A simple high-performance liquid chromatographic determination of mexiletine in human plasma at therapeutic levels

Summary A method for the quantitative determination of mexiletine in human plasma by high-performance liquid chromatography has been described. The plasma samples are buffered to pH 12 and extracted on Clin-Elut columns with diethylether-ethylacetate (1:1), after addition of the internal standard, the 2,4,6 methyl analogue of mexiletine. The minimum detectable amount of mexiletine is 50 ng in 0.5 ml plasma. Recovery is between 96–114% and the relative standard deviation at 1.5 ml^–1 level of mexiletine is 2.1% Accurate determinations of human plasma levels were performed after oral or intravenous treatment.Part of the work was presented at the 29. Kongreß der Deutschen Gesellschaft für Laboratoriumsmedizin, Hamburg 1985.     

Liquid chromatographic analysis of mexiletine in serum, with alternate application to tocainide, procainamide, and N-acetylprocainamide.

A simple and precise high performance liquid chromatographic method for the determination of mexiletine in human serum or plasma is described. Following addition of N-propionylprocainamide as internal standard the specimens are extracted, under basic conditions, into methylene chloride. After removal of the aqueous layer the drug is back-extracted into dilute acid, which is then injected directly for analysis. The extraction efficiency is 79% for both mexiletine and internal standard, and the assay is linear to 4 mg/L (twice upper therapeutic concentration). Inter-run coefficients of variation are 3.0% or less. The relative retention time of mexiletine to internal standard averages 1.3. An adaptation of this method is described for an alternate application to the analysis of tocainide, procainamide and N-acetylprocainamide.     

A sensitive determination method for mexiletine derivatized with dansyl chloride in rat plasma utilizing a HPLC peroxyoxalate chemiluminescence detection system.

A sensitive determination method for a non-fluorescent anti-arrhythmic drug, mexiletine, in rat plasma is presented utilizing a HPLC peroxyoxalate chemiluminescence (PO-CL) detection system. After an internal standard (4-methylmexiletine, 4.35 pmol) and 0.1 N sodium hydroxide solution were added to 5 microL rat plasma, the solution was poured onto an Extrelut 1 column. Both mexiletine and the internal standard were eluted with diethy ether and then the eluate was evaporated to dryness. The residue was dissolved in 0.2 M borate buffer (pH 8.5) and mixed with dansyl chloride (75 nmol) in acetronitrile. After standing of 90 min at room temperature, 0.5 N HCl was added to the reaction mixture to stop the reaction and a 2/45 aliquot of the mixture was subjected to a HPLC PO-CL detection system using bis(4-nitro-2(3,6,9-trioxadecyloxycarbonyl)phenyl) oxalate (TDPO) and hydrogen peroxide. The calibration curve for mexiletine in rat plasma was linear over the range 20-100 ng/mL plasma (20.6-103 fmol/injection). The detection limit (S/N = 2) was 1.0 fmol over the whole procedure. The method was applied to the measurement of the time courses of plasma mexiletine concentration after oral administration of the drug [25 mg (115.9 mumol)/kg] to rats.     

Simultaneous determination of mexiletine and four hydroxylated metabolites in human serum by high-performance liquid chromatography and its application to pharmacokinetic studies.

A high-performance liquid chromatographic method has been developed for the simultaneous determination of mexiletine and its four hydroxylated metabolites in human serum. The method involves a single-step extraction of mexiletine, hydroxymethylmexiletine, p-hydroxymexiletine and their corresponding alcohols with diisopropyl ether-dichloromethane-propan-2-ol (2.5:1.5:0.5, v/v). Separation of the compounds on a deactivated Supelcosil LC8-DB column is accomplished by high-performance liquid chromatography with ultraviolet detection at 203 nm. Overall the recovery of each compound is reproducible and greater than 75%. The lower limit of detection is 2 ng/ml for mexiletine and its metabolites. The application of the method is shown by measuring the concentrations in serum of mexiletine and its metabolites over 24 h in a healthy volunteer after a single intravenous injection of the drug and by monitoring serum concentrations in patients receiving long-term treatment by mouth of the drug.     

