Identification of Metabolism and Excretion Differences of Procymidone between Rats and Humans Using Chimeric Mice: Implications for Differential Developmental Toxicity.

A metabolite of procymidone, hydroxylated-PCM, causes rat-specific developmental toxicity due to higher exposure to it in rats than in rabbits or monkeys. When procymidone was administered to chimeric mice with rat or human hepatocytes, the plasma level of hydroxylated-PCM was higher than that of procymidone in rat chimeric mice, and the metabolic profile of procymidone in intact rats was well reproduced in rat chimeric mice. In human chimeric mice, the plasma level of hydroxylated-PCM was less, resulting in a much lower exposure. The main excretion route of hydroxylated-PCM-glucuronide was bile (the point that hydroxylated-PCM enters the enterohepatic circulation) in rat chimeric mice, and urine in human chimeric mice. These data suggest that humans, in contrast to rats, extensively form the glucuronide and excrete it in urine, as do rabbits and monkeys. Overall, procymidone's potential for causing teratogenicity in humans must be low compared to that in rats.

Evaluation of single and multiple doses of a novel mGlu2 agonist, a potential antipsychotic therapy, in healthy subjects.

AIMS: The safety, tolerability, pharmacokinetics (PK) and pharmacodynamics of single and multiple doses of a novel mGlu2 agonist were assessed in healthy males. METHODS: In two, Phase 1 investigator- and subject-blind, placebo-controlled studies, oral doses of prodrug LY2979165 were evaluated: single doses (20-150 mg, N = 30) and multiple once-daily (QD) doses (20-400 mg; N = 84), using a titration regimen. The plasma and urine PK of LY2979165 and active moiety, 2812223, were measured. Cerebrospinal fluid (CSF) was collected to determine PK and neurotransmitter levels. Safety parameters were assessed throughout. RESULTS: Nausea and vomiting were dose limiting following single doses; dose titration allowed higher doses to be tested over 14 days. The most common adverse events related to LY2979165 were dizziness, vomiting, nausea, somnolence and headache. The plasma PK of 2812223 were approximately linear with minimal accumulation with QD dosing. Conversion of LY2979165 to 2812223 was extensive, with minimal LY2979165 measurable in plasma. There was no effect of food on the PK of LY2979165 and 2812223. After 60 mg LY2979165 single-dose, 2812223 exposure in CSF was approximately 2-6% and plasma exposure and peak concentrations were approximately four-fold higher than the mGlu2 agonist in vitro EC50 value. No consistent effects were observed on CSF neurotransmitter levels. CONCLUSIONS: Oral doses of LY2979165 up to 60 mg as a single dose and up to 400 mg given as multiple QD doses, using a titration regimen, were well tolerated with linear PK. Overall, these data support further clinical evaluation of LY2979165.

Human metabolism of Δ3-carene and renal elimination of Δ3-caren-10-carboxylic acid (chaminic acid) after oral administration.

We studied the human in vivo metabolism of Δ(3)-carene (CRN), a natural monoterpene which commonly occurs in the human environment. Four healthy human volunteers were orally exposed to a single dose of 10 mg CRN. Each volunteer gave one urine sample before administration and subsequently collected each urine sample within 24 h after administration. The concentration of the proposed CRN metabolites Δ(3)-caren-10-ol (CRN-10-OH), Δ(3)-caren-10-carboxylic acid (chaminic acid, CRN-10-COOH), and Δ(3)-caren-3,4-diol (CRN-3,4-OH) were determined using a very specific and sensitive GC-MS/MS procedure. Other CRN metabolites were investigated using GC-PCI-MS Q1 scan analyses. CRN-10-COOH was detected in each urine sample with maximum concentration (113.0-1,172.9 µg L(-1)) 2-3 h after administration, whereas CRN-10-OH and CRN-3,4-OH were not detected in any of the samples. The renal excretion kinetics of CRN-10-COOH showed an elimination half-life of about 3 h. The cumulative excretion of CRN-10-COOH within 24 h after exposure correlated with about 2 % of the applied dose. The GC-PCI-MS Q1 scan analysis indicated several additional human CRN metabolites; thereof, six spectra enabled the prediction of the corresponding chemical structure. The results of the study indicate that CRN-10-COOH is a relevant product of the human in vivo metabolism of CRN. The oxidation of its allylic methyl group proceeds until the acidic structure without interruption. Thus, the generation of the alcoholic intermediate appeared to be the rate-determining step of this metabolic route. Nevertheless, the proportion of CRN-10-COOH in the CRN metabolism is low, and other oxidative metabolites are likely. This hypothesis was confirmed by the discovery of additional human CRN metabolites, whose predicted chemical structures fit in with further oxidative products of CRN metabolism.

