Vinod R. Shah

Simultaneous High-Performance Liquid Chromatographic Analysis of Cefprozil Diastereomers in a Pharmacokinetic Study

Cefprozil, a new oral cephalosporin, consists of a 90:10 cis:trans isomer mixture. Sensitive, specific and reproducible high performance liquid chromatographic methods have been developed for the simultaneous quantification of the two stereoisomers of cefprozil in plasma and urine samples from human and rats. Cephalexin acted as the internal standard. Plasma protein was precipitated with acetonitrile and trichloracetic acid with subsequent extraction of acetonitrile. After vortexing and centrifuging, the aqueous phase was injected onto a reverse phase C8 column. Urine samples were acidified with sodium acetate buffer (pH 3.8) and then directly injected onto a reverse phase C18 column. The detector was set at 280 nm. These methods were applied to determine protein binding of both isomers in human and rat sera, and to perform a pharmacokinetic study in human. Results showed that both isomers bound moderately to serum proteins with no interference by the other isomer. The pharmacokinetic study in human indicated that cefprozil was well absorbed and the cis and trans isomers have similar pharmacokinetics.

Sensitive liquid chromatographic–mass spectrometric assay for the simultaneous quantitation of nefazodone and its metabolites hydroxynefazodone m-chlorophenylpiperazine and triazole-dione in human plasma using single-ion monitoring

A sensitive, selective, accurate, precise and reproducible high-performance liquid chromatographic-mass spectrometric (LC-MS) assay was developed and validated for the simultaneous determination of nefazodone (NEF), hydroxynefazodone (OH-NEF), m-chlorophenylpiperazine (mCPP), and triazole-dione (Dione) in human plasma using trazodone (TRZ) as the internal standard (I.S.). The method involved simultaneous protein precipitation with acetonitrile and liquid-liquid extraction with methylene chloride, after which the organic layer was evaporated to dryness. The residue was reconstituted in 25% acetonitrile in 10 mM ammonium formate (pH 4.0), and an aliquot was injected onto a BDS Hypersil C18 column at a flow-rate of 0.3 ml/min. The mobile phase comprising of 10 mM ammonium formate (pH 4) and acetonitrile in 55:45 (v/v) was used in an isocratic condition. The mass spectrometer was programmed to admit the protonated molecules at m/z 197.0 (mCPP), 372.0 (I.S.) 470.4 (NEF), 458.1 (Dione), and 486.2 (OH-NEF). Standard curves were linear (r2 > or = 0.995) over the concentration range of 4-1000 ng/ml for Dione and 2-500 ng/ml for other analytes. The lowest standard concentrations were the lower limit of quantitation for each analyte. The mean predicted quality control (QC) concentrations for all analytes deviated less than -12.1% from the corresponding nominal values; the intra-assay and inter-assay precision of the assay for all analytes were within 7.0% relative standard deviation. All analytes including I.S. were stable in the injection solvent at room temperature for at least 24 h. The extraction recovery of the various analytes ranged from 67.3 to 86.5%. The validated assay was applied to a pharmacokinetic study of nefazodone.

Oral Absolute Bioavailability and Intravenous Dose-Proportionality of Cefprozil in Humans

