Pharmacokinetic evaluation of drug interactions with anti-HIV drugs, II: Effect of 2',3'-dideoxyinosine (ddI) on zidovudine kinetics in monkeys.

The pharmacokinetic basis of a drug interaction between zidovudine (AZT) and 2',3'-dideoxyinosine (ddI) was investigated in normal monkeys. Five animals received 20 mg/kg of AZT intragastrically in the absence and presence of ddI. ddI was administered intravenously to produce steady-state ddI plasma concentrations for 30 min. Plasma and urine samples were analyzed for AZT, its major glucuronide metabolite, GAZT, and ddI by high-performance liquid chromatography (HPLC). Resultant AZT and GAZT concentration data were analyzed by noncompartmental methods. Statistical analysis indicated no differences in AZT's apparent total clearance, apparent volume of distribution at steady-state, and elimination half-life due to ddI, however, the mean apparent total clearance decreased from 2.92 to 1.67 L/h/kg, and the mean apparent volume of distribution at steady-state decreased from 5.79 to 3.43 L/kg in the presence of ddI. Incomplete urine collections in most animals prevented conclusions from being made about ddI's effect on renal elimination parameters. Nonetheless, the urinary GAZT/AZT ratio, a parameter not influenced by incomplete urine collection, was significantly reduced in the presence of ddI. Although additional studies will be useful to characterize the full importance of the interaction, there is evidence to suggest that both renal and metabolic elimination of AZT and renal elimination of GAZT may be inhibited by ddI.

Pharmacokinetics of phenobarbital following single and repeated doses.

Serum levels of phenobarbital, and also urinary excretion of phenobarbital and p-hydroxyphenobarbital, were examined after single and repeated oral doses of phenobarbital to three male subjects. Serum levels of phenobarbital at steady state were approximately ten times as high as those after a single dose. The overall elimination rate constant for loss of phenobarbital from serum, Kel, was significantly reduced after repeated doses, and Cmax infinity values calculated from single-dose data poorly predicted observed Cmax infinity values. Five-day urinary excretion of phenobarbital and p-hydroxyphenobarbital accounted for 16 and 21 per cent, respectively, of the initial dose. Due to extensive drug accumulation, 83 per cent of the final dose was excreted in five-day urine as phenobarbital and 85 per cent, as p-hydroxyphenobarbital. Comparison of plasma and renal clearances indicated that the rate of phenobarbital metabolism was reduced owing to repeated dosing, while the rate of urinary excretion of parent drug was unchanged.

Bioavailability of oral and intramuscular phenobarbital.

The absorption of phenobarbital was compared in healthy adult subjects after oral and intramuscular therapeutic doses. Serum levels of phenobarbital were determined for 21 days after dosing by means of radioimmunoassay. Serum levels were similar from both dosage routes, with peak levels occurring at 1-3 hours after dosing and then declining slowly with an elimination half-life of about 90 hours. The overall efficiency of phenobarbital absorption from intramuscular doses was approximately 80 per cent of that from equivalent oral doses. Except in cases where oral dosing is not appropriate, there is no clinical advantage in giving phenobarbital intramuscularly to adult patients.

An extensive exploration of the rat brain for sites mediating prostaglandin-induced hyperthermia.

Experiments designed to ascertain the site or sites within the brain where prostaglandins act to evoke hyperthermia were carried out. At an ambient temperature of 20 degrees C, microinjections of PGE1 (50-100 ng) or saline vehicle were made in restrained rats. The 272 sites tested formed a matrix encompassing much of the subcortical tissue rostral to the medulla. The sensitivity of each site to PGE1 was evaluated on the basis of the dose required to elicit a criterion level of effect and on the intensity of the effect evoked by a fixed dose. Injections of PGE1 into a sensitive site typically produced a rapidly developing, short lasting, monophasic rise in body temperature. Saline injected at these sites produced no initial hyperthermia. Almost all of the active sites were located within the anterior hypothalamic/preoptic region (AH/PO). The sites of greatest reactivity were found in the ventral aspect of the tissue lying between the anterior commissure and the optic chiasm. These data suggest that the AH/PO region is an important and probably the only supramedullary site of PGE1 action in the rat brain.