Sensitive monitoring of monoterpene metabolites in human urine using two-step derivatisation and positive chemical ionisation-tandem mass spectrometry.

A gas chromatographic-positive chemical ionisation-tandem mass spectrometric (GC-PCI-MS/MS) method for the simultaneous determination of 10 oxidative metabolites of the monoterpenoid hydrocarbons α-pinene, (R)-limonene, and Δ(3)-carene ((+)-3-carene) in human urine was developed and tested for the monoterpene biomonitoring of the general population (n=36). The method involves enzymatic cleavage of the glucuronides followed by solid-supported liquid-liquid extraction and derivatisation using a two-step reaction with N,O-bis(trimethylsilyl)-trifluoroacetamide and N-(trimethylsilyl)imidazole. The method proved to be both sensitive and reliable with detection limits ranging from 0.1 to 0.3 µg L(-1). In contrast to the frequent and distinct quantities of (1S,2S,4R)-limonene-1,2-diol, the (1R,2R,4R)-stereoisomer could not be detected. The expected metabolite of (+)-3-carene, 3-caren-10-ol was not detected in any of the samples. All other metabolites were detected in almost all urine samples. The procedure enables for the first time the analysis of trace levels of a broad spectrum of mono- and bicyclic monoterpenoid metabolites (alcohols, diols, and carboxylic acids) in human urine. This analytical procedure is a powerful tool for population studies as well as for the discovery of human metabolism and toxicokinetics of monoterpenes.

Determination of benzimidazole- and bicyclic hydantoin-derived selective androgen receptor antagonists and agonists in human urine using LC-MS/MS.

Selective androgen receptor modulators (SARMs) represent a novel class of drugs with tissue-specific agonistic and antagonistic properties, which are prohibited in sports from January 2008 according to the World Anti-Doping Agency. Preventive approaches to restrict the use of SARMs include early implementation of target analytes into doping control screening assays. Five model SARMs were synthesized, four of which are analogs to prostate-specific androgen receptor antagonists with a 5,6-dichloro-benzimidazole nucleus. The fifth SARM is a muscle-tissue specific agonist with a bicyclic hydantoin structure (BMS-564929). Dissociation pathways after negative electrospray ionization were studied using an LTQ-Orbitrap mass analyzer, and diagnostic product ions and common fragmentation patterns were employed to establish a screening procedure that target the intact SARMs as well as putative metabolic products. Sample preparation based on solid-phase extraction and subsequent LC-MS/MS measurement allowed for detection limits of 1-20 ng/mL, intra- and interday precisions of between 2.4 and 13.2% and between 6.5 and 24.2%, respectively. Recoveries varied from 89 to 106%, and tests for ion suppression or enhancement effects were negative for all analytes. [figure: see text]

Pharmacokinetics, metabolism, and excretion of the intestinal peptide transporter 1 (SLC15A1)-targeted prodrug (1S,2S,5R,6S)-2-[(2'S)-(2-amino)propionyl]aminobicyclo[3.1.0.]hexen-2,6-dicarboxylic acid (LY544344) in rats and dogs: assessment of first-pass bioactivation and dose linearity.

