Surfactant-assisted dispersive liquid-liquid microextraction combined with field-amplified sample stacking in capillary electrophoresis for the determination of mexiletine and lidocaine.

A sensitive method for the determination of mexiletine and lidocaine using surfactant-assisted dispersive liquid-liquid microextraction coupled with capillary electrophoresis was developed. Triton X-100 and dichloromethane were used as the dispersive agent and extraction solvent, respectively. After the extraction, mexiletine and lidocaine were analyzed using capillary electrophoresis with ultraviolet detection. The detection sensitivity was further enhanced through the use of field-amplified sample stacking. Under optimal extraction and stacking conditions, the calibration curves were linear over a concentration range of 0.05-1.00 µM for mexiletine and 0.03-1.00 µM for lidocaine. The limits of detection (signal-to-noise ratio of 3) were 0.01 and 0.01 µM for mexiletine and lidocaine, respectively. An approximately 1141- to 1250-fold improvement in sensitivity was observed for the two analytes compared with the injection of a standard solution without the surfactant-assisted dispersive liquid-liquid microextraction and field-amplified sample stacking procedures. This developed method was successfully applied to the determination of mexiletine and lidocaine in human urine and serum samples. Both precision and accuracy for urine and serum samples were less than 8.7 and 6.7%, respectively. The recoveries of the two analytes from urine and serum samples were 54.7-64.9% and 16.1-56.5%, respectively.

Mexiletine carbonyloxy beta-D-glucuronide: a novel metabolite in human urine.

1. The study was performed to isolate and characterize a glucuronic acid conjugate of mexiletine that releases mexiletine on acid hydrolysis from urine samples obtained from healthy volunteers following a single oral dose of mexiletine. 2. The [M-H]- ion of the isolated metabolite was observed at m/z 398 in the negative electrospray ionization mass spectrum. This mass number was 44 higher than that of the product generated when mexiletine is subjected to direct glucuronidation. In positive-ion mode, collision-induced dissociation of the quasimolecular ion [M+NH4]+, m/z 417, gave product ions at m/z 224, 180 and 58. These mass spectral data indicated that the metabolite contained a carbonyloxy moiety in its structure in addition to mexiletine and a glucuronic acid moiety. 3. The presence of this carbonyloxy moiety was further supported by the following chemical reactions. When the metabolite was hydrolysed with an aqueous solution of 1 M sodium hydroxide at room temperature, mexiletine was released, whereas the N-methoxycarbonyl derivative of mexiletine was obtained after treatment of the metabolite with methanolic sodium hydroxide solution. 4. The results indicated that the structure of the isolated metabolite was the N-carbonyloxy beta-D-glucuronic acid conjugate of mexiletine.

Pharmacokinetic and pharmacodynamic interaction between mexiletine and propafenone in human beings.

BACKGROUND AND OBJECTIVE: Mexiletine and propafenone are often used concomitantly and are metabolized by the same cytochrome P450 isozymes, namely CYP2D6, CYP1A2, and probably CYP3A4. Our objective was to study the potential pharmacokinetic and electrophysiological interactions between mexiletine and propafenone. METHODS: Fifteen healthy volunteers, 8 extensive metabolizers and 7 poor metabolizers of CYP2D6, received oral doses of mexiletine 100 mg two times daily from day 1 to day 8 and oral doses of propafenone 150 mg two times daily from day 5 to day 12. Interdose studies were performed at steady-state on mexiletine alone (day 4), mexiletine plus propafenone (day 8), and propafenone alone (day 12). RESULTS: In subjects in the extensive metabolizer group, coadministration of propafenone decreased oral clearances of R-(-)-mexiletine (from 41+/-11 L/h to 28+/-7 L/h) and S-(+)-mexiletine (from 43+/-15 L/h to 29+/-11 L/h) to an extent such that these values were no longer different between the extensive and the poor metabolizer groups. Propafenone coadministration also decreased partial metabolic clearances of mexiletine to hydroxymethylmexiletine, p-hydroxymexiletine, and m-hydroxymexiletine in extensive metabolizers by 71%, 67%, and 73%, respectively. In contrast, propafenone did not alter the kinetics of mexiletine enantiomers in subjects in the poor metabolizer group except for a slight decrease in the formation of hydroxymethylmexiletine. Pharmacokinetic parameters of propafenone were not changed during concomitant administration of mexiletine in subjects of either phenotype. Finally, electrocardiographic parameters (QRS duration, QTc, RR, and PR intervals) were not modified during the combined administration of the drugs. CONCLUSION: Propafenone is a potent CYP2D6 inhibitor that may cause an increase in plasma concentrations of coadministered CYP2D6 substrates.