Determination and Pharmacokinetics of Mexiletine in Rabbit Plasma by Gas Chromatography with Mass Spectrometry

This paper describes a gas chromatography/mass spectrophotometry method for determination of mexiletine in rabbit plasma. Mexiletine and internal standard metoprolol were extracted from rabbit plasma with a mixture of ethylacetate and diethylether at basic pH with liquid-liquid extraction. Calibration curves were linear over the concentration range 45–2000 ng/mL. Intra- and inter-day precision (relative standard deviation) for mexiletine in rabbit plasma were less than 6.9%, and accuracy (relative error) was better than 6.8%. Recovery of mexiletine from rabbit plasma averaged 92.6%. This method was successfully applied to six rabbits which had given an oral capsule of 200 mg mexiletine.     

Fluorimetric determination of mexiletine in serum by high-performance liquid chromatography using pre-column derivatization with fluorescamine

A simple, specific and sensitive micro-scale method for the assay of the antiarrhythmic agent mexiletine in human serum is described. The method uses high-performance liquid chromatography, with pre-column fluorimetric derivatization by fluorescamine. Following extraction with diethyl ether, mexiletine and 4-methylmexiletine (an internal standard) were derivatized with fluorescamine under weakly alkaline condition (pH 9.0) and chromatographed on a reversed-phase column with aqueous methanol-2-propanol as the mobile phase. The two fluorescent derivatives of mexiletine and the internal standard were separated as clear single peaks, and no interfering peaks were observed on the chromatograms. The detection limit for mexiletine was 0.005 micrograms/ml from only 100 microliters of serum, and the calibration curves in the range 0.01-5 micrograms/ml were linear, with an overall coefficient of variation of less than 5%. The analytical recovery of a known amount of mexiletine added to serum was almost 100%. This method proved to be effective in the rapid monitoring of the serum concentrations in patients who received this potent antiarrhythmic drug.     

High-performance liquid chromatographic assay for mexiletine hydroxylation in microsomes of human liver

A simple high-performance liquid chromatographic assay, using fluorescence detection, is described for determining simultaneously the production of the two major hydroxylated metabolites of mexiletine in human liver microsomes. The detection limits of hydroxymethylmexiletine and p-hydroxymexiletine are 0.35 and 0.08 nmol/ml, respectively. The assay is specific, reproducible and allows the simultaneous kinetic characterization of the reactions in small amounts of liver tissue. The assay may be used to acquire a better knowledge of the kinetic behaviour of mexiletine and of its metabolites, and to investigate if the large inter-individual variations of the mexiletine pharmacokinetics are of metabolic origin, due to variations of its hydroxylation processes.     

Selective and sensitive high-performance liquid chromatographic assay for the metabolites of nomifensine in human plasma

A selective high-performance liquid chromatographic method for the determination of the three metabolites of nomifensine in human plasma is described. All metabolites and the internal standard, mexiletine, are extracted with diethyl ether and then back-extracted into an acidic aqueous phase. After subsequent extraction into diethyl ether the metabolites are analysed by high-performance liquid chromatography. A reversed-phase C18 column is used with a mobile phase of dioxane-methanol-potassium phosphate buffer (pH 2.25). The sensitivity of the method is 0.007 micromol/l for all metabolites. Extraction efficiencies are 84.6%, 75.8%, and 78.2% for 4'-hydroxynomifensine, 4'-hydroxy-3'-methoxynomifensine and 3'-hydroxy-4'-methoxynomifensine, respectively. The reproducibility of the method is good, the coefficients of variation (%) varying between 2.1% and 9.9% in the concentration range 0.05-1.00 micromol/l. The procedure was applied to human plasma samples from a volunteer who had received a single oral dose of nomifensine. The method is accurate and sensitive for pharmacokinetic studies on the metabolites of nomifensine.     

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.     