The peptidyl prodrug (1S,2S,5R,6S)-2-[(2'S)-(2-Amino)propionyl]a-minobicyclo[3.1.0.]hexen-2,6-dicarboxylic acid, also known as LY544344, was discovered to improve the oral bioavailability of the parent drug (+)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylic acid (LY354740), a potent group II metabotropic glutamate receptor agonist. This prodrug has been shown to deliver high plasma concentrations of the active drug via intestinal peptide transporter 1 (SLC15A1) (PepT1)-mediated intestinal transport and presystemic hydrolysis in preclinical species. The current data describe the pharmacokinetic behavior of LY544344 and LY354740, with a specific focus on the first-pass activation processes and dose linearity in rats and dogs. The PepT1 transporter makes an attractive prodrug target because of its high capacity and relatively broad substrate specificity. This was demonstrated by the wide dose proportionality observed in both species (up to 1000 mg/kg in rats and 140 mg/kg in dogs). After oral administration of LY544344, absorption and bioactivation were extensive and rapid, with greater than 97% of prodrug hydrolysis occurring before its appearance in the hepatic portal vein. Systemic activation was likewise extensive, with 100% conversion of a 7-mg/kg intravenous dose in dogs. Radiolabeled studies confirmed that hydrolysis to LY354740 was the only metabolic pathway and that the excretion pattern of the active drug was not altered by administration of the prodrug. These results demonstrate the nearly ideal prodrug properties of LY544344 and further validate the utility of the peptide transporter-directed approach to prodrug design.

Single-drop liquid-phase microextraction for the determination of hypericin, pseudohypericin and hyperforin in biological fluids by high performance liquid chromatography.

The analysis of hypericin, pseudohypericin (collectively called in this study hypericins) and hyperforin in biological fluids is reported using single-drop liquid-phase microextraction in conjunction with HPLC-UV-fluorescence detection. A new option for analysis of the active principle constituents in biological samples is proposed, reducing the steps required prior to analysis. There are several parameters which determine the mass transfer such as the extraction solvent, drop and sample volumes, extraction time and temperature, pH and ionic strength, stirring rate and depth of needle tip in the bulk solution. These parameters were chosen to optimize the performance in the current study. The method was validated with respect to precision, accuracy and specificity. The intra-day precision values were below 2.3% for the high concentration level of control samples and 6.2% for the low level. The respective inter-day precision values were calculated to be below 4.4 and 7.1%, respectively, for the two concentration levels. Accuracy of the method, calculated as relative error, ranged from -2.6 to 7.0%. It was demonstrated that as long as the extraction procedure is consistently applied, quantitative analysis is performed accurately and reproducibly in human urine and plasma samples. Limits of quantitation (LOQs) in urine were calculated to be 3, 6 and 12 ng/ml for pseudohypericin, hypericin and hyperforin, respectively. Slightly higher limits were measured in plasma, i.e. 5, 12 and 20 ng/ml, for the respective analytes.

Metabolism and disposition of a potent group II metabotropic glutamate receptor agonist, in rats, dogs, and monkeys.

Metabolism and disposition of MGS0028 [(1R,2S,5S,6S)-2-amino-6-fluoro-4-oxobicyclo[3.1.0]hexane-2,6-dicarboxylic acid monohydrate], a potent group II metabotropic glutamate receptor agonist, were examined in three preclinical species (Sprague-Dawley rats, beagle dogs, and rhesus monkeys). In rats, MGS0028 was widely distributed and primarily excreted in urine as parent and as a single reductive metabolite, identified as the 4R-isomer MGS0034 [(1R,2S,4R,5S,6S)-2-amino-6-fluoro-4-hydroxybicyclo[3.1.0]-hexane-2,6-dicarboxylic acid]. MGS0028 had a low brain to plasma ratio at efficacious doses in rats and was eliminated more slowly in rat brain than in plasma. Exposure increased proportionally (1--10 mg/kg p.o.) in rats, with bioavailability>60% at all doses. However, bioavailability was only approximately 20% in monkeys, and MGS0034 was found in relatively high abundance in plasma. In dogs, oral bioavailability was >60%, and the metabolite was not detected. In vitro metabolism was examined in liver subcellular fractions (microsomes and cytosol) from rat, dog, monkey, and human. Reductive metabolism was observed in rat, monkey, and human liver cytosol incubations, but not in dog liver cytosol incubations. No metabolism of MGS0028 was detected in incubations with liver microsomes from any species. Similar to in vivo results, MGS0028 was reduced in cytosol stereospecifically to MGS0034. The rank order of in vitro metabolite formation (monkey >> rat approximately human >> dog) was in agreement with in vivo observations in rats, dogs, and monkeys. Based on the observation of species difference in reductive metabolism, rat and monkey were recommended to be the preclinical species for further characterization prior to testing in humans. Finally, allometric scaling predicts that human pharmacokinetic parameters would be acceptable for further development.