Stereoselective disposition of the antiarrhythmic agent mexiletine during the concomitant administration of caffeine.

Caffeine consumption is extensive in industrialized countries and its role in drug-drug interactions is often overlooked. CYP1A2, the major cytochrome P450 isoform involved in the metabolism of caffeine, has also been implicated in the formation of N-hydroxymexiletine, the major metabolite of mexiletine. Therefore, the objective of this study was to assess the effects of a clinically relevant dosage of caffeine on the stereoselective disposition of mexiletine. Fourteen healthy volunteers--10 extensive metabolizers (EMs) and 4 poor metabolizers (PMs) of CYP2D6--received a single 200 mg oral dose of racemic mexiletine hydrochloride on two occasions (1 week apart): once by itself and once during administration of caffeine (100 mg four times daily). Serial blood and urine samples were collected and pharmacokinetic parameters were estimated. Although the total clearance of mexiletine was not significantly altered by the coadministration of caffeine in EMs and PMs, a stereoselective decrease (16% in EMs and 14% in PMs) in the urinary recovery of N-hydroxymexiletine from the R-(-)-enantiomer was observed. Also, the partial metabolic clearance of R-(-)-mexiletine to N-hydroxymexiletine glucuronide was reduced from 126 +/- 48 mL/min to 106 +/- 32 mL/min and 152.6 (73.4-196.2) mL/min to 109 (77-127) mL/min by the coadministration of caffeine in EMs and PMs, respectively. Consequently, the R/S ratio for urinary recovery and the partial metabolic clearance of mexiletine to N-hydroxymexiletine were 28% lower during the coadministration of caffeine. In conclusion, data obtained in this study indicate that coadministration of caffeine does not lead to clinically significant changes in mexiletine plasma concentrations. However, results obtained suggest that CYP1A2 is involved in the formation of N-hydroxymexiletine.

Determination of the enantiomeric composition of mexiletine and its four hydroxylated metabolites in urine by enantioselective capillary gas chromatography.

The enantiomers of mexiletine and four of its hydroxylated metabolites were directly separated by capillary gas chromatography using a heptakis(6-O-tert-butyl-dimethylsilyl-2,3-di-O-methyl)-beta- cyclodextrin column. The method was applied to the analysis of urine samples from cancer patients who were treated with racemic mexiletine as part of a study of the use of mexiletine in the relief of neuropathic pain. Samples analyzed before and after deconjugation of the urine with beta-glucuronidase/arylsulfatase showed a high stereoselectivity in the formation and conjugation of these compounds.

Pharmacokinetics of mexiletine enantiomers in healthy human subjects. A study of the in vivo serum protein binding, salivary excretion and red blood cell distribution of the enantiomers.

