No abstract

ABSTRACT:Tigecycline, a novel, first-in-class glycylcycline antibiotic, has been approved for the treatment of complicated intra-abdominal infections and complicated skin and skin structure infections. The pharmacokinetics, metabolism, and excretion of [ 14 C]tigecycline were examined in healthy male volunteers. Tigecycline has been shown to bind to bone; thus, to minimize the amount of radioactivity binding to bone and to maximize the recovery of radioactivity, tigecycline was administered intravenously (30-min infusion) as a single 100-mg dose, followed by six 50-mg doses, every 12 h, with the last dose being [ 14 C]tigecycline (50 Ci). After the final dose, the pharmacokinetics of tigecycline in serum showed a long halflife (55.8 h) and a large volume of distribution (21.0 l/kg), whereas radioactivity in serum had a shorter half-life (6.9 h) and a smaller volume of distribution (3.3 l/kg). The major route of elimination was feces, containing 59% of the radioactive dose, whereas urine contained 32%. Unchanged tigecycline was the predominant drugrelated compound in serum, urine, and feces. The major metabolic pathways identified were glucuronidation of tigecycline and amide hydrolysis followed by N-acetylation to form N-acetyl-9-aminominocycline. The glucuronide metabolites accounted for 5 to 20% of serum radioactivity, and approximately 9% of the dose was excreted as glucuronide conjugates within 48 h. Concentrations of N-acetyl-9-aminominocycline were approximately 6.5% and 11% of the tigecycline concentrations in serum and urine, respectively. Excretion of unchanged tigecycline into feces was the primary route of elimination, and the secondary elimination pathways were renal excretion of unchanged drug and metabolism to glucuronide conjugates and N-acetyl-9-aminominocycline.

Metabolite identification studies remain an integral part of pre-clinical and clinical drug development programs. Analysis of biological matrices, such as plasma, urine, feces and bile, pose challenges due to the large amounts of endogenous components that can mask a drug and its metabolites. Although direct infusion nanoelectrospray using capillaries has been used routinely for proteomic studies, metabolite identification has traditionally employed liquid chromatographic (LC) separation prior to analysis. A method is described here for rapid metabolite profiling in biological fluids that involves initial sample clean-up using pipette tips packed with reversed-phase material (i.e. ZipTips) to remove matrix components followed by direct infusion nanoelectrospray on an LTQ/Orbitrap mass spectrometer using a protonated polydimethylcyclosiloxane cluster ion for internal calibration. We re-examined samples collected from a prazosin metabolism study in the rat. Results are presented that demonstrate that sub parts-per-million accuracies can be achieved on molecular ions, facilitating identification of metabolites, and on product ions, facilitating structural assignments. The data also show that the high-resolution measurements (R = 100,000 at m/z 400) enable metabolites of interest to be resolved from endogenous components. The extended analysis times available with nanospray enables signal averaging for 1 min or more that is valuable when metabolites are present in low concentrations as encountered here in plasma and brain. Using this approach, the metabolic fate of a drug can be quickly obtained. A limitation of this approach is that metabolites that are structural isomers cannot be distinguished, although such information can be collected by LC/MS during follow-on experiments.

ABSTRACT:Bazedoxifene is a selective estrogen receptor modulator under development for the prevention and treatment of osteoporosis. The disposition of [ 14 C]bazedoxifene was determined in six healthy postmenopausal women after administration of a single oral dose of 20 mg (200 Ci). After dosing, blood was collected at frequent intervals, and urine and fecal samples were collected for up to 10 days. Aliquots of plasma, blood, urine, and fecal homogenates were analyzed for concentrations of radioactivity. Bazedoxifene metabolite profiles in plasma and feces were determined by highperformance liquid chromatography with radioactivity flow detection; metabolite structures were confirmed by liquid chromatography-mass spectrometry. Bazedoxifene was rapidly absorbed, exhibiting a mean peak plasma concentration of 3.43 ng/ml at 1.2 h postdose. The total mean recovery of the radioactive dose in excreta was 85.6%, with the majority recovered in feces (84.7%) and only a small fraction (0.81%) in urine. Radiochromatograms of plasma revealed that glucuronidation was the major metabolic pathway; little or no cytochrome P450-mediated metabolism was evident. The majority of circulating radioactivity was constituted by metabolites, with bazedoxifene-5-glucuronide being the predominant metabolite (up to 95%). Bazedoxifene-4-glucuronide was a minor metabolite (up to 20%), and unchanged bazedoxifene represented 0 to 13% of the radioactivity in most plasma samples. Unchanged bazedoxifene was the major radioactive component in feces, however, reflecting unabsorbed drug and/or glucuronides that were hydrolyzed by intestinal bacterial enzymes. [ 14 C]Bazedoxifene was generally well tolerated. These findings demonstrated that, after oral administration in healthy postmenopausal women, bazedoxifene was rapidly absorbed, metabolized via glucuronidation, and excreted predominantly in feces.

ABSTRACT:Prazosin (2-[4-(2-furanoyl)-piperazin-1-yl]-4-amino-6,7-dimethoxyquinazoline) is an antihypertensive agent that was introduced to the market in 1976. It has since established an excellent safety record. However, in vitro metabolism of prazosin has not been investigated. This study describes the in vitro biotransformation of prazosin in liver microsomes from rats, dogs, and humans, as well as rat and human cryopreserved hepatocytes and characterization of metabolites using liquid chromatography/tandem mass spectrometry. The major in vivo biotransformation pathways reported previously in rats and dogs include demethylation, amide hydrolysis, and O-glucuronidation. These metabolic pathways were also confirmed in our study. In addition, several new metabolites were characterized, including a stable carbinolamine, an iminium species, and an enamine-all formed via oxidation of the piperazine ring. Two ring-opened metabolites generated following oxidative cleavage of the furan ring were also identified. Using semicarbazide hydrochloride as a trapping agent, an intermediate arising from opening of the furan ring was captured as a pyridazine product. In the presence of glutathione, three glutathione conjugates were detected in microsomal incubations, although they were not detected in cryopreserved hepatocytes. These data support ring opening of the furan via a reactive ␥-keto-␣,␤-unsaturated aldehyde intermediate. In the presence of UDP-glucuronic acid, prazosin underwent conjugation to form an N-glucuronide not reported previously. Our in vitro investigations have revealed additional metabolic transformations of prazosin and have shown the potential of prazosin to undergo bioactivation through metabolism of the furan ring to a reactive intermediate.Prazosin (Fig. 1), a short-acting vasodilator discovered in the mid1970s, has been widely used in treating hypertension and congestive heart failure (Constantine, 1974;Althuis and Hess, 1977;Stanaszek et al., 1983). It was the first of a new class of direct-acting vasodilators acting by ␣-adrenoreceptor blockade. Prazosin was introduced to the U.S. market as MINIPRESS by Pfizer (New York, NY) in 1976. Prazosin is well tolerated, with the most common side effect associated with treatment being postural hypotension. Although no longer a major drug among the antihypertensive agents, based on prazosin's ability to antagonize centrally located ␣ 1 -adrenergic receptors, a new indication for treatment of post-traumatic stress disorders (PTSD) encountered during civilian life is being explored in clinical studies (Taylor and Raskind, 2002). In addition, prazosin is also being investigated in treating combat-related nightmares characteristic of PTSD among soldiers recently returned from Operation Iraqi Freedom (Daly et al., 2005).In vitro metabolism of prazosin in animals or humans has not been reported. Early metabolism studies with 14 C-labeled prazosin (label in quinazoline ring) in rats and dogs revealed that it undergoes extensive hepatic metabolism with biliary excret...