Teruhisa Ichihashi

Effect of Oily Vehicles on Absorption of Mepitiostane by the Lymphatic System in Rats

Abstract— [14C]Mepitiostane in various vehicles was administered to the small intestine of anaesthetized rats with cannulated thoracic ducts, and the effect of lipids on lymphatic absorption was examined. The extent of lymphatic absorption was greatest when administered in triolein and sesame oil, which are triglycerides of long‐chain fatty acids. Absorption in the presence of other vehicles was in the order of 10% Tween 80 aqueous solution > monolein > oleic acid ∼ oleic acid/monolein (2:1 mol/mol) > aqueous suspension. Differences between the extents of lymphatic absorption of mepitiostane in the various formulations were not due to variation in the lymph flow but to the increased secretion of chylomicron and very low density lipoproteins. During absorption of mepitiostane from the small intestine, oil affected not only the penetration into epithelium cells and the metabolism in them, but also the partition between blood and lymph.

Effect of Bile on Absorption of Mepitiostane by the Lymphatic System in Rats

Abstract— The effects of bile and site of gastrointestinal absorption on the lymphatic absorption of the highly lipophilic drug, mepitiostane were examined using thoracic duct‐cannulated rats. The lymphatic absorption from the small intestine was very small in the absence of bile compared with that when bile was present. The lymphatic absorption was greatest when drug was administered to the upper small intestine with bile, was smaller for the lower regions of the small intestine, and was negligible for the stomach and the large intestine. A correlation was observed between the extent of lymphatic absorption and the secretion of chylomicron and very low density lipoproteins after administration to various regions with or without bile. The portal absorption data of mepitiostane confirmed that site specificity occurs in the partition of drug between blood and lymph.

Intrinsic lymphatic partition rate of mepitiostane, epitiostanol, and oleic acid absorbed from rat intestine.

Mepitiostane (MP), epitiostanol (EP), and oleic acid were administered to the jejunal loop of mesenteric vein- and thoracic duct-cannulated rats, and the intrinsic lymphatic partition rate (ILPR) of the absorbed compounds was directly determined. When 14C-EP was administered to the jejunal loop, recovery of unchanged EP in the mesenteric blood and the lymph was 7.9 and 0.03% of the administered dose, respectively. In contrast, following administration of 14C-MP, recovery of unchanged MP in the mesenteric blood and the lymph was 1.2 and 15.0%, respectively. Thus, following passage through the mucosal cell, 99.6% of the unchanged EP was partitioned into the blood and 0.4% into the lymph, while for unchanged MP, 7.6% was partitioned into the blood and 92.4% into the lymph. When 14C-oleic acid was administered to the jejunal loop, most of the penetrating oleic acid was incorporated into triglycerides in epithelial cells and transferred exclusively into the lymph. However, of the unchanged oleic acid, only 37.6% was partitioned into the lymph and 62.4% into the blood. The ILPR was 92.4% for MP, 0.4% for EP, and 37.6% for oleic acid. We conclude that the ILPR values indicate the true lymphotropic property of the compounds.

A quantitative concept of the mechanism of intestinal lymphatic transfer of lipophilic molecules.

The partition of mepitiostane, testosterone, and some structurally related compounds between lymph and blood in rat jejunum (lymph–blood partition ratio; LBPR) was determined, and the quantitative relationship between LBPR and lipophilicity was examined. When the ΔRm values (hydrophobic parameter derived from the mobility) relative to testosterone were <0.2, their logLBPRs remained approximately constant in the range of −2 to −3. When the ΔRm values of the compounds were >0.2, a linear correlation (r = 0.986, n = 8) was observed between these values and the logLBPRs. The LBPR, but not the extent of lymphatic absorption, of lipophilic molecules was determined strictly by the superlipophilicity, and for high partitioning into the lymph (>50% of the absorbed amount), the ΔRm value had to be >0.50 (5.65 as the logP value). The relationship between LBPR and superlipophilicity could be explained on the basis of the theoretical equations derived from absorption kinetics based on a dynamic partitioning model.

Factors Determining the Intrinsic Lymphatic Partition Rate of Epitiostanol and Mepitiostane

Substitution of the steroid epitiostanol (EP) at position 17 with methoxycyclopentane yields the extremely lipophilic mepitiostane (MP) with preferential partitioning into the lymph. Most of the MP in the lymph was associated with the core lipids of chylomicrons and very low-density lipoproteins (VLDL), as was also the case for EP. However, the dialysis velocity of EP and MP from lymph to plasma differed greatly; EP, but not MP, was transferred from the lymph to the plasma. This difference was attributed to differences in their unbound fraction in the lymph. Lymphatic transfer and the logP value of several tested steroids correlated well. Therefore, the oral EP prodrug, MP, partitioned into the lymph because of its superlipophilicity and resultant retention in the core lipids of chylomicrons and VLDL.

EPIMERIZATION OF MOXALACTAM BY ALBUMIN AND SIMULATION OF IN VIVO EPIMERIZATION BY A PHYSIOLOGICALLY BASED PHARMACOKINETIC MODEL

We investigated the mechanism of epimerization (R to S or S to R) of moxalactam in serum of rats, dogs, and humans. The epimerization of moxalactam occurred in the serum of these animals, but not in the serum filtrate. The albumin fraction of human serum purified by gel filtration catalysed the epimerization of moxalactam at an identical rate to serum, but other fractions (i.e., lipoproteins and globulins) showed slower epimerization. alpha 1-acid glycoprotein, which was eluted in the same fraction with albumin by G-200 gel filtration, did not epimerize moxalactam. The presence of 2 mM warfarin decreased the binding of R- and S-moxalactam and decreased the epimerization of moxalactam in human serum. These results demonstrate moxalactam was epimerized on the warfarin binding site on albumin in serum. Additionally, a physiologically based pharmacokinetic model shows that the epimerization of moxalactam after administration in dogs is simulated by the epimerization in serum.

Epimerization Kinetics of Moxalactam in Frozen Urine and Plasma Samples

The epimerization of moxalactam (LMOX) in frozen urine and plasma samples was studied during long-term storage. The R/S ratio at equilibrium [(R/S)eq] at −10°C was similar in urine and in rat and human plasma ultrafiltrate but differed from that in water. The (R/S)eq values in human plasma and its ultrafiltrate differed slightly, while they were the same in rat plasma and in its ultrafiltrate. The difference for the human plasma and ultrafiltrate may result from differences in plasma protein binding between R- and S-epimers in the liquid region of the frozen plasma. The change of R/S ratio in frozen human plasma continued below the collapse temperature of LMOX aqueous solution, where the liquid region appeared still to exist as determined by NMR measurement. Consequently, the biological LMOX samples should be preserved at or below −70°C to prevent changes in the R/S ratio.

Epimerization kinetics of moxalactam in frozen solution.

The epimerization rate constants of R- and S-epimers of moxalactam (LMOX) in a frozen aqueous solution decreased as the temperature decreased. The reaction proceeded in the unfrozen region remaining in the frozen solution, without being affected by the ice. The reaction stopped completely below the collapse temperature of the LMOX aqueous solution. The ratio of R- and S-epimers at equilibrium, which was equal to the ratio of the epimerization rate constant, increased as the temperature decreased. This change in the ratio at equilibrium could be ascribed to the difference in the activation energy between the two epimers.