1. The disposition kinetics of serum free (unbound) and total mexiletine enantiomers were studied in 12 healthy subjects following oral administration of 200 mg racemic mexiletine hydrochloride. The disposition of the enantiomers of mexiletine in urine, saliva, and red blood cells was also examined. 2. The mean peak serum total mexiletine concentration of 217 +/- 69 ng/ml for R(-)-mexiletine was found to be significantly greater than a mean of 197 +/- 56 ng/ml for S(+)-mexiletine. The mean serum total R(-)-mexiletine concentrations were also found to be significantly greater than those for S(+)-mexiletine during the first 6 h following drug administration. The oral absorption, as well as the rapid and the terminal disposition kinetic parameters between the mexiletine enantiomers, were not significantly different. 3. Comparative in vitro serum protein binding of mexiletine enantiomers examined by ultrafiltration and equilibrium dialysis indicated a pH-dependent stereoselective binding of the enantiomers to serum proteins. A serum pH ranging from 6.3 to 9.4 was found to correlate with serum protein binding of the enantiomers from approximately 30-80% respectively. Within the same serum pH range, the serum free drug R(-)/S(+) ratios were found to decrease from 1.0 to 0.7 respectively. At serum pH7.4, a mean serum free fraction of 0.57 +/- 0.7 and 0.56 +/- 0.6 were observed for R(-) and S(+)-mexiletine respectively. 4. The overall mean saliva/serum-free mexiletine enantiomer area under the concentration-time curve ratios of 6.10 +/- 2.82 and 7.49 +/- 3.48 for R(-)- and S(+)-mexiletine respectively were found to be significantly different. The overall mean saliva R(-)/S(+) enantiomer ratio of 0.89 +/- 0.02 (mean +/- SE) over 48 h suggested a stereoselective disposition of the mexiletine enantiomers in saliva. 5. The mean mexiletine red blood cells to serum-free drug concentration ratios among 11 subjects studied were found to range from 0.6 to 1.4 for R(-)-mexiletine and from 0.6 to 1.8 for S(+)-mexiletine. The overall mean ratios of 0.85 +/- 0.06 and 0.84 +/- 0.08 (mean +/- SE) over 48 h for R(-)- and S(+)-mexiletine respectively were both slightly but significantly different from unity. This data together with an overall red blood cell mean R(-)/S(+)-mexiletine concentration ratio of 0.91 +/- 0.13 suggested a non-stereoselective and passive diffusion of the enantiomers into red blood cells. 6. The cumulative amounts of unchanged R(-)- and S(+)-mexiletine in the urine were found to be variable among the 12 subjects with a mean percent urinary recovery of 3.49 +/- 3.35% for R(-)-mexiletine and 3.68 +/- 3.94% for S(+)-mexiletine.

Direct analysis of the enantiomers of mexiletine and its metabolites in plasma and urine using an HPLC-CSP.

A high-performance liquid chromatographic assay has been developed for the quantification of the enantiomers of mexiletine and its four major metabolites, in plasma and in urine. Mexiletine and all metabolites were determined, after derivatization of mexiletine and its hydroxymetabolites, p-hydroxymexiletine and hydroxymethylmexiletine, using a Chiralpak AD chiral stationary phase, based on a carbamoyl derivative of amylose. o-phthalaldehyde was chosen as derivatization reagent to increase the sensitivity of detection, to achieve separation of all compounds in one chromatographic system, and to avoid interferences.

Role of polymorphic debrisoquin 4-hydroxylase activity in the stereoselective disposition of mexiletine in humans.

It was reported previously that mexiletine undergoes stereoselective disposition in humans and that formation of three of its major metabolites co-segregates with polymorphic debrisoquin 4-hydroxylase (CYP2D6) activity. In this study, the hypothesis was tested that the CYP2D6-mediated oxidation pathways of mexiletine are responsible for the stereoselective disposition of the racemate in humans. Fourteen healthy subjects (10 extensive metabolizers [EMs] and 4 poor metabolizers [PMs]) participated in this study. They received a single 200-mg oral dose of racemic mexiletine hydrochloride on two occasions: once alone and once during administration of low-dose quinidine (50 mg four times a day). Blood and urine samples were obtained over 48 hr after the administration of mexiletine and analyzed by a stereoselective high-performance liquid chromatography assay. As reported previously, RS-mexiletine disposition was altered by a genetically determined (PM) or drug-induced (quinidine) decrease in CYP2D6 activity. In contrast, R/S ratio of the apparent total and nonrenal clearances of mexiletine and the R/S ratio of the urinary recovery of both enantiomers were similar in EMs and PMs. Moreover, these ratios were unaltered by quinidine administration. Partial metabolic clearance of N-hydroxymexiletine glucuronide, a non-CYP2D6 dependent metabolite, was highly stereoselective; the R/S ratio was 11.3 +/- 3.4. This ratio was similar in subjects with either an EM or a PM phenotype and was not altered by quinidine administration. Thus, the results obtained in this study suggest that non-CYP2D6-dependent metabolic pathways are responsible for the stereoselective disposition of mexiletine in humans.