Determination of monobromobimane derivatives of phenylmercapturic and benzylmercapturic acids in urine by high-performance liquid chromatography and fluorimetry.

A method was developed for the determination in human urine of S-phenylmercapturic (PMA) and S-benzylmercapturic (BMA) acids, metabolites respectively of benzene and toluene. PMA and BMA were determined, after alkaline hydrolysis, to give respectively thiophenol and benzylmercaptan, and coupling of the thiol-containing compounds with monobromobimane (MB), by reversed-phase HPLC on a diphenyl-silica bonded cartridge (100 x 4.6 mm I.D., 5 microm particle size) with fluorimetric detection. Wavelengths for excitation and emission were 375 and 480 nm, respectively. The recovery of PMA and BMA from spiked urines was >90% in the 10-500 microg/l range; the quantification limits were respectively 1 and 0.5 microg/l; day-to-day precision at 42 microg/l was C.V. <7%. The suitability of the proposed procedure for the biological monitoring of exposure to low-level airborne concentrations of benzene and toluene, was evaluated by analyzing the urinary excretion of PMA and BMA in subjects exposed to different sources of aromatic hydrocarbons, namely occupationally-unexposed referents (non-smokers, n=15; moderate smokers, n=8; mean number of cigarettes smoked per-day=17 cig/day) and non-smoker workers occupationally exposed to toluene in maintenance operations of rotogravure machines (non-smokers, n=17). Among referents, non-smokers showed values of PMA ranging from <1 to 4.6 microg/l and BMA from 1.0 to 10.4 microg/l; in smokers, PMA values ranging from 1.2 to 6.7 microg/l and BMA from 9.3 to 39.9 microg/l, were observed. In occupationally exposed non-smoker subjects, BMA median excretion value (23.6 microg/l) was higher than in non-smoker referents (3.5 microg/l) (P<0.001) and individual BMA values (y, microg/l) were associated and increased with airborne toluene concentration (x, mg/m3) according to the equation y=6.5+0.65x (r=0.69, P<0.01, n=17). The proposed analytical method appears to be a sensitive and specific tool for biological monitoring of low-level exposure to benzene and toluene mixtures in occupational and environmental toxicology laboratory.

Sex differences in the metabolism of (+)-S-145, a novel thromboxane A2 receptor antagonist in rat.

1. After the oral administration of 5 mg/kg S-1452 to rat, the plasma levels of (+)-S-145 were similar between the male and female, but there were sex differences in the profiles of its beta-oxidized and hydroxylated metabolites in plasma. 2. beta-Oxidation of (+)-S-145 determined in vitro was slightly higher in the female than in the male, and agreed with the plasma levels of the beta-oxidized metabolites. 3. 5-Hydroxylation activities of (+)-S-145 and beta-oxidized metabolites by rat liver microsomes were significantly higher in the male than in the female, but marked sex differences were not observed in 6-hydroxylation activities. These results revealed that differences in monooxygenase activities directly account for the sex differences in the plasma level of 5-hydroxylated metabolites, and that the peroxisomal beta-oxidation enzyme system also affected the plasma level of 6-hydroxylated metabolites. 4. Biliary excretion was higher in the male than in the female, and quantitative identification of metabolites in bile indicated that this was based on the prominent excretion of taurine conjugates in the male rat. This conclusion was supported by the fact that taurine conjugation activity was higher in male liver homogenates than in the female.

Pharmacokinetics and plasma protein binding of the new potent class III antiarrhythmic agent 3-[4-(1H-imidazol-1-yl)benzoyl]-7-isopropyl-3, 7-diazabicyclo[3.3.1]nonane dihydroperchlorate.

