The pharmacokinetics and tissue distribution of methotrexate (MTX) were investigated after intravenous (i.v.) and intramuscular (i.m.) injection of free MTX (treatment I), MTX-bearing neutral liposomes (large unilamellar vesicles), OLUVs (treatment II) and free MTX mixed manually with empty OLUVs (treatment III), 4mg/kg as free MTX to rats. After i.v. infusion in 1 min, the plasma concentrations of MTX (C-p), area under the plasma concentration-time curve (AUC, 173 vs. 1110 mu g ml/min), terminal half-life (t(1/2), 24.0 min vs. not determined), mean residence time (MRT, 13.0 vs. 165 min) and apparent volume of distribution at steady state (V-ss, 289 vs. 584 ml/kg) increased significantly; however, total body clearance (Cl, 23.1 vs. 3.61 ml/min/kg), renal clearance (Cl-R, 8.38 vs. 1.88 ml/min/kg) and nonrenal clearance (Cl-NR, 14 6 vs. 1 66 ml/min/kg) decreased significantly from treatment II when compared with the values from treatment I. This could be due to the fact that some of the MTX-bearing OLUVs were entrapped in tissues and the others were present in plasma (increase in MRT and V-SS from treatment II). MTX was slowly released from the MTX-bearing OLUVs (increase in t(1/2) from treatment II), With the present HPLC assay, the concentrations of MTX represent the sum of the free MTX and MTX in MTX-bearing OLUVs (increase in C-p and AUC, and decrease in Cl from treatment II). Some pharmacokinetic parameters of MTX, such as t(1/2) (24.0 vs. 58.2 min), MRT (13.0 vs. 23.3 min) and V-SS (289 vs. 456 ml/kg) were significantly different after i.v. administration of empty OLUVs (between treatments I and III); however, the differences seemed to be smaller than those between treatments I and II. After i.m. administration, t(1/2) (37.2 min vs. not determined) and the total amounts of MTX excreted in urine (X(u), 319 vs. 171 mu g) were significantly different after treatments I and II. After both i.v. and i.m. administration, the amount of MTX remaining per gram of tissue, and/or tissue to plasma ratio (T/P) of MTX were significantly reduced in the kidney, small intestine, large intestine or stomach from treatment II when compared with those from treatment I. This implies that the side-effects of MTX on the kidney and gastrointestinal tract could be reduced after i.v. or i.m. administration of MTX-bearing OLUVs rather than free MTX. The mean encapsulation efficiency of MTX in MTX-bearing OLUVs was 3.88% and the MTX was released slowly from MTX-bearing OLUVs when incubated in phosphate-buffered saline, rat plasma and rat liver homogenate.