Influence of debrisoquine hydroxylation phenotype on the pharmacokinetics of mexiletine.

Marked interindividual variation has been observed in the pharmacokinetics of the antiarrhythmic agent mexiletine. The fact that its urinary excretion is dependent on urinary pH may account, in part, for such variation. The influence that genetic differences in hepatic metabolism of the debrisoquine-type may have on mexiletine pharmacokinetics was considered in this study. The pharmacokinetics and urinary excretion of mexiletine (250 mg administered intravenously) were investigated in 5 rapid extensive metabolisers (EM), 5 slow EM and 5 poor metabolisers (PM) of debrisoquine, under conditions of controlled urinary pH. Mexiletine disposition kinetics was found to be altered in PM individuals. These subjects showed higher total area under the curve (AUC), (15.7 versus 8.16 micrograms.h.ml-1) prolonged elimination half-lives (in serum and urine) (serum: 18.5 versus 11.6 h, urine: 19.2 versus 11.7 h) and lower total clearance values compared with EM (216 versus 450 ml.min-1). In this respect, slow EM individuals generally presented intermediate values of those pharmacokinetic parameters. A higher incidence of adverse-effects was also observed among slow EM and PM subjects. It is concluded that genetic differences in mexiletine oxidation of the debrisoquine-type have an influence on its observed pharmacokinetic variability. The clinical consequences are discussed.

Single-dose quinidine treatment inhibits mexiletine oxidation in extensive metabolizers of debrisoquine.

Urinary elimination of unchanged mexiletine, p-hydroxymexiletine (PHM), hydroxymethylmexiletine (HMM) and mexiletine N-glucuronide conjugate (MGC) was investigated before and after treatment with quinidine. All subjects were phenotyped as extensive metabolizers for debrisoquine oxidation. The total recovery of mexiletine and metabolites was significantly reduced after quinidine pretreatment. It is concluded that pretreatment with a very low dose of quinidine inhibits markedly the elimination of both major mexiletine metabolites (PHM and HMM) and likely decreases the overall elimination of mexiletine. That should lead to changes in mexiletine disposition and have clinical consequences during combination therapy with both drugs.

Reversed-phase LC resolutions of chiral antiarrhythmic agents via derivatization with homochiral isothiocyanates.

The search for new antiarrhythmic agents has been intense, because the established drugs for the treatment of cardiac arrhythmias are neither uniformly effective nor well-tolerated. Among the recently introduced new antiarrhythmic agents are tocainide (TOC), mexiletine (MEX), flecainide (FLE), and propafenone (PRO). Each of these drugs is a chiral amine used clinically as the racemic mixture. We have examined the high-performance liquid chromatographic chiral resolution of the above four drugs via derivatization with homochiral derivatizing agents (HDAs). The amino functionality of the drugs was reacted with four homochiral isothiocyanates, 2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl isothiocyanate (TAGIT), (R)-alpha-methylbenzyl isothiocyanate (RAMBI), (S)-1-(1-naphthyl)ethyl isothiocyanate (SNEIT), and (R)-1-(2-naphthyl)ethyl isothiocyanate (RBEIT). Complete separation of the two peaks (resolution factor R = 1.5) was achieved with all four HDAs for TOC, with TAGIT, RBEIT, and RAMBI for MEX, with TAGIT and SNEIT for PRO, and only with TAGIT for FLE. SNEIT was used to develop analytical procedures for the determination of the enantiomeric composition of TOC in human urine and blood serum. The four HDAs offer several advantages over many other HDAs and should be useful in studies of enantioselective drug action and disposition.

The pharmacokinetics of the enantiomers of mexiletine in humans.