Diurnal oscillations of CSF valproate in monkey.

Valproate (VPA) was administered to four rhesus monkeys by constant-rate intravenous infusion for two weeks under controlled conditions. Plasma and CSF samples were collected for a period of 27 hours at 3-hour intervals during steady-state and post-infusion periods. The mean correlation coefficient between total plasma and CSF VPA concentrations was found to be 0.78 +/- 0.09. The CSF VPA levels reflected the changes in free VPA in plasma but the two were not equivalent. Diurnal fluctuations in CSF VPA concentration were similar to those found in plasma but the inter-animal variation was greater in CSF than in plasma.

Plasma protein binding interaction between valproic and salicylic acids in rhesus monkeys.

The effects of three levels of salicylic acid on the steady-state plasma concentrations of free and total valproic acid were examined in catheterized rhesus monkeys. Valproate was infused intravenously for a total of 41 hr, and salicylate was added after the first 8 hr. The three salicylate infusions were randomly assigned to each monkey. Valproate free fraction was determined by equilibrium dialysis. Statistically significant increases in valproate free fraction and total body clearance were observed after addition of salicylic acid. The increase in valproate clearance was positively correlated with the molar ratio of salicylate to valproate steady-state plasma concentrations. There was no significant change in valproate free concentration after salicylate treatment. The proposed mechanism of this in vivo interaction includes plasma protein binding displacement with no change in valproate intrinsic clearance.

Pharmacokinetic evaluation of drug interactions with zidovudine. I: Probenecid and zidovudine in monkeys.

Pharmacokinetic evaluation of a drug interaction between zidovudine (AZT) and probenecid was conducted in monkeys. Six animals received 20 mg/kg of AZT as single intragastric (ig) and iv doses in the absence and presence of 50 mg/kg of probenecid administered ig. Plasma concentrations of AZT and its 5'-glucuronide metabolite (AZTG) were quantitated for 12 h by HPLC. Amounts of AZT and AZTG in urine were also measured, as were probenecid plasma concentrations. Non-compartmental methods were used to obtain pharmacokinetic parameters for AZT and AZTG. In the presence of probenecid, the total clearance of AZT decreased by 50%, renal clearance decreased, and elimination half-life increased. The volume of distribution at steady-state and systemic bioavailability of AZT were not significantly altered by probenecid. The areas under the plasma concentration-time curves and terminal half-lives of AZTG were increased, and renal clearances of AZTG were decreased. The alterations in AZT and AZTG pharmacokinetic parameters are consistent with inhibition of metabolism and renal tubular secretion by probenecid. Since AZT was administered by both oral and iv routes, clearance, volume of distribution, and bioavailability parameters were independently determined. Based on data reported for humans on the zidovudine-probenecid interaction, monkeys appear to be appropriate animal models for the evaluation of zidovudine drug interactions.

Effects of humidity and temperature on in vitro dissolution of carbamazepine tablets.

The rate and extent of dissolution of various approved marketed carbamazepine (CBZ) tablets exposed to 33, 52, 75, and 97% relative humidities at both room temperature and 40 degrees C, and saturated water vapor at room temperature were compared to fresh unstressed tablets. The dissolution data indicate that exposure of CBZ tablets to high humidity and temperature can have a profound effect on tablet disintegration and dissolution. The dissolution rates of some batches of CBZ products exposed to 97% humidity at 40 degrees C or saturated water vapor at room temperature were drastically reduced in only 6-7 days.

Bioavailability of regular and controlled-release chlorpheniramine products.

