Population pharmacokinetic modeling and simulation to support qualification of pyridoxic acid as endogenous biomarker of OAT1/3 renal transporters.

Renal clearance of many drugs is mediated by renal organic anion transporters OAT1/3 and inhibition of these transporters may lead to drug-drug interactions (DDIs). Pyridoxic acid (PDA) and homovanillic acid (HVA) were indicated as potential biomarkers of OAT1/3. The objective of this study was to develop a population pharmacokinetic model for PDA and HVA to support biomarker qualification. Simultaneous fitting of biomarker plasma and urine data in the presence and absence of potent OAT1/3 inhibitor (probenecid, 500 mg every 6 h) was performed. The impact of study design (multiple vs. single dose of OAT1/3 inhibitor) and ability to detect interactions in the presence of weak/moderate OAT1/3 inhibitors was investigated, together with corresponding power calculations. The population models developed successfully described biomarker baseline and PDA/HVA OAT1/3-mediated interaction data. No prominent effect of circadian rhythm on PDA and HVA individual baseline levels was evident. Renal elimination contributed greater than 80% to total clearance of both endogenous biomarkers investigated. Estimated probenecid unbound in vivo OAT inhibitory constant was up to 6.4-fold lower than in vitro values obtained with PDA as a probe. The PDA model was successfully verified against independent literature reported datasets. No significant difference in power of DDI detection was found between multiple and single dose study design when using the same total daily dose of 2000 mg probenecid. Model-based simulations and power calculations confirmed sensitivity and robustness of plasma PDA data to identify weak, moderate, and strong OAT1/3 inhibitors in an adequately powered clinical study to support optimal design of prospective clinical OAT1/3 interaction studies.

Physiologically Based Pharmacokinetic Models of Probenecid and Furosemide to Predict Transporter Mediated Drug-Drug Interactions.

PURPOSE: To provide whole-body physiologically based pharmacokinetic (PBPK) models of the potent clinical organic anion transporter (OAT) inhibitor probenecid and the clinical OAT victim drug furosemide for their application in transporter-based drug-drug interaction (DDI) modeling. METHODS: PBPK models of probenecid and furosemide were developed in PK-Sim®. Drug-dependent parameters and plasma concentration-time profiles following intravenous and oral probenecid and furosemide administration were gathered from literature and used for model development. For model evaluation, plasma concentration-time profiles, areas under the plasma concentration-time curve (AUC) and peak plasma concentrations (Cmax) were predicted and compared to observed data. In addition, the models were applied to predict the outcome of clinical DDI studies. RESULTS: The developed models accurately describe the reported plasma concentrations of 27 clinical probenecid studies and of 42 studies using furosemide. Furthermore, application of these models to predict the probenecid-furosemide and probenecid-rifampicin DDIs demonstrates their good performance, with 6/7 of the predicted DDI AUC ratios and 4/5 of the predicted DDI Cmax ratios within 1.25-fold of the observed values, and all predicted DDI AUC and Cmax ratios within 2.0-fold. CONCLUSIONS: Whole-body PBPK models of probenecid and furosemide were built and evaluated, providing useful tools to support the investigation of transporter mediated DDIs.

Phase I randomized clinical trial of N-acetylcysteine in combination with an adjuvant probenecid for treatment of severe traumatic brain injury in children.

