Susceptibility to metabolism is a common issue with the tert-butyl group on compounds of medicinal interest. We demonstrate an approach of removing all the fully sp 3 C−Hs from a tert-butyl group: replacing some C−Hs with C−Fs and increasing the s-character of the remaining C−Hs. This approach gave a trifluoromethylcyclopropyl group, which increased metabolic stability. Trifluoromethylcyclopropyl-containing analogues had consistently higher metabolic stability in vitro and in vivo compared to their tert-butyl-containing counterparts.

We investigated the effects on LH secretion of infusing gamma-aminobutyric acid (GABA) agonists muscimol and baclofen (GABAA and GABAB receptor agonists, respectively) into either the medial preoptic area (mPOA) or the arcuate-ventromedial region (ARC-VMR) of the hypothalamus of castrated rams during the nonbreeding season. Bilateral microdialysis of artificial cerebrospinal fluid for 4 h followed by treatment with artificial cerebrospinal fluid, baclofen (1 mM), or muscimol (1 mM in the ARC-VMR, 250 microM in the mPOA) for 4 h was carried out on three separate occasions in random order. Simultaneously, jugular venous blood was collected at 10-min intervals, and plasma later was assayed for LH. The estimated dose of baclofen delivered to each unilateral microdialysis site was 7.9 micrograms; for muscimol, it was 1.1 micrograms for the mPOA and 4.5 micrograms for the ARC-VMR. In the mPOA, baclofen had no detectable effect, whereas muscimol had a delayed suppressive effect on mean LH (P < 0.01). In the ARC-VMR muscimol rapidly reduced mean LH (P < 0.01). In contrast, baclofen increased mean LH (P = 0.01) and LH pulse amplitude (P = 0.05) without altering the LH interpulse interval (P > 0.10). These results support the involvement of GABAA receptors in the mPOA in regulating LH secretory patterns. More importantly, both GABAA and GABA(B) receptors in the ARC-VMR appear to differentially modulate LH and, presumably, GnRH release. Whether GABA acts directly on the GnRH neuron or indirectly through other neural systems remains to be determined.

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ABSTRACT:1-[4-Aminomethyl-4-(3-chlorophenyl)-cyclohexyl]-tetrahydro-pyrimidin-2-one, 1, was developed as an inhibitor of dipeptidyl peptidase-4 enzyme. Biotransformation studies with 1 revealed the presence of an N-carbamoyl glucuronide metabolite (M1) in rat bile and urine. N-Carbamoyl glucuronides are rarely observed, and little is understood regarding the mechanism of N-carbamoyl glucuronidation. The objectives of the current investigation were to elucidate the structure of the novel N-carbamoyl glucuronide, to investigate the mechanism of N-carbamoyl glucuronide formation in vitro using stable labeled CO 2 , UDP glucuronosyltransferase (UGT) reaction phenotyping, and to assess whether M1 was formed to the same extent in vitro across species-mouse, rat, hamster, dog, monkey, and human. Structure elucidation was performed on a mass spectrometer with accurate mass measurement and MS n capabilities.13 C-labeled carbon dioxide was used for identification of the mechanism of N-carbamoyl glucuronidation. Mechanistic studies with 13 C-labeled CO 2 in rat liver microsomes revealed that CO 2 from the bicarbonate buffer (in equilibrium with exogenous CO 2 ) may be responsible for the formation of M1. M1 was formed in vitro in liver microsomes from multiple species, mainly rat and hamster, followed by similar formation in dog, monkey, mouse, and human. M1 could be detected in UGT1A1, UGT1A3, and UGT2B7 Supersomes in a CO 2 -rich environment. In conclusion, our study demonstrates that formation of M1 was observed in microsomal incubations across various species and strongly suggests incorporation of CO 2 from the bicarbonate buffer, in equilibrium with exogenous CO 2 , into the carbamoyl moiety of the formed N-carbamoyl glucuronide.