F. Varin

Determination of midazolam and two of its metabolites in human plasma by high-performance liquid chromatography.

A high-performance liquid chromatographic assay coupled with UV detection (254 nm) has been developed for the determination of midazolam and two of its hydroxylated metabolites, 1-hydroxymidazolam (1-OH) and 4-hydroxymidazolam (4-OH), in human plasma. Following a novel solid-phase extraction procedure, midazolam and its metabolites are well recovered from plasma. The analytes were extracted with C1 cartridges and the extracts were evaporated to dryness. The dry residues were dissolved in 200 microliters of mobile phase [0.02 M ammonium phosphate monobasic buffer-methanol-acetonitrile (60:35:5, v/v) (300 ml), 600 microliters of 0.2 M tetrabutylammonium bromide solution, adjusted to a pH* (apparent pH) of 4.10]. The separation of the analytes was performed on a Spherisorb C8 column (10 cm x 4.6 mm I.D.) maintained at 30 degrees C. The mobile phase was pumped at a flow-rate of 1.5 ml/min. The method has a lower limit of quantitation of 15 ng/ml of plasma for midazolam and proved to be reproducible (inter-assay precision 5.4%) and accurate (94 +/- 5%) over the therapeutic range of concentrations.

High-performance liquid chromatography-electrochemical detection of vecuronium and its metabolites in human plasma.

A high-performance liquid chromatographic assay coupled with electrochemical detection has been developed for the determination of vecuronium and its three putative deacetylated metabolites in human plasma. A novel solid-phase extraction procedure allowed good recovery of both vecuronium and its metabolites, together with ease and speed of execution. This method was sensitive, reproducible and accurate over the therapeutic range of concentrations of vecuronium and its metabolites, and was applied successfully to a study of the pharmacokinetics of vecuronium in anaesthetized patients.

Determination of succinylcholine in human plasma by high-performance liquid chromatography with electrochemical detection.

An alternative HPLC method for the quantification of the depolarizing neuromuscular blocking agent succinylcholine in human plasma is described. Drug spiked plasma and patient plasma samples were extracted using a C1 solid-phase cartridge. Succinylcholine was separated on a Cyano column and quantitated using electrochemical detection at a potential of 450 mV and 750 mV. Mobile phase consisted of a mixture of phosphoric acid-acetonitrile-methanol (45:35:25) adjusted to an apparent pH of 5. Standard curves for the quantitation were linear in the range of 250-8000 ng/ml. Between-day and within-day relative standard deviations were 5.1% and 1.7%, respectively. Mean drug recovery and accuracy was 68% and 104%, respectively.

Determination of atracurium and laudanosine in human plasma by high-performance liquid chromatography

A high-performance liquid chromatographic method coupled with fluorometric detection has been developed for the determination of atracurium and its major end-product laudanosine in human plasma. The method enables good separation of atracurium from its metabolites after direct precipitation of plasma proteins. The assay is sensitive, reproducible and linear for atracurium concentrations ranging from 31.25 to 8000 ng/ml. In a clinical setting, drugs commonly administered during anesthesia did not interfere with the assay. This method provides a simple and time-saving alternative to existing methods.

Determination of opiates and other basic drugs by high-performance liquid chromatography with electrochemical detection☆

A procedure is described for the extraction and determination of morphine (M), hydromorphone (HM), codeine (C) and metoclopramide (MCP) present in human plasma. The drugs are separated by reversed-phase liquid chromatography and detected amperometrically at a glassy carbon electrode. The method provides high sensitivity and selectivity and has been used successfully in bioavailability studies.

High-performance liquid chromatographic assays with fluorometric detection for mivacurium isomers and their metabolites in human plasma

Two high-performance liquid chromatographic assays coupled with fluorometric detection have been developed for the determination of mivacurium isomers (trans-trans, cis-trans and cis-cis) and their monoester and alcohol metabolites in human plasma. A novel solid-phase extraction procedure allowed good recovery of the mivacurium isomers (mean 98%) and their monoester metabolites (mean 83%), whereas the alcohol metabolites were analyzed after direct precipitation of plasma proteins. For all analytes, these assays proved to be sensitive (LOQ 3.9-15.6 ng/ml), reproducible (C.V. < 15%) and accurate (> 94%) over the therapeutic range of concentrations of mivacurium and its metabolites. These two methods were applied successfully to a pharmacokinetic study of mivacurium after a bolus dose of 0.15 mg/kg in anesthetized patients.

Determination of neostigmine in human plasma and cerebrospinal fluid by high-performance liquid chromatography with ultraviolet detection.

A high-performance liquid chromatographic assay coupled with ultraviolet detection has been developed for the determination of neostigmine in human plasma and cerebrospinal fluid. A novel solid-phase extraction procedure was first used for this analyte and allowed good recovery (89+/-4.5%) together with ease and speed of execution. The method was sensitive, reproducible (C.V.<4.5%) and accurate (100+/-6.6%) over the range 2.6-167.0 ng/ml neostigmine concentrations in plasma or cerebrospinal fluid, and was applied successfully to study the pharmacokinetics of neostigmine in patients suffering from chronic postoperative abdominal pain.