1. This study examined the pharmacokinetics of the enantiomers of mexiletine in five healthy subjects who were each given a single, 300 mg, oral dose of racemic mexiletine hydrochloride. 2. The time course of the concentration ratio between the R(-) and the S(+) enantiomers (R/S) in plasma showed a progressive decrease, with a mean +/- S.D. ratio of 1.37 +/- 0.11 at 1 h and 0.64 +/- 0.11 at 48 h. Similarly, the R/S ratios in urine were 1.38 +/- 0.42 and 0.55 +/- 0.12 at 1 h and 72 h, respectively. 3. The terminal elimination half-life of S(+)mexiletine was 11.0 +/- 3.80 h, which was significantly greater (P less than 0.05) than that of the R(-) enantiomer, 9.10 +/- 2.90 h. S(+)Mexiletine also showed a significantly greater apparent volume of distribution (P less than 0.01) and renal clearance (P less than 0.05) than R(-)mexiletine. There was no significant difference in the apparent oral total drug clearance of the enantiomers. 4. The disposition of mexiletine enantiomers in man was stereoselective, and the differences observed between the enantiomers may be due largely to differences in their serum protein binding.

Stereoselective analysis of the enantiomers of mexiletine by high-performance liquid chromatography using fluorescence detection and study of their stereoselective disposition in man.

A sensitive, stereoselective high-performance liquid chromatographic assay was developed for the resolution of the enantiomers of mexiletine as their 2-naphthoyl derivatives on a Pirkle type 1A chiral phase column. Detection of the derivatives was accomplished with a fluorescent detector. Maximum recovery of the enantiomers from plasma was 83% and was observed when plasma proteins were precipitated with a mixture of barium hydroxide-zinc sulphate. The calibration curve in plasma was linear over the concentration range 5-750 ng/ml for each enantiomer (r2 = 0.999) and in urine the linear range was 0.25-7.5 micrograms/ml (r2 = 0.999) for each enantiomer. The minimum detectable quantity of each enantiomer in plasma was 5 ng/ml at a signal-to-noise ratio of 5:1, representing 100 pg injected. A preliminary pharmacokinetic study was undertaken in one healthy male volunteer following an oral dose of 300 mg of racemic mexiletine hydrochloride. The apparent elimination half-lives determined from the plasma data were 12.1 and 14.1 h for the R(-) and S(+) enantiomers, respectively. The cumulative urinary excretion amounts of R(-)- and S(+)-mexiletine were found to be 8.01 and 10.46 mg, respectively. The plasma data indicated that a cross-over of the enantiomer ratios occurred at approximately 8 h. The urinary excretion of the enantiomers was consistent with the pattern found in plasma.

Mexiletine disposition: individual variation in response to urine acidification and alkalinisation.

The disposition of mexiletine has been studied in five subjects on two occasions with urine pH controlled at 5.0 and at 8.0. With acid urine total plasma clearance was similar in all subjects (462 to 497 ml min-1) and the plasma half-life ranged from 3.8 to 9.2 h (mean 6.7 h). With alkaline urine the total plasma clearance varied considerably (239 to 441 ml min-1); the mean half-life, 9.7 h, (range 7.6 to 12.7 h) was not significantly different from that in the acid urine study. Renal clearance fell greatly in every subject on alkalinisation of the urine. The total plasma clearance fell by a similar amount in two. In the remaining three the fall in total clearance was much smaller because of an increase in non-renal clearance. The reduction in total plasma clearance only just achieved statistical significance. The increase in predicted steady-state plasma mexiletine concentrations during infusion with change in urine pH from 5 to 8 varied between +5% and +95% (mean +39%). Changes in urine pH have a predictable effect upon renal clearance of mexiletine. However, disposition is changed in an unpredictable manner and inter-subject variation in distribution volume and non-renal clearance are important factors.

Determination of mexiletine in biological fluids by high-performance liquid chromatography.