BACKGROUND: There are no therapies shown to improve outcome after severe traumatic brain injury (TBI) in humans, a leading cause of morbidity and mortality. We sought to verify brain exposure of the systemically administered antioxidant N-acetylcysteine (NAC) and the synergistic adjuvant probenecid, and identify adverse effects of this drug combination after severe TBI in children. METHODS: IRB-approved, randomized, double-blind, placebo controlled Phase I study in children 2 to 18 years-of-age admitted to a Pediatric Intensive Care Unit after severe TBI (Glasgow Coma Scale [GCS] score ≤8) requiring an externalized ventricular drain for measurement of intracranial pressure (ICP). Patients were recruited from November 2011-August 2013. Fourteen patients (n = 7/group) were randomly assigned after obtaining informed consent to receive probenecid (25 mg/kg load, then 10 mg/kg/dose q6h×11 doses) and NAC (140 mg/kg load, then 70 mg/kg/dose q4h×17 doses), or placebos via naso/orogastric tube. Serum and CSF samples were drawn pre-bolus and 1-96 h after randomization and drug concentrations were measured via UPLC-MS/MS. Glasgow Outcome Scale (GOS) score was assessed at 3 months. RESULTS: There were no adverse events attributable to drug treatment. One patient in the placebo group was withdrawn due to adverse effects. In the treatment group, NAC concentrations ranged from 16,977.3±2,212.3 to 16,786.1±3,285.3 in serum and from 269.3±113.0 to 467.9±262.7 ng/mL in CSF, at 24 to 72 h post-bolus, respectively; and probenecid concentrations ranged from 75.4.3±10.0 to 52.9±25.8 in serum and 5.4±1.0 to 4.6±2.1 µg/mL in CSF, at 24 to 72 h post-bolus, respectively (mean±SEM). Temperature, mean arterial pressure, ICP, use of ICP-directed therapies, surveillance serum brain injury biomarkers, and GOS at 3 months were not different between groups. CONCLUSIONS: Treatment resulted in detectable concentrations of NAC and probenecid in CSF and was not associated with undesirable effects after TBI in children. TRIAL REGISTRATION: ClinicalTrials.gov NCT01322009.

Differences in the pharmacokinetics of 4-amino-3-chlorophenyl hydrogen sulfate, a metabolite of resatorvid, in rats and dogs.

The pharmacokinetics of 4-amino-3-chlorophenyl hydrogen sulfate, M-III of resatorvid, in rats and dogs were investigated using radiolabeled M-III ([(14)C]M-III). The elimination half-life of (14)C in the plasma of rats was approximately 1/30 of that of dogs after intravenous dosing of [(14)C]M-III at 0.5 mg/kg to rats and dogs. The in vitro and in vivo plasma protein binding ratios of M-III were relatively high and were the same in both species. The intrinsic clearance (CL(int)) of M-III in rats was much higher than the glomerular filtration rate in rats. Furthermore, the concentration of [(14)C]M-III in the kidney of rats was much higher than that in the plasma. On the contrary, in dogs, the concentration of [(14)C]M-III in the kidney was very much lower than that in the plasma. These results indicated that M-III was effectively taken up into the kidney and was excreted into the urine in rats; however, in dogs, ineffective renal uptake of M-III was presumed. When [(14)C]M-III and probenecid were simultaneously and continually infused intravenously to rats, the CL(int) of M-III decreased with increasing plasma concentrations of probenecid, indicating that kidney uptake of M-III in rats was inhibited by probenecid. It was also thought that uptake by the organic anion transport system(s) in the basolateral membrane is involved in the renal uptake of M-III in rats. The pharmacokinetic differences of M-III between rats and dogs are considered to be mainly caused by the difference in the urinary excretion via the renal distribution processes.

G-protein coupled receptor 35 (GPR35) activation and inflammatory pain: Studies on the antinociceptive effects of kynurenic acid and zaprinast.

G-protein coupled receptor 35 (GPR35) is a former "orphan receptor" expressed in brain and activated by either kynurenic acid or zaprinast. While zaprinast has been studied as a phosphodiesterase inhibitor, kynurenic acid (KYNA) is a tryptophan metabolite and has been proposed as the endogenous ligand for this receptor. In the present work, we showed that GPR35 is present in the dorsal root ganglia and in the spinal cord and in order to test the hypothesis that GPR35 activation could cause analgesia, we administered suitable doses of zaprinast or we increased the local concentration of KYNA by administering a precursor (kynurenine) or by inhibiting its disposal from the CNS (with probenecid). We used the "writhing test" induced by acetic acid i.p. injection in mice. KYNA and kynurenine plasma and spinal cord levels were measured with HPLC techniques. Kynurenine (30, 100, 300 mg/kg s.c.) increased plasma and spinal cord levels of KYNA and decreased the number of writhes in a dose dependent manner. Similarly, probenecid was able to increase KYNA levels in plasma and spinal cord, to reduce the number of writes and to amplify kynurenine effects. Furthermore, zaprinast had antinociceptive effects in the writhing test without affecting KYNA levels. In agreement with its affinity for GPR35 receptor (approximately 10 times higher than that of KYNA), zaprinast action occurred at relatively low doses. No additive actions were obtained when kynurenine and zaprinast were administered at maximally active doses. Our results suggest that GPR35 could be an interesting target for innovative pharmacological agents designed to reduce inflammatory pain. This article is part of a Special Issue entitled 'Trends in neuropharmacology: in memory of Erminio Costa'.