Dose-dependency of pharmacokinetic/pharmacodynamic parameters after intravenous bolus doses of cisatracurium

BACKGROUNDnPharmacokinetic/pharmacodynamic (PK/PD) parameters of neuromuscular blocking agents (NMBAs) are generally assumed to be dose-independent. To our knowledge, there are very few clinical reports where the PK/PD parameters of a NMBA were derived separately for each dose group during a formal dose-ranging study. The primary objective of this study was to challenge a potential dose-dependency of cisatracurium PK/PD parameters by conducting a well-controlled experimental study.nnnMETHODSnEight dogs were anaesthetized with pentobarbital and mechanically ventilated. Two doses of cisatracurium (1.5xED(95) and 6xED(95)) were administered in a randomized cross-over design after an appropriate washout period. Neuromuscular function was monitored using train-of-four (TOF) stimulation. Arterial blood was sampled continuously for the first minute after cisatracurium injection and at frequent intervals thereafter. Cisatracurium plasma concentrations were determined by high performance liquid chromatography analysis. PK/PD modelling of individual data sets was performed with NONMEM using a non-parametric approach and a descriptive sigmoid E(max) model.nnnRESULTSnCisatracurium PKs were linear over the dose range studied. Using non-parametric PK/PD analysis, mean values for plasma-effect compartment equilibration delay (k(e0)) were 0.0600 vs 0.1278 min(-1) (P<0.05) and sensitivity (EC(50)) were 323 vs 235 ng ml(-1) (P<0.05) for the high and low doses, respectively.nnnCONCLUSIONSnA dose-dependent effect on the PK/PD parameters of cisatracurium has important clinical implications as an accurate estimate of the EC(50) is desirable. PK/PD parameters derived after intubating bolus doses of cisatracurium would be more reliable.

Early Pharmacokinetics of Midazolam

SummaryThe influence of sampling site and sampling schedule on the early pharmacokinetics of a lipophilic drug, midazolam, was studied in 6 anaesthetised patients following an intravenous bolus injection of 0.05 mg/kg. Arterial and venous blood samples were drawn simultaneously over a period of 180 minutes. An extensive sampling procedure was used during the initial distribution phase; specifically, 12 arterial and 4 venous samples within the first 2 minutes. Midazolam concentrations were measured by high-performance liquid chromatography. Compared with peak venous concentrations, peak arterial concentrations were 2-fold greater and were reached 1 minute earlier. Although arterial concentrations exceeded corresponding venous concentrations for up to 90 minutes, a significant arteriovenous difference was observed during the first 2 minutes only. Consequently, noncompartmental pharmacokinetic parameters (mean residence time, volume of distribution at steady-state, total body clearance) obtained from arterial and venous values did not differ significantly. In addition, similar results were obtained for pharmacokinetic parameters using either the complete (continuous sampling over 2 minutes) or limited (assuming 1- and 2-minute samples only) arterial data set. It is concluded that both the sampling site and the sampling schedule altered the initial plasma concentration-time profile of midazolam. Although neither of these factors had a strong influence on the overall pharmacokinetic parameters of midazolam, their potential effect on the pharmacokinetic/pharmacodynamic relationship of midazolam during the onset phase should be considered.

Studies on the pharmacokinetics of cisatracurium in anesthetized dogs: in vitro-in vivo correlations.

Cisatracurium undergoes primarily temperature and pH-dependent Hofmann elimination in humans. This study was conducted to describe the pharmacokinetics of cisatracurium in anesthetized dogs and determine whether its in vitro degradation rate in plasma is predictive of its in vivo elimination rate, as this is the case in humans. Nine dogs were anesthetized with pentobarbital and administered different bolus doses of cisatracurium in a randomized cross-over design. Arterial blood was collected at frequent intervals after each bolus injection. In vitro degradation rate (k(in vitro)) of cisatracurium was determined in each dog blank plasma. Plasma concentrations were determined by HPLC. Pharmacokinetic analyses were performed using two compartmental models assuming central or both central and peripheral elimination. Mean in vivo terminal elimination rate of cisatracurium (16.4 +/- 2.7 min) was twofold faster than mean in vitro degradation rate (32.9 +/- 3.7 min) in our dogs. Organ clearance was 6.12 +/- 1.69 mL/min.kg and accounted for 56 +/- 12% of the total body clearance. Apparent volume of distribution, an exit site-dependent parameter, averaged 212 or 184 mL/kg whether or not peripheral elimination was accounted for in the model. The in vitro rate of degradation in plasma is not of predictive value for the in vivo elimination rate of cisatracurium in anesthetized dogs. Organ clearance plays a more important role in the elimination of cisatracurium in dogs than in humans. Increased biliary excretion and/or presence of renal secretion are potential mechanisms that need to be explored.