Determination of bupropion using liquid chromatography with fluorescence detection in pharmaceutical preparations, human plasma and human urine

A novel pre-column derivatization reversed-phase high-performance liquid chromatography with fluorescence detection is described for the determination of bupropion in pharmaceutical preparation, human plasma and human urine using mexiletine as internal standard. The proposed method is based on the reaction of 4-chloro-7-nitrobenzofurazan (NBD-Cl) with bupropion to produce a fluorescent derivative. The derivative formed is monitored on a C18 (150 mm × 4.6 mm i.d., 5 μm) column using a mobile phase consisting of methanol-water 75:25 (v/v), at a flow-rate of 1.2 mL/min and detected fluorimetrically at λ(ex) = 458 and λ(em) = 533 nm. The assay was linear over the concentration ranges of 5-500 and 10-500 ng/mL for plasma and urine, respectively. The limits of detection and quantification were calculated to be 0.24 and 0.72 ng/mL for plasma and urine, respectively (inter-day results). The recoveries obtained for plasma and urine were 97.12% ± 0.45 and 96.00% ± 0.45, respectively. The method presents good performance in terms of precision, accuracy, specificity, linearity, detection and quantification limits and robustness. The proposed method is applied to determine bupropion in commercially available tablets. The results were compared with an ultraviolet spectrophotometry method using t- and F-tests.     

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.     

High pressure liquid chromatographic assay for mexiletine in serum

A highly sensitive, rapid, and specific high pressure liquid chromatographic assay for the analysis of the antiarrhythmic agent mexiletine is reported. The method involves extraction of mexiletine with organic solvent followed by analysis using fluorescence detection. The minimum measurable limit is 1 ng and inter- and intra-day coefficients of variation are less than 5.8%. The method is useful for pharmacokinetic studies and routine serum monitoring of mexiletine.     

A practical and efficient route for the highly enantioselective synthesis of mexiletine analogues and novel beta-thiophenoxy and pyridyl ethers

A practical and efficient procedure for the enantioselective synthesis of mexiletine analogues with use of 10% of spiroborate ester 6 as chirality transfer agent is presented. A variety of mexiletine analogues were prepared in good yield with excellent enantioselectivities (91-97% ee) from readily available starting materials. The developed methodology was also successfully applied for the synthesis of novel beta-amino ethers containing thiophenyl and pyridyl fragments.     

Enantiomeric separation of basic drugs with partially filled serum albumin as chiral selector in capillary electrophoresis.

A reliable method is presented for the chiral separation of three basic drugs (mexiletine, chlorpheniramine and propranolol) with serum albumins (human and porcine, HSA and PSA) as chiral selectors by capillary electrophoresis in combination with the partial filling technique. Based on the systematic optimization of operation variables, the chiral separation of mexiletine, chlorpheniramine and propranolol was achieved in the pH 7.4 phosphate buffer by using HSA, PSA and PSA as selectors, respectively. The chiral recognition ability of HSA and PSA was compared. HSA and PSA show a different chiral recognition ability for each of the three drugs. In addition, the association constants between enantiomeric drugs and proteins were determined to be 2.00 and 3.80 x 10(2) M(-1) for mexiletine and HSA, 0.59 and 1.12 x 10(3) M(-1) for chlorpheniramine and PSA, and 0.87 and 1.42 x 10(3) M(-1) for propranolol and PSA. The method for the chiral separation and determination of association constants possesses the advantages of simple performance, effective avoiding of the interference of the UV detection from protein, and lowering of the reagent consumption.     

First synthesis and full characterization of mexiletine N-carbonyloxy β-d-glucuronide

A simple, convergent synthesis of the N-carbonyloxy β-d-glucuronide of mexiletine (sodium salt) in moderate yield is described. The compound is now available as an authentic reference standard for analytical studies, enabling more detailed investigation on the metabolism of mexiletine.     

Preparation of highly enantiopure stereoisomers of 1-(2,6-dimethylphenoxy)-2-aminopropane (mexiletine)

Mexiletine [1-(2,6-dimethylphenoxy)-2-aminopropane], an orally effective antiarrhythmic agent, exhibits enantioselective pharmacokinetics and pharmacodynamics during mexiletine therapy. The purpose of this paper is to emphasize the advantage of tetrahydropyranyl-protected mandelic acid (THPMA) in the resolution of mexiletine enantiomers. Both enantiomers of mexiletine were obtained in 99% enantiomeric excess. Judging by the differential shielding effects in the ¹H and ¹³C NMR analyses, we have observed the opposite predominant conformation for the mexiletine mandelates in comparison with the O-methylmandelates.     