Competitive inhibition of renal tubular secretion of gemifloxacin by probenecid.

Probenecid interacts with transport processes of drugs at several sites in the body. For most quinolones, renal clearance is reduced by concomitant administration of probenecid. The interaction between gemifloxacin and probenecid has not yet been studied. We studied the extent, time course, site(s), and mechanism of this interaction. Seventeen healthy volunteers participated in a randomized, two-way crossover study. Subjects received 320 mg gemifloxacin as an oral tablet without and with 4.5 g probenecid divided in eight oral doses. Drug concentrations in plasma and urine were analyzed by liquid chromatography-tandem mass spectrometry. WinNonlin was used for noncompartmental analysis, compartmental modeling, and statistics, and NONMEM was used for visual predictive checks. Concomitant administration of probenecid increased plasma gemifloxacin concentrations and amounts excreted in urine compared to baseline amounts. Data are average estimates (percent coefficients of variation). Modeling showed a competitive inhibition of the renal tubular secretion of gemifloxacin by probenecid as the most likely mechanism of the interaction. The estimated K(m) and Vmax for the saturable part of renal elimination were 9.16 mg/liter (20%) and 113 mg/h (21%), respectively. Based on the molar ratio, the affinity for the renal transporter was 10-fold higher for gemifloxacin than for probenecid. Since probenecid reached an approximately 200-times-higher area under the molar concentration-time curve from 0 to 24 h than gemifloxacin, probenecid inhibited the active tubular secretion of gemifloxacin. Probenecid also reduced the nonrenal clearance of gemifloxacin from 25.2 (26%) to 21.0 (23%) liters/h. Probenecid inhibited the renal tubular secretion of gemifloxacin, most likely by a competitive mechanism, and slightly decreased nonrenal clearance of gemifloxacin.

Population pharmacokinetics of oseltamivir when coadministered with probenecid.

Oseltamivir is a potent, selective, oral neuraminidase inhibitor for the treatment and prophylaxis of influenza. Plasma concentrations of the active metabolite, oseltamivir carboxylate, are increased in the presence of probenecid, suggesting that the combination could allow for the use of reduced doses of oseltamivir. To investigate this proposal, we developed a population pharmacokinetic model and simulated the pharmacokinetics of candidate combination regimens of oral oseltamivir (45 mg and 30 mg twice a day) plus oral probenecid (500 mg/6 hourly). Probenecid plus oseltamivir 45 mg achieved all the pharmacokinetic parameters expected of oseltamivir alone, but combination with oseltamivir 30 mg and dose interval extension approaches did not. An oseltamivir-probenecid combination may compromise tolerability and enhance the potential for drug interactions. In addition, increased dosing requirements may affect compliance and attainment of optimal oseltamivir exposure, potentially facilitating the emergence of viral strains with reduced susceptibility to oseltamivir. These factors, set alongside increased capacity for oseltamivir production, should be carefully considered before an oseltamivir-probenecid combination is used.

Pharmacokinetic and pharmacodynamic interaction between allopurinol and probenecid in healthy subjects.