A high-performance liquid chromatographic method for the determination of the anti-arrhythmic drug mexiletine in human plasma, urine, and cerebrospinal fluid is described. Following extraction with diethyl ether, mexiletine and the internal standard 4-methylmexiletine were derivatized with 2,4-dinitrofluorobenzene. Analyses were performed using an alternating on-column enrichment technique on small Perisorb RP-2 30-40 micrometers pre-columns with pre-column backflushing for direct injection onto the analytical column of Spherisorb ODS 5 micrometers. Complete separation from endogenous constituents of plasma, urine or cerebrospinal fluid was achieved by isocratic reversed-phase ion-pair chromatography with 1-heptanesulfonic acid (0.005 M; PIC B7)-acetonitrile-tetrahydrofuran (42:48:10, v/v) as eluent. Interferences from other drugs were not observed. The limit of detection was 10 ng/ml (C.V. 6.5%). Day-to-day coefficients of variation were below 9%. The applicability of this rapid method for pharmacokinetic studies and clinical routine is demonstrated.

Mexiletine, a new antiarrhythmic agent, for treatment of premature ventricular complexes.

This double-blind crossover study was designed to compare the safety and efficacy or mexiletine, a new class I antiarrhythmic agent, with those of a placebo in reducing premature ventricular complexes. Twelve patients who had a median of 294 such complexes/hour were admitted to the study. Eleven completed 4 weeks of trial with mexiletine and placebo with ambulatory electrocardiographic (Holter) recordings taken at the end of each treatment period. The doses given were designed to reduce the frequency of premature ventricular complexes by 50 percent or more from the baseline value. Mexiletine significantly reduced the rate of premature ventricular complexes by comparison with placebo (-66 percent versus 3 percent, p = 0.032). In addition, mexiletine reduced the median number/hour of ventricular couplets observed. After 4 weeks of therapy, 2, 2, 1 and 6 patients, respectively, were taking 100, 200, 300 and 400 mg of mexiletine every 8 hours. Mexiletine produced no significant change in baseline values including electrocardiographic intervals, blood pressure or heart rate. The most frequently observed adverse effects were digestive difficulties (eight patients taking mexiletine, four taking placebo) and central nervous system effects (seven taking mexiletine, five taking placebo). These data show the efficacy and safety of mexiletine in the treatment of premature ventricular complexes in a majority of patients.

The effect of spontaneous changes in urinary pH on mexiletine plasma concentrations and excretion during chronic administration to healthy volunteers.

1. The effect of spontaneous changes in urinary pH on renal excretion and plasma concentration of mexiletine has been examined during chronic administration of subtherapeutic doses of the drug to healthy volunteers. 2. Significant correlations were found between urinary pH and the plasma concentration and renal excretion of mexiletine. 3. Prediction of plasma mexiletine concentrations from our data suggests that the amount of mexiletine in plasma would increase by more than 50% following a rise in urinary pH similar to that which occurred spontaneously in our subjects. 4. Factors which influence urinary pH should be considered when the dosage of mexiletine is chosen. Extremes of urinary pH may account for some cases of inefficacy of the drug, and for the occurrence of unwanted effects at conventional doses.

New electron-capture gas-liquid chromatographic method for the determination of mexiletine plasma levels in man.

A method for the determination of mexiletine in human plasma by gas-liquid chromatography with electron-capture detection is described. Plasma samples are extracted at pH 12 with dichloromethane after addition of the internal standard, the 2,4-methyl analogue of mexiletine. A derivative is obtained using heptafluorobutyric anhydride; according to gas chromatography-mass spectrometry it is a monoheptafluorobutyryl compound. The minimum detectable amount of mexiletine is 5 pg. Accurate determinations of human plasma levels were performed after oral or intravenous treatment.

The identification and analysis of mexiletine and its metabolic products in man.

Sensitive and specific gas-liquid chromatographic methods were developed for the analysis of mexiletine and its metabolites in urine of man. The identity of the g.l.c. peaks was established by mass-spectrometry. The hydroxylamine (Va) was qualitatively identified and determined quantitatively after conversion to the more stable oxime (Vb). Selective extraction procedures, t.l.c. and derivatization with hexamethyldisilazane (HMDS) and trifluoroacetic anhydride (TFA) were used in the qualitative identification of the major metabolites (VI-IX), particularly in distinguishing the basic products VI and VII from their corresponding alcoholic products VIII and IX. The limit of detection of the g.l.c. method was 6 to 12 ng ml-1 for compounds I-IV, and 40 to 50 ng ml-1 for compounds Vb-IX.