The absolute bioavailability (F) and dose proportionality of cefprozil were investigated in a parallel design study with an embedded two‐way crossover leg. Twenty‐four healthy male subjects divided into 3 dosing groups received a single 250‐, 500‐, or 1000‐mg dose of cefprozil by a 30‐minute intravenous infusion. Subjects assigned to the 500‐mg dose group also received a 500‐mg oral dose of cefprozil in crossover manner with a wash‐out period of 7 days between each treatment. Cefprozil consists of cis and trans isomers in an approximate 90:10 ratio. Serial blood and urine samples were collected and analyzed for the concentrations of the cis and trans isomers of the cephalosporin using high‐pressure liquid chromatographic assay with UV detection methods. After the 250‐, 500‐, and 1000‐mg intravenous administration of cefprozil, the peak concentrations were 13.2, 26.0, and 48.5 μg/mL, and area under the plasma concentration versus time proxies were 17.2, 31 A, and 58.1 μg · hour/mL, respectively, for the cis isomer increasing in a dose proportional manner. Total body clearance, renal clearance, and volume of distribution at steady state, adjusted for body weight, were not significantly different among all groups. Mean residence time, elimination half‐life, and urinary recovery were invariant with the dose. Based on the plasma and urine data, the estimates of F were 89% and 94% for the cis isomer, respectively. The plasma concentrations of the trans isomer were about 1/10th of the cis isomer, and all parameters were similar to those observed for the cis isomer. In summary, cefprozil exhibits linear pharmacokinetics and is essentially completely absorbed after oral administration.

High-performance liquid chromatographic-electrochemical assay for the quantitation of BMS-181885 in monkey plasma.

A high-performance liquid chromatographic-electrochemical assay was developed and validated for the quantitation of BMS-181885 (I), an anti-migraine agent, in monkey plasma. The assay involved a solid-phase extraction of I and BMY-46317 (internal standard; I.S.) on a 1-ml cyano cartridge using the automatic solid-phase extraction cartridge (ASPEC) system. Immediately following the conditioning of the cyano column (3 ml of methanol and 2 ml of 1% glacial acetic acid), plasma (0.25 ml) was loaded on to the column. The column was then washed with a 3 ml of 0.1 M ammonium acetate buffer (pH 6). The final elution of the analytes was performed using 2 ml of methanol. The eluate was then evaporated to dryness (gentle stream of nitrogen at 40 degrees C) and the residue was dissolved in the mobile phase and injected on to a YMC basic column (15 cm x 4.6 mm; 5 microm particle size) at a flow-rate of 1 ml/min. A mixture of 0.1 M ammonium acetate at pH 6-acetonitrile-methanol (70:20:10, v/v) was used as the mobile phase. Standard curves, with a lower limit of quantitation of 2 ng/ml of I were linear (r2> or =0.998; range: 2-50 ng/ml). Based on the analysis of the quality control (QC) samples, the assay was both accurate and precise. The stability of I was established following freeze-thaw cycles and storage at or below -20 degrees C. The extraction recovery of I from monkey plasma was about 82%. The validated assay method was applied to determine the pharmacokinetics of I in monkeys following a single 1 mg/kg intravenous dose.

A Simple HPLC Assay, with Ultraviolet Detection, For Determination of a Monobactam Antibiotic

Abstract BMS-180680 is a monobactam antibiotic currently under development for the treatment of gram negative bacteria including Pseudomonas aeruginosa. Simple, rapid, sensitive, precise, and reproducible assay methods have been developed for the quantification of BMS-180680 in dog and rat plasma using aztreonam as the internal standard. The assay methods involve a single-step protein precipitation by addition of acetonitrile, followed immediately by centrifugation, after which the supernatant is evaporated to dryness (65°C) under a gentle stream of nitrogen. The residue is re-constituted in the mobile phase, transferred to a micro-WISP vial and injected on to a Zorbax It, C-18 HPLC column preceded by a guard column. Mobile phase comprised 17% acetonitrile and 83% of 40 mM dibasic ammonium phosphate and 6 mM tetrabutylammonium hydrogen sulfate. A small amount of tetrahydrofuran (20 mL/liter of mobile phase) was added and the apparent pH of the mobile phase was adjusted to 4.1 with 85% phosphoric acid. The...

Pharmacokinetic assessment of the sites of first-pass metabolism of BMS-181101, an antidepressant agent, in rats.

The relative contribution of the gut and the liver to the first‐pass metabolism of BMS‐181101 (3‐[3‐[4‐(5‐methoxy‐4‐pyrimidinyl)‐1‐piperazinyl]propyl]‐5‐fluoro‐1H‐indole dihydrochloride), a potential antidepressant agent, has been evaluated in rats.