BACKGROUND AND OBJECTIVE: Combination therapy with allopurinol and probenecid is used to treat tophaceous gout in patients who do not respond sufficiently to allopurinol alone. However, the potential interaction between these drugs has not been systematically investigated. The objective of this study was to investigate the pharmacokinetics and hypouricaemic effect of oxypurinol (the active metabolite of allopurinol) and probenecid when administered alone and in combination in healthy subjects. METHODS: An open-label, randomized, three-way crossover clinical trial was conducted in 12 healthy adults. Subjects were randomized to receive treatment for 7 days with allopurinol (150 mg twice daily), probenecid (500 mg twice daily) or combination therapy with both drugs, with a 7-day washout period between treatments. Venous blood samples were collected predose (at 0 hours) and 1, 2, 3, 4, 6, 8, 10 and 12 hours after dosage for determination of oxypurinol and/or probenecid concentrations. Plasma and urinary urate concentrations were determined on each study day and at the end of each washout period. Pharmacokinetic and pharmacodynamic parameters were analysed using two-way ANOVA. RESULTS: Coadministration of allopurinol and probenecid significantly reduced average steady-state plasma oxypurinol concentrations (mean+/-SD: allopurinol alone 9.7+/-2.1 mg/L vs combination 5.1+/-1.0 mg/L, p<0.001). Probenecid concentrations were unaffected. Plasma urate concentrations decreased (p<0.01) during allopurinol therapy (0.16+/-0.05 mmol/L), probenecid therapy (0.13+/-0.02 mmol/L) and combination therapy (0.09+/-0.02 mmol/L) compared with baseline (0.30+/-0.05 mmol/L). CONCLUSION: Coadministration of allopurinol and probenecid to healthy subjects had a greater hypouricaemic effect than either allopurinol or probenecid alone, despite a reduction in plasma oxypurinol concentrations when the drugs were taken concomitantly.

Tubular transporters and clearance of adefovir.

Adefovir is transported by the organic anion transporter (OAT1) and the multidrug resistant protein (MRP2, 4 and 5). We studied adefovir clearance in rat after inhibition of transporters by probenecid and in mutant transport-deficient (TR-) rats, in which MRP2 is lacking. After treatment by probenecid or placebo, pharmacokinetics of adefovir 10mg/kg was studied via population nonlinear mixed effect modeling. The fraction of drug excreted in the urine was low. Renal clearance of adefovir was significantly lower (P < 0.05) in probenecid TR- rats (0.03+/-0.02l/h) than in normal control (0.09+/-0.05l/h), in normal probenecid (0.10+/-0.07l/h) and in TR- control rats (0.13+/-0.07l/h). In vivo in rats MRP2 mutation alone did not affect adefovir clearance suggesting that MRP2 does not play a critical role in the secretion of adefovir. Additional pharmacological inhibition of transporters decreased renal clearance, which may reflect inhibition of compensating transport mechanisms activated when MRP2 is lacking.

Is the monkey an appropriate animal model to examine drug-drug interactions involving renal clearance? Effect of probenecid on the renal elimination of H2 receptor antagonists.

The renal drug-drug interaction between famotidine (an H(2) receptor antagonist) and probenecid has not been reproduced in rats. We have proposed that this is caused by a species difference in the transport activity by human/rat organic anion transporter (OAT) 3 and the expression of organic cation transporter (OCT) 1 in the rodent kidney. Since monkey OATs (mkOATs) exhibit similar transport activities to human orthologs, it is hypothesized that in vivo studies in monkeys will allow a more precise prediction of renal drug-drug interactions in humans. Famotidine and cimetidine were efficiently taken up by mkOAT3-expressing human embryonic kidney cells (Km, 154 and 71 microM, respectively), and their uptake was strongly inhibited by probenecid (Ki, 3.0-5.7 microM). Quantification of mkOCT1 and mkOCT2 mRNAs in the monkey kidney using real-time reverse transcription-polymerase chain reaction revealed their predominant expression in the liver and kidney, respectively. Crossover studies were conducted in cynomolgus monkeys. Famotidine was given by i.v. administration, with or without probenecid. Probenecid treatment caused a 65% reduction in the renal clearance (0.426 +/- 0.079 versus 0.165 +/- 0.027 l/h/kg) and a 90% reduction in the tubular secretion clearance (0.275 +/- 0.075 versus 0.0230 +/- 0.0217 l/h/kg), whereas it had no effect on the renal clearance of cimetidine. In contrast to the species-dependent effect of probenecid, allometric scaling using animal data (rat, dog, and monkey) successfully predicted the renal and tubular secretion clearance of famotidine in humans. These results suggest that monkeys are more appropriate animal species for predicting the renal drug-drug interactions in humans.