The bioavailability of chlorpheniramine regular-release versus controlled-release products was compared using 15 human subjects. The dosage forms evaluated were an 8-mg barrier coated-bead capsule, an 8-mg repeat action tablet, two 4-mg tablets, and 4- and 8-mg syrups. Single doses of each product were administered orally in a 5-way crossover study, plasma samples were collected at specific time intervals, and chlorpheniramine levels assayed by HPLC. Pharmacokinetic analysis was based on a two-compartment open model. The average plasma elimination half-life of chlorpheniramine was calculated to be approximately 18.3 hr. The controlled-release products gave a higher Cmax than the 4-mg syrup, but less than two 4-mg tablets. The controlled-release products also extended the time necessary to attain peak drug levels compared to the 4- and 8-mg syrups. The area under the curve (AUC) data for the controlled-release products was not equivalent to equal amounts of the regular-release products. The study indicated that while the controlled-release chlorpheniramine products were successful in prolonging the time course of absorption, this was at the expense of incomplete bioavailability of the drug.

Analytical methods validation: bioavailability, bioequivalence and pharmacokinetic studies. Conference report.

This is a summary report of the conference on Analytical Methods Validation: Bioavailability, Bioequivalence and Pharmacokinetic Studies. The conference was held from December 3 to 5, 1990 in the Washington, DC area and was sponsored by the American Association of Pharmaceutical Scientists, US Food and Drug Administration, Federation International Pharmaceutique, Health Protection Branch (Canada) and Association of Official Analytical Chemists. The purpose of the report is to represent our assessment of the major agreements and issues discussed at the conference. The report is also intended to provide guiding principles for validation of analytical methods employed in bioavailability, bioequivalence and pharmacokinetic studies in man and animals. The objectives of the conference were: 1. To reach a consensus on what should be required in analytical methods validation and the procedures to establish validation; 2. To determine processes of application of the validation procedures in the bioavailability, bioequivalence and pharmacokinetic studies; 3. To develop a report on analytical methods validation (which may be referred to in developing future formal guidelines). Acceptable standards for documenting and validating analytical methods with regard to processes, parameters or data treatments were discussed because of their importance in assessment of pharmacokinetic, bioavailability and bioequivalence studies. Other topics which were considered essential in the conduct of pharmacokinetic studies or in establishing bioequivalency criteria, including measurement of drug metabolites and stereoselective determinations, were also deliberated.

Perspectives on microsampling: DBS.

The concept of microsampling and particularly the dried blood spot methodology has been widely known to the scientific community for considerable time. Yet, there is no formal standard approach available to date for the pharmaceutical community to benefit from, in its regulatory interactions. This article discusses the various aspects of these issues and provides a framework within which a standard set of procedures can be adopted.

US FDA/EMA harmonization of their bioanalytical guidance/guideline and activities of the Global Bioanalytical Consortium.

The 2011 annual conference of the American Association of Pharmaceutical Scientists, held in Washington DC, USA, hosted a roundtable entitled: 'Update of the US FDA/European Medicines Agency (EMA) harmonization of their bioanalytical guidance - Global Bioanalytical Consortium activity and impact on small and large molecules.' The roundtable was initiated with a presentation from CT Viswanathan on the history of the revision of the FDA guideline on bioanalytical method validation. It was followed by a presentation by Jan Welink who presented an update on the final European Medicines Agency guideline on bioanalytical method validation with relevance to ongoing harmonization efforts. The final presentation was by Fabio Garofolo on the progress of the Global Bioanalytical Consortium harmonization teams for small and large molecules. Brian Booth and Sam Haidar of the FDA updated the audience on the status of the revision of the FDA bioanalytical guidance. The roundtable was moderated by Stephen Lowes.

2012 white paper on recent issues in bioanalysis and alignment of multiple guidelines.

Over 400 professionals representing pharmaceutical companies, CROs, and multiple regulatory agencies participated in the 6th Workshop on Recent Issues in Bioanalysis (WRIB). Like the previous sessions, this event was in the format of a practical, focused, highly interactive and informative workshop aiming for high-quality, improved regulatory compliance and scientific excellence. Numerous 'hot' topics in bioanalysis of both small and large molecules were shared and discussed, leading to consensus and recommendations among panelists and attendees representing the bioanalytical community. The major outcome of this year's workshop was the noticeable alignment of multiple bioanalytical guidance/guidelines from different regulatory agencies. This represents a concrete step forward in the global harmonization of bioanalytical activities. The present 2012 White Paper acts as a practical and useful reference document that provides key information and solutions on several topics and issues in the constantly evolving world of bioanalysis.