GLG-V-13 (3-[4-(1H-imidazol-1-yl)benzoyl]-7-isopropyl-3,7-diazabicyclo [3.3.1] nonane dihydroperchlorate, CAS 155029-33-7) has been shown to be a potent class III antiarrhythmic agent. The oral and intravenous pharmacokinetics and plasma protein binding of GLG-V-13 in dogs and in rabbits have now been investigated. Plasma GLG-V-13 concentration-time profiles, following an i.v. bolus dose of 6 mg/kg, were fitted to a 2-compartment model. The volume of distribution at steady state (Vd(ss)), the total systemic (ClB), and the elimination half-life (t1/2 beta) were 4.441 l/kg, 1.113 l/h/kg, and 2.485 h in dogs and 3.723 l/kg, 1.548 l/h/kg, and 1.401 h in rabbits. Following i.v. dosing, approximately 9.38% of the parent compound was excreted in dogs urine (0-72 h). Changes in plasma GLG-V-13 concentrations, after oral administration of GLG-V-13 (6 mg/kg), were best described by the 1-compartment pharmacokinetic model. The tmax and Cmax were 1.69 h, 0.54 mg/l in dogs and 1.44 h, 0.35 mg/l in rabbits. On oral administration, GLG-V-13 was moderately eliminated (t1/2kel' 1.867 h-1 in dogs and 3.961 h-1 in rabbits, respectively). Oral bioavailability was estimated to be 53.2% +/- 11.3% in dogs and 66.7% +/- 7.7% in rabbits. About 8.74% of the oral dose (6 mg/kg) was excreted via the dog urine (0-72 h). In vitro binding of GLG-V-13 to dog plasma protein was 29.4 +/- 9.90% (from 0.5 to 4 mg/l). Ex vivo binding of GLG-V-13 to dog plasma protein was 10.4 +/- 7.20%.(ABSTRACT TRUNCATED AT 250 WORDS)

Quantitative measurement of zacopride in human plasma and urine by combined gas chromatography/negative ion chemical ionization mass spectrometry.

A highly sensitive and specific assay was developed for routine analysis of zacopride at the femtomole level in human plasma and urine. Zacopride and the deuterated internal standard [(2H3)zacopride] were measured by gas chromatography/negative ion chemical ionization mass spectrometry with methane as the reagent gas. A multiple-step liquid-liquid extraction procedure was used to isolate the two compounds of interest from complex biological matrices. Zacopride was converted to the fluorinated derivative with pentafluoropropionic anhydride. The mass spectrometer was tuned to monitor the very intense [M-HF]- ion at m/z 435 which was generated into the ion source by a dissociative capture process. This assay was performed with 1 ml of plasma or 0.2 ml of urine, and the quantification limit of the method was calculated as 10 pg ml-1 using a suitable statistical test. The very low relative standard deviation and mean percentage error values calculated during the within-day or between-day repeatability assays have clearly demonstrated the ruggedness of the technique for the routine quantitative measurement of zacopride in plasma and urine. Some preliminary results on the pharmacokinetics of this potent drug are presented to illustrate the applicability of this new powerful gas chromatographic/mass spectrometric method.

High-performance liquid chromatographic determination of BRB-I-28, a novel antiarrhythmic agent, in dog plasma and urine.

A sensitive reversed-phase high-performance liquid chromatographic (HPLC) technique with ultraviolet detection has been developed to determine the concentration of BRB-I-28 (I), a novel antiarrhythmic agent, in dog plasma and urine. The mobile phase was acetonitrile-methanol-37.5 mM phosphate buffer, pH 6.8-triethylamine (50:50:75:0.1, v/v). The compound was extracted from dog plasma and urine with chloroform after alkalinization with sodium hydroxide. The extraction recovery was 83% from plasma and 84% from urine. Good linearity (r > 0.996) was observed throughout the ranges 0.1-12.0 micrograms/ml (plasma) and 0.1-8.0 micrograms/ml (urine). Intra- and inter-assay variabilities were less than 4%. The lower limit of quantitation was 0.08 microgram/ml in either plasma or urine. HPLC analysis of plasma and urine samples from a dog treated with I has demonstrated that the method was accurate and reproducible.

Capillary gas chromatographic method for the determination of the thromboxane A2 receptor antagonist S-1452 and its metabolites in human urine.