Carrier-mediated uptake of H2-receptor antagonists by the rat choroid plexus: involvement of rat organic anion transporter 3.

The choroid plexus (CP) acts as a site for the elimination of xenobiotic organic compounds from the cerebrospinal fluid (CSF). The purpose of the present study is to investigate the role of rat organic anion transporter 3 (rOat3; Slc22a8) in the uptake of H(2)-receptor antagonists (cimetidine, ranitidine, and famotidine) by the isolated rat CP. Saturable uptake of cimetidine and ranitidine was observed in rOat3-LLC with K(m) values of 80 and 120 microM, respectively, whereas famotidine was found to be a poor substrate. The steady-state concentration of the H(2)-receptor antagonists in the CSF was significantly increased by simultaneously administered probenecid, although it did not affect their brain and plasma concentrations. Saturable uptake of cimetidine and ranitidine was observed in the isolated rat CP with K(m) values of 93 and 170 microM, respectively, whereas 50% of the uptake of famotidine remained at the highest concentration examined (1 mM). The K(i) value of ranitidine for the uptake of cimetidine by the isolated CP (50 microM) was similar to its own K(m) value, suggesting that they share the same transporter for their uptake. The inhibition potency of organic anions such as benzylpenicillin, estradiol 17beta-glucuronide, p-aminohippurate, and estrone sulfate for the uptake of cimetidine by the isolated rat CP was similar to that for benzylpenicillin, the uptake of which has been hypothesized to be mediated by rOat3, whereas a minimal effect by tetraethylammonium excludes involvement of organic cation transporter(s). These results suggest that rOat3 is the most likely candidate transporter involved in regulating the CSF concentration of H(2)-receptor antagonists at the CP.

Impact of Mrp2 on the biliary excretion and intestinal absorption of furosemide, probenecid, and methotrexate using Eisai hyperbilirubinemic rats.

PURPOSE: This study assesses the impact of rat multidrug resistance-associated protein 2 (Mrp2) on the biliary excretion and oral absorption of furosemide, probenecid, and methotrexate using Eisai hyperbilirubinemic rats (EHBR). METHODS: To assess Mrp2-mediated biliary excretion, rats received a 2-h intravenous infusion of furosemide, probenecid, or methotrexate. Blood and bile samples were collected at specified intervals. To assess Mrp2's impact on oral absorption, rats received furosemide, probenecid, or methotrexate orally at 5 mg/kg. Jugular and portal blood samples were obtained at timed intervals. All samples were analyzed by LC-MS/MS. Pharmacokinetic parameters were estimated using WinNonlin and standard pharmacokinetic equations. RESULTS: Thirty seven- and 39-fold reductions in biliary clearance were observed in EHBR as compared to control rats for probenecid and methotrexate, respectively. Biliary clearance was comparable between EHBR and control rats for furosemide. In all cases, no significant difference in absorption was observed between EHBR and control rats. CONCLUSIONS: This study provides the first evidence that Mrp2 mediates the biliary excretion of probenecid but not furosemide. Additionally, Mrp2 apparently has a less profound impact on intestinal absorption than biliary excretion of its substrates. Furthermore, alteration in systemic clearance in EHBR indicates that a potential compensatory mechanism may occur in EHBR.

Rapid internal acyl migration and protein binding of synthetic probenecid glucuronides.