2011 White paper on recent issues in bioanalysis and regulatory findings from audits and inspections.

The 5th Workshop on Recent Issues in Bioanalysis (WRIB) was organized by the Calibration and Validation Group as a 2-day full immersion workshop for pharmaceutical companies, CROs and regulatory agencies to discuss, review, share perspectives, provide potential solutions and agree upon a consistent approach to recent issues in the bioanalysis of both small and large molecules. High quality, better compliance to regulations and scientific excellence are the foundation of this workshop. As in the previous editions of this significant event, recommendations were made and a consensus was reached among panelists and attendees, including industry leaders and regulatory experts representing the global bioanalytical community, on many 'hot' topics in bioanalysis. This 2011 White Paper is based on the conclusions from this workshop, and aims to provide a practical reference guide on those topics.

Regulatory observations in bioanalytical determinations.

The concept of measuring analytes in biological media is a long-established area of the quantitative sciences that is employed in many sectors. While academic research and R&D units of private firms have been in the forefront of developing complex methodologies, it is the regulatory environment that has brought the focus and rigor to the quality control of the quantitative determination of drug concentration in biological samples. In this article, the author examines the regulatory findings discovered during the course of several years of auditing bioanalytical work. The outcomes of these findings underscore the importance of quality method validation to ensure the reliability of the data generated. The failure to ensure the reliability of these data can lead to potential risks in the health management of millions of people in the USA.

Workshop report and follow-up--AAPS Workshop on current topics in GLP bioanalysis: Assay reproducibility for incurred samples--implications of Crystal City recommendations.

The Conference Report of the 3rd AAPS/FDA Bioanalytical Workshop (Crystal City III) endorsed the concept that assay methods supporting bioanalytical data in submissions must demonstrate assay reproducibility by using incurred samples. The present Workshop was convened to provide a forum for discussion and consensus building about incurred sample assay reproducibility for both nonclinical and clinical studies. Information about current regulatory perspectives on incurred sample reanalysis (ISR) was presented, implications of ISR for both large and small molecules were discussed, and the steering committee put forth recommendations for performing ISR. These recommendations from the Workshop, along with the subsequent evolution of approaches leading to a robust ISR program, may be used by scientists performing bioanalytical assays for regulated studies to provide additional confirmation of assay reproducibility for incurred samples.

Quantitative bioanalytical methods validation and implementation: best practices for chromatographic and ligand binding assays.

The Third AAPS/FDA Bioanalytical Workshop, entitled "Quantitative Bioanalytical Methods Validation and Implementation: Best Practices for Chromatographic and Ligand Binding Assays" was held on May 1-3, 2006 in Arlington, VA. The format of this workshop consisted of presentations on bioanalytical topics, followed by discussion sessions where these topics could be debated, with the goal of reaching consensus, or identifying subjects where addition input or clarification was required. The discussion also addressed bioanalytical validation requirements of regulatory agencies, with the purpose of clarifying expectations for regulatory submissions. The proceedings from each day were reviewed and summarized in the evening sessions among the speakers and moderators of the day. The consensus summary was presented back to the workshop on the last day and was further debated. This communication represents the distillate of the workshop proceedings and provides the summary of consensus reached and also contains the validation topics where no consensus was reached.

Effect of central cholinergic blockade on the hyperthermia evoked by prostaglandin E1 injected into the rostral hypothalamus of the rat.

The possibility of a cholinergic involvement in the hyperthermic action produced by the injection of prostaglandin E1 (PGE1) into the anterior hypothalamic/preoptic region of the rat brain was examined. Intracerebral injection of either atropine or mecamylamine prior to the injection of PGE1 failed to attenuate the PGE1-induced hyperthermia. Both atropine and mecamylamine by themselves produced a rise in colonic temperature. It thus seems unlikely that PGE1 evokes hyperthermia in the rat by releasing endogenous acetylcholine at muscarinic or nicotinic synapses in the rostral hypothalamus. The possibility that PGE1 acts by inhibiting the release of acetylcholine within this region requires additional investigation.