A capillary gas chromatographic method using a sulphur-specific detector (Hall's electrolytic conductivity detector) was established to determine the thromboxane A2 antagonist S-1452 and its metabolites in human urine. The target species were the free acid (+)-S-145 of the drug and its nine metabolites: the three hydroxyl forms of (+)-S-145 (I, II and III), bis-nor-(+)-S-145 (IV) the hydroxylated forms of IV (V and VI), tetranor-(+)-S-145 (VII) and the hydroxylated forms of VII (VIII and IX). These ten compounds, which have the same sulphur-containing functional group in common, were determined simultaneously. Their conjugated forms, which were assumed to be glucuronides, were also assayed after hydrolysis. The first derivatization was esterification with diazomethane. The second, for the hydroxylated compounds, was trimethylsilylation with bis(trimethylsilyl)trifluoroacetamide. The ten analytes appeared as separate peaks without mutual interference during 5 min. Hall's detector distinguished the ten analytes selectively from the other urinary components, which removed the need for complex clean-up procedures and led to higher sensitivity with a lower noise level. The method is sensitive enough for the assay of substances present at more than 0.1 micrograms/ml of urine. All the compounds could be determined with a high level of precision and accuracy, with 2-5% relative standard deviation and within +/- 5% deviation from the actual value. Day-to-day measurements verified the reproducibility of the method. Recovered substances were quantified by following the time course, and the analytical data together with previously obtained plasma data clarified the metabolism pharmacokinetically.

Identification of urinary metabolites of 2-methyl-3-(1,4,5,6-tetrahydronicotinoyl)pyrazolo[1,5-a]pyridine in rat, rabbit and dog.

The metabolism of KC-764 (2-methyl-3-(1,4,5,6-tetrahydronicotinoyl)pyrazolo[1,5-a]pyridine, CAS 94457-09-7) in rat, rabbit and dog was studied. The urine of animals dosed with 14C-KC-764 was extracted with ethyl acetate after treatment with beta-glucuronidase and arylsulfatase. The metabolites were purified by TLC and HPLC from the extract. Unchanged KC-764 and 16 metabolites were isolated and their structures were identified or proposed by NMR and MS spectrometry. The metabolism of KC-764 took place by the oxidation of the tetrahydropyridine ring, 6,7-position and 2-methyl group of the pyrazolopyridine ring, and their combinations. The oxidation of the tetrahydropyridine ring was predominant in dog, whereas the oxidation of the pyrazolopyridine ring was more important in rabbit. Rat produced the various metabolites by their combination. 6-Oxo and 6-ureido derivatives of the tetrahydropyridine ring were common major metabolites in all animal species studied.

Pharmacokinetics of the new antiplatelet agent 2-methyl-3-(1,4,5,6-tetrahydronicotinoyl)pyrazolo[1,5-a]pyridine in human subjects.

The pharmacokinetics of KC-764 (2-methyl-3-(1,4,5,6-tetrahydronicotinoyl)pyrazolo[1,5-a]pyridine, CAS 94457-09-7) was studied in healthy male adult volunteers after single ascending oral dose and multiple dosing for 7 days. Serum KC-764 concentration attained the peak in 1 h and declined with a half-life of about 2 h at a single oral dose of 5, 10, 20 and 40 mg. No dose dependent pharmacokinetics of KC-764 was demonstrated. Three metabolites were detected in serum, but their concentrations were lower than that of KC-764. 48-h urinary recoveries after single doses were 41.6-46.6% of dose, not being dose-dependent. Urinary recovery of unchanged KC-764 was 1.1-1.6% of dose. Three metabolites were present in greater amount in urine than unchanged KC-764 and two metabolites were less than KC-764. There was little daily variation of serum concentrations and urinary excretion of KC-764 and its metabolites in the multiple dosing (20 mg twice a day) study. The daily and total urinary recovery were same as those after single doses. Food reduced Cmax and tended to delay tmax, but did not influence AUC0----infinity and urinary recovery. Serum protein binding of KC-764 was about 60%, being not dependent on total serum concentration.

Pharmacokinetics of the new antiplatelet agent 2-methyl-3-(1,4,5,6-tetrahydronicotinoyl)pyrazolo[1,5-a]pyridine in laboratory animals.