Internal acyl migration reactions of drug 1-O-acyl-beta-D-glucopyranuronates (1beta-acyl glucuronides) are of interest because of their possible role in covalent binding to proteins and consequent adverse effects. The reactivity of the synthetic probenecid 1beta-acyl glucuronide (PRG), the principal metabolite of probenecid (PR) in humans, has been investigated in terms of acyl migration, hydrolysis, and covalent binding to proteins in phosphate buffer (pH 7.4) and human plasma at 37 degrees C. PRG primarily degraded by acyl migration according to apparent first-order kinetics and the 2-, 3-, and 4-acyl isomers sequentially appeared as both alpha- and beta-anomeric forms. In addition, small amounts of PRG and extremely labile 1alpha-acyl isomer existed in the equilibrated mixture favoring the 2alpha/beta-acyl isomer, that provided significant information regarding the mechanism of acyl migration. All of the positional isomers and anomers were characterized using preparative HPLC and NMR spectroscopy. Acyl migration was observed to predominate over hydrolysis in both media although the extent of hydrolysis in plasma was larger than that in the buffer. The overall degradation half-lives (h) in the buffer and plasma were 0.27 +/- 0.003 and 0.17 +/- 0.007, respectively. The covalent binding rapidly proceeded mainly via the Schiff's base mechanism and reached a plateau after 2 h of incubation. The maximal binding was 146 +/- 4.8 pmol/mg of protein, and ca. 10% of the initial concentration of PRG. These results indicated that PRG is most labile and susceptible to acyl migration of all the drug acyl glucuronides reported to date in the physiological conditions, and highly reactive to plasma proteins, that could provide a possible explanation for the immunologically based adverse effects of PR.

The role of probenecid-sensitive organic acid transport in the pharmacokinetics of N-methyl-D-aspartate receptor antagonists acting at the glycine(B)-site: microdialysis and maximum electroshock seizures studies.

The purpose of the present study was to determine whether the probenecid-sensitive organic acid transporter is responsible for the short duration of action of a new group of N-methyl-D-aspartate receptor glycine(B)-site antagonists, MRZ 2/570, 2/571, and 2/576. A prolongation of their anticonvulsant activity from 60 to 180 to 240 min, was found in mice after pretreatment with probenecid (200 mg/kg i.p.). Microdialysis studies in rats showed that this is likely due to a change in central nervous system concentrations of these drugs because cotreatment with probenecid caused an increase in the brain extracellular fluid half-life (0.5- to 4-fold) and the brain area under the curve (1.8- to 3.6-fold). In serum the half-life of MRZ 2/576 (30 mg/kg) was also increased by coadministration of probenecid from 15.6 +/- 1.3 to 40.6 +/- 6.0 min. At steady state (MRZ 2/576, 20 mg/kg/h i.v.), brain extracellular fluid concentration was elevated 2.5-fold by concomitant administration of probenecid. These results clearly show that these glycine(B)-site antagonists are rapidly cleared from the systemic circulation and the central nervous system by the probenecid-sensitive organic acid transport system. Moreover, the present data show that MRZ 2/570, 2/571, and 2/576 reach the brain in concentrations (1.34-2.32 microM) above the range of their in vitro potencies at the glycine site of the N-methyl-D-aspartate receptor (0.1-1.0 microM).

Complete two-dimensional separation for analysis of acidic compounds in plasma using column-switching reversed-phase liquid chromatography.

A complete two-dimensional separation technique for the determination of acidic compounds in plasma was developed by using column-switching reversed-phase liquid chromatography. This technique was based on solute peak enrichment at the top of the second column during heart-cutting and an ion-pair chromatographic separation in the second column using tetrabutylammonium ion, where different separation modes in the first and second columns and solute peak enrichment at the top of the second column during heart-cutting were achieved coincidentally. Retention behaviors of two solutes, zidovudine-beta-D-glucuronide (AZT-beta-D-Gluc) and probenecid, in the first and second column and solute peak enrichment at the top of the second column were investigated for establishment of the system. Different retention behaviors of the solutes in the first and second column, which were evaluated by changes in capacity factor versus acetonitrile concentration in the mobile phases, and peak enrichment could be accomplished by using ion-pair chromatography in the second column. System suitability was confirmed by assessing the number of theoretical plates (N) of the second column for the solutes after heart-cutting. The N values in the second column after column switching were almost same as those in the case that the solutes were directly injected onto the second column. These results indicate that complete two-dimensional separation should be achieved by using this system. Furthermore, this technique was applied to method development for the determination of AZT-beta-D-Gluc and probenecid in rat plasma. The peaks of each analyte in the plasma extract obtained by deproteinization were well separated from those of endogenous substances, and easy determination of the analytes could be accomplished at the ng/ml level only by changing the acetonitrile concentration in the mobile phases.