微流控芯片毛细管电泳激光诱导荧光检测法测定片剂中盐酸美西律的含量

建立了微流控芯片毛细管电泳激光诱导荧光检测法测定片剂中盐酸美西律含量的方法,对衍生条件和电泳条件进行了系统的考察。盐酸美西律经异硫氰酸荧光素(FITC)40℃衍生6h,以20 mmol/L硼砂为电泳缓冲溶液,进样30s后,分离电压2000V,可在1 min内完成一次检测。方法的检出限为0.022 mg/L、线性范围0.108～1.079 mg/L、相关系数0.994,加标回收率为99.7%～102.3%,方法适用于盐酸美西律的检测和质量控制。     

Spectrophotometric and Spectrofluorometric Determination of Heptaminol and mexiletine in Their Dosage Forms

Abstract

Two simple, rapid specific methods were developed for the determination of heptaminol and mexiletine and their dosage forms. The methods are based on the reaction of either heptaminol or mexiletine with acetylacetone-formaldehyde reagent to give a yellow chromophore measurable spectrophotometrically at 344 or 338 nm or flurometrically at 480 nm for heptaminol and mexiletine, respectively. The color was stable for at least 1/2 h. Beer's law was valid within a concentration range of 15–30 and 8–20 μg ml^?1 spectrophotometrically and 0.2–0.8 and 0.4–1.0 μg ml^?1 fluorometrically for heptaminol and mexiletine, respectively.     

Development of an HPLC method for the determination of mexiletine in human plasma and urine by solid-phase extraction

A sensitive and specific high-performance liquid chromatography (HPLC) method has been developed and validated for the quantification of mexiletine (MEX) in human plasma and urine. It uses solid-phase extraction (SPE) followed by an automated reversed-phase HPLC with a pre-column derivatization with 4-chloro-7-nitrobenzofurazan (NBD-CI) and UV-vis Absorbance detection. The process was set as: the UV-vis Absorbance wavelength was set at 458 nm. Chromatographic separation was performed on a Phenomenex-C₁₈ Column (Aqua, 150 mm × 4.6 mm i.d. with 5 μm particle size) with the mobile phase consisting of acetonitrile and water (80:20, v/v), and the flow rate was set at 1.0 mL min⁻¹. Calibration of the overall analytical procedure gave a linear signal (r > 0.9998) over a MEX concentration range of 0.2-2.0 μg mL⁻¹ in human plasma and urine. The detection limit in plasma and urine was 0.1 μg mL⁻¹. Intra- and inter-day precision of the assay at three concentrations within this range were 0.31-2.50%. The high specificity and sensitivity have been achieved by this fast method (total run-time <6 min). The method has been successfully validated in human plasma and urine and it has been shown to be precise, accurate and reliable.     

LC Method with Precolumn Derivatization for Analysis of Mexiletine in Pharmaceutical Preparations

A isocratic, selective and accurate LC method of analysis of mexiletine in pharmaceutical preparations has been developed and validated. The method is based on derivatization of mexiletine with 4-chloro-7-nitrobenzofurazan in pH 9.0 borate buffer to yield a yellow product. Chromatography was performed on a C₁₈ column (150 × 4.6 mm i.d.) with acetonitrile–water 80:20 (v/v) as mobile phase at a flow rate of 1.0 mL min^?1. UV–visible absorbance detection was performed at 458 nm. The retention time of the mexiletine derivative was 4.10 min, and response was a linear function of concentration in the range 0.5–4.0 μg mL^?1 (r = 0.9998). The limits of detection and quantification were 0.05 and 0.15 μg mL^?1, respectively. Method validation revealed precision, sensitivity, and robustness were acceptable. Low RSD values are indicative of high precision, and high recovery values are indicative of the accuracy of the method. Results obtained by use of the proposed method for analysis of the mexiletine content of pharmaceutical a preparation were compared with those obtained by use of the official method. The method has been used for analysis of pharmaceutical preparations.