KC-764 (2-methyl-3-(1,4,5,6-tetrahydronicotinoyl)pyrazolo [1,5-a]pyridine CAS 94457-09-7) and its metabolites in serum and urine were determined after intravenous and oral administration in mice, rats, rabbits and dogs at a dose of 5 mg/kg. KC-764 was rapidly eliminated from serum in all species. The biological half-lives of unchanged KC-764 after intravenous administration in mice, rats, rabbits and dogs were 1.31, 0.29, 1.94 and 1.20 h, respectively. 2-Methyl-3-(1,4,5,6-tetrahydro-6-oxonicotinoyl)pyrazolo-[1,5-a]pyr idine was a common major metabolite in serum of all species, although 6,7-dihydro-6,7-dihydroxy-2-methyl-3-(1,4,5,6- tetrahydro-6-oxonicotinoyl) pyrazolo-[1,5-a]pyridine (M-8) was more abundant in rabbits. Urinary recovery of unchanged KC-764 was as low as 0.4-2.2% in all species. The major urinary metabolite was 2-methyl-3-(1,4,5,6-tetrahydro-6-ureidonicotinoyl)pyrazolo-[1,5-a] pyridine in mice, rats and dogs, but M-8 was in rabbits. KC-764 was rapidly and well absorbed by oral administration, and extensively metabolized in all species tested.

Determination of 6-chloro-3-(3-cyclopropyl 1,2,4-oxadiazol-5-yl)-5-methyl-imidazo less than 1,5-a greater than-quinoxalin-4(5h)-one in rat serum, urine and brain by solid-phase extraction and liquid chromatography.

A simple, rapid, and accurate liquid chromatographic method with ultraviolet detection and solid-phase extraction is described for the quantitation of 6-chloro-3-(3-cyclopropyl 1,2,4-oxadiazol-5-yl)-5-methyl-imidazo less than 1,5-a greater than-quinoxalin-4(5h)-one (I, U-80447) in rat serum, urine and brain. Linear calibration curves were obtained in the concentration ranges of 5 ng ml-1-20 micrograms ml-1 (serum), 20 ng ml-1-20 micrograms ml-1 (urine), and 50 ng g-1-200 micrograms g-1 (brain). Intra- and inter-assay precision and accuracy were all found to be less than 10% at the three concentrations evaluated. The absolute extraction recovery each from serum, urine and brain was greater than or equal to 90%. Application of this method to the quantitation of the title compound in rat serum and brain for a pharmacokinetic study is reported.

Preliminary results of biliary excretion of ramipril after T-drainage in cholecystectomy patients.

Four cholecystectomy patients, aged between 52 and 56 years, weighing between 64 and 90 kg, received 5 mg of ramipril as a single dose in order to investigate the pharmacokinetics and excretion pattern of ramipril. All patients had a T-drainage that allowed bile collection. Serum was collected at regular intervals, bile was collected hourly for 6 h followed by a 6- and 12-h fraction, and urine was collected every 2 h for 8 h followed by a 4- and 12-h fraction. The concentrations of ramipril and ramiprilat in serum and ramipril, ramiprilat, ramipril glucuronide, ramiprilat glucuronide, diketopiperazine, and diketopiperazine acid in bile and urine were determined and the amounts excreted in urine and bile over 24 h were calculated. There were great interindividual differences in maximum concentrations as well as in the time to reach maximum concentrations in plasma and bile as well as in the excretion pattern between urine and bile. The highest concentrations in bile were found for diketopiperazine acid (3,080 ng/ml) and ramipril glucuronide (2,414 ng/ml). In general, only minimal amounts of unchanged ramipril (prodrug) were detected in the bile. In the urine, the major metabolites excreted were diketopiperazine acid, ramiprilat, and diketopiperazine in amounts of 537, 188, and 124 micrograms, respectively. In bile, the main substances excreted were diketopiperazine acid and ramiprilat glucuronide, which amounted to 501 and 314 micrograms, respectively. Biliary excretion may be the explanation for the noncomplete urinary recovery of ramipril and its metabolites.