Quantitative evaluation of brain distribution and blood-brain barrier efflux transport of probenecid in rats by microdialysis: possible involvement of the monocarboxylic acid transport system.

This study was performed to evaluate quantitatively the brain distribution and the efflux transport across the blood-brain barrier of probenecid, using in vivo microdialysis and in situ brain perfusion techniques. The brain interstitial fluid (ISF)-to-plasma cerebrospinal fluid (CSF)-to-plasma and brain tissue-to-plasma unbound concentration ratios of probenecid at steady state were less than unity, which suggests restricted distribution in the brain. An uphill concentration gradient from ISF to plasma and a downhill concentration gradient from CSF to ISF were observed. Kinetic analysis revealed that the efflux clearance from brain ISF to plasma (0.0373 ml/min/g brain) was significantly greater than the influx clearance from plasma to brain (0.00733 ml/min/g brain). The ratio of the ISF concentration (Cisf) to the plasma unbound concentration (Cp,f) of probenecid was increased 2- to 3-fold by salicylate (3.7 mM) and benzoate (3.6 mM), which are accepted as substrates of the monocarboxylic acid transport system, compared with the same ratio for the control. In addition, the ratio Cisf/Cp,f was increased by treatment with N-ethylmaleimide, a sulfhydryl-modifying agent, whereas p-aminohippuric acid and choline did not produce increasing effects on Cisf/Cp,f. These data suggest that the restricted distribution of probenecid in the brain may be ascribed to efficient efflux from the brain ISF, which may be regulated by the monocarboxylic acid transport system at a relatively high ISF concentration.

Pharmacokinetics of bacampicillin in equids.

Bacampicillin hydrochloride is an ester prodrug that is hydrolyzed to ampicillin after its absorption from the gastrointestinal tract. It was administered intragastrically at a dose rate of 13.5 mg/kg of body weight to ponies and horses, and was highly bioavailable (F = 41.0%), compared with other penicillins in adult horses. The high peak ampicillin plasma concentration of 6.1 +/- 0.5 micrograms/ml achieved and persistence of the antibiotic at concentration of 0.3 +/- 0.1 microgram/ml 6 hours after its intragastric administration, suggest that bacampicillin hydrochloride may reach suitable bactericidal concentrations for treatment of infections caused by susceptible microorganisms. In a separate experiment, dichlorvos, an organophosphate compound that inhibits some of the esterase activity in plasma, was administered orally to the same animals at a dose rate of 40 mg/kg, followed by intragastric administration of bacampicillin hydrochloride at a dose of 13.5 mg/kg. Plasma pseudocholinesterase and erythrocyte acetylcholinesterase activities were reduced to < 5% of reference (predichlorvos) values after dichlorvos administration. However, rate of hydrolysis of bacampicillin into ampicillin was not affected. Consequently, the disposition and fate of bacampicillin when administered intragastrically 1 day after dichlorvos administration were similar to the values obtained after administration of bacampicillin alone. Intragastric coadministration of probenecid at a dose rate of 75 mg/kg and bacampicillin at 13.5 mg/kg limited absorption of the antibiotic from the gastrointestinal tract. This suggests existence of a common transport mechanism for bacampicillin and probenecid in the gastrointestinal wall, and precludes use of this combination for treatment. The bioavailable fraction of ampicillin after combination treatment indicated prolonged residence time in the plasma, presumably as a consequence of reduced renal tubular secretion.

Screening of potential transport systems for methyl mercury uptake in rat erythrocytes at 5 degrees by use of inhibitors and substrates.

