 58.7% +/- 3.13% and 1.44 +/- 0.033 mM-1 and 71.7% +/- 4.38% and 2.18 +/- 0.09 mM-1 for the human and rat plasma, respectively. For HSA this binding was saturable with an affinity constant of 4.75 +/- 0.66 mM-1. After intravenous administration (250 mg/kg) the in vivo rat plasma binding of methotrexate was roughly 20% higher (93.6%) than the in vitro methotrexate plasma binding. The in vivo relative recovery of methotrexate was found to be 12.6% +/- 1.8% versus 25.4% +/- 3.3% in vitro. This study showed that the protein content can directly affect microdialysis probe recovery. However, by taking into account this recovery, microdialysis allows to measure the protein binding of methotrexate.
本研究的目的，一是使用微透析法估算甲氨蝶呤的蛋白结合率，二是确定透析液中蛋白质含量对透析膜回收率的影响。采用水回收率法和浓度差法估算体外回收率。相对回收率与甲氨蝶呤浓度无关：50微摩尔/升和300微摩尔/升时分别为39.3%±2.86%和39.2±1.27%。观察到蛋白质的存在对透析膜回收率有显著影响：缓冲液、人血清白蛋白（HSA）（600微摩尔/升）和人血浆的回收率分别为55.9%±2.7%、42.3%±7.5%和45.5%±0.1%。甲氨蝶呤与人及大鼠血浆蛋白的结合呈现非饱和现象。人血浆和大鼠血浆的结合百分比及相应的总结合容量分别为58.7%±3.13%和1.44±0.033毫摩尔⁻¹以及71.7%±4.38%和2.18±0.09毫摩尔⁻¹。对于HSA，这种结合是可饱和的，亲和常数为4.75±0.66毫摩尔⁻¹。静脉注射（250毫克/千克）后，甲氨蝶呤在大鼠体内的血浆结合率比体外甲氨蝶呤血浆结合率约高20%（93.6%）。甲氨蝶呤的体内相对回收率为12.6%±1.8%，而体外为25.4%±3.3%。本研究表明，蛋白质含量可直接影响微透析探针的回收率。然而，考虑到这种回收率后，微透析能够测量甲氨蝶呤的蛋白结合率。