The current study was designed to screen the potential transport systems for methyl mercury (MeHg) uptake by isolated erythrocytes from rats at 5 degrees. Several inhibitors and substrates were used to test which potential transport system might be involved in MeHg uptake. Probenecid was used to test the organic anion transport system, valinomycin was used to test the effect of the membrane potential, D-glucose and cytochalasin B were used to test the facilitated diffusive D-glucose transport system and colchicine and vinblastine were used to test the microtubule system. The effects of Ca++, Mg++ and Na+ on MeHg uptake have been examined. Ouabain, ATP and glucose were used to test the active transport system, cysteine for the cysteine-facilitated transport system, glycine for system Gly, DL-methionine for system L, and MeHgCl and 4',4-diisothiocyano-2',2-stilbenedisulfonic acid (DIDS) for the Cl- ion transport system. The results showed that MeHg uptake might be involved in the following transport systems at 5 degrees: 1) organic anion transport system; 2) facilitated diffusive D-glucose transport system; 3) cysteine-facilitated transport system; 4) Cl- ion transport system. Moreover, the transport systems for MeHg uptake were sensitive to the membrane potential. Although the mechanisms of interaction of transport systems have not been fully clarified, evidence has been presented which support the existence of several simultaneous transport systems for MeHg uptake.

Sex-dependent and independent renal excretion of nilvadipine metabolites in rat: evidence for a sex-dependent active secretion in kidney.

1. To clarify the mechanism of sex-dependent and independent kidney secretion of major nilvadipine metabolites (3, 7) in rat, renal clearance corrected for protein binding and glomerular filtration rate (GFR) was measured in both sexes. The effect of probenecid, an inhibitor of organic anion transport, on these measurements was also investigated. 2. Clear sex-dependent active secretion was observed in the renal excretion of 3 (3-carboxylic acid pyridine derivative). In the female rat, 3, clearance was approximately 32-fold greater than GFR and was markedly decreased by probenecid. Conversely, in the male rat, renal clearance of 3 was only a fraction of GFR and was unaffected by probenecid. 3. Sex-independent active secretion was observed in the renal excretion of 7 (5-carboxylic acid pyridine derivative). In both sexes of rat 7 clearance was about 22-fold greater than GFR and was markedly reduced by probenecid. 4. A clear presence of sex-dependent and independent active secretion mechanisms in the kidney has been demonstrated in rat. The female rat is able to eliminate 3 and 7 in urine by an active secretion mechanism that is inhibited by probenecid. In the male rat, a transport mechanism for 7 is present, but either lacks or is apparently inactive for 3.

Should probenecid be used to reduce the dicloxacillin dosage in orthopaedic infections? A study of the dicloxacillin-saving effect of probenecid.

Reduction in the dosage of dicloxacillin from 500 mg to 250 mg 3 times a day would mean lowering of costs and less side-effects in orthopaedic infections. In this cross-over study, the serum concentrations of dicloxacillin were measured in 9 patients after administration of dicloxacillin 500 mg 3 times a day (dicloxacillin 500 mg) and after co-administration of 250 mg dicloxacillin and 250 mg probenecid 3 times per day (dicloxacillin 250 mg+probenecid 250 mg). Concentrations were measured every hour after the tablet intake. The mean maximum serum concentrations of dicloxacillin were 17.1 micrograms/ml (dicloxacillin 500 mg) and 12.2 micrograms/ml (dicloxacillin 250 mg+probenecid 250 mg), respectively (P < 0.05). Serum concentrations above 3 micrograms/ml were obtained during 285 min. in both regimes, but the individual variations were biggest during in the dicloxacillin 250 mg+probenecid 250 mg treatment. Serum concentrations above 5 micrograms/ml were in mean measured during 228 min. (dicloxacillin 500 mg) and 190 min. (dicloxacillin 250 mg+probenecid 250 mg), respectively (P < 0.05). The clinical significance of these findings is being discussed. In theory, treatment with dicloxacillin 250 mg+probenecid 250 mg may be as sufficient as dicloxacillin 500 mg.