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In an acidic buffered solution, erythrosine B can react with amiodarone to form an association complex, which not only generates great enhancement in resonance Rayleigh scattering (RRS) spectrum of erythrosine B at 346.5 nm but also results in quenching of fluorescence spectra of erythrosine B at λ emission  = 550.4 nm/λ excitation  = 528.5 nm. In addition, the formed erythrosine B-amiodarone complex produces a new absorbance peak at 555 nm. The spectral characteristics of the RRS, absorbance, and fluorescence spectra, as well as the optimum analytical conditions, were studied and investigated. As a result, new spectroscopic methods were developed to determine amiodarone by utilizing erythrosine B as a probe. Moreover, the ICH guidelines were used to validate the developed RRS, photometric, and fluorimetric methods. The enhancements in the absorbance and the RRS intensity and the decrease in the fluorescence intensity of the used probe were proportional to the concentration of amiodarone in ranges of 2.5-20.0, 0.2-2.5, and 0.25-1.75 μg/mL, respectively. Furthermore, limit of detection values were 0.52 ng/mL for the spectrophotometric method, 0.051 μg/mL for the RRS method, and 0.075 μg/mL for the fluorimetric method. Moreover, with good recoveries, the developed spectroscopic procedures were applied to analyze amiodarone in its commercial tablets., (© 2024 John Wiley & Sons Ltd.)

This research explored the HPLC fingerprints of Hypericum attenuatum Choisy, which has anti-arrhythmic activity. HPLC was adopted to perform a determination of chemical fingerprints of H. attenuatum specimens acquired through seven distinct sources. The anti-arrhythmic activity of each H. attenuatum sample was obtained through pharmacodynamics experiments in animals. A regression analysis and correlation analysis were utilized to calculate the relationship of the peak and pharmacological effectiveness with the identified peak. Peaks numbered 5, 7, 13 and 14 in the fingerprint were regarded as the likely anti-arrhythmic agents. The fingerprint was compared with reference standards for identification of the correlative peaks. Liquid chromatography-time-of-flight-mass spectrometry was applied to identify its structure. As a consequence, a universal model was established for the utilization of HPLC to investigate anti-arrhythmic activity and the spectrum-effect relationship among H. attenuatum. This model is available for the discovery of the major bioactive constituents of Hypericum., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The potential for inhibiting recrystallization with Eudragit® L (EUD-L), hypromellose acetate succinate (HPMC-AS), and polyvinylpyrrolidone-co-vinylacetate (PVP-VA) on amorphous felodipine (FLD) at low polymer loading was investigated in this study. The physical stabilities of the FLD/polymer amorphous solid dispersions (ASDs) were investigated through storage at 40 °C. The HPMC-AS and PVP-VA strongly inhibited FLD recrystallization, although EUD-L did not effectively inhibit the FLD recrystallization. The rotating frame 1 H spin-lattice relaxation time ( 1 H-T 1ρ ) measurement clarified that EUD-L was not well mixed with FLD in the ASD, which resulted in weak inhibition of recrystallization by EUD-L. In contrast, the HPMC-AS and PVP-VA were well mixed with the FLD in the ASDs. Solid-state 13 C spin-lattice relaxation time ( 13 C-T 1 ) measurements at 40 °C showed that the molecular mobility of the FLD was strongly suppressed when mixed with polymer. The reduction in the molecular mobility of FLD was in the following order, starting with the least impact: FLD/EUD-L ASD, FLD/HPMC-AS ASD, and FLD/PVP-VA ASD. FLD mobility at the storage temperature, evaluated by 13 C-T 1 , showed a good correlation with the physical stability of the amorphous FLD. The direct investigation of the molecular mobility of amorphous drugs at the storage temperature by solid-state NMR relaxation time measurement can be a useful tool in selecting the most effective crystallization inhibitor at low polymer loading., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

For the first time, three phase hollow fiber liquid phase microextraction using an influential, and green middle phase comprised a new relatively-hydrophobic deep eutectic solvent (three-phase HF-LPME-DES) was developed for trace analyses of antiarrhythmic drugs in biological and environmental samples. The extraction solvent was easily synthesized by mixing the green and cheap raw materials, namely choline chloride and 1-phenylethanol (ChCl: Ph-ETOH), in the ambient temperature. Good compatibility to pores of hollow-fiber, high ability for extraction of ionizable organic compounds with no need to any carrier agents, and easy availability in the laboratory environment turned this new proposed deep eutectic intermediate to a worthy generation of the supported liquid membrane (SLM). Final determination was accurately done by high performance liquid chromatography-ultraviolet detection (HPLC-UV). After effective statistical optimization of main parameters, the valid analytical features were found to be: wide linear dynamic ranges (LDRs) of 0.8 to 500 ng mL -1 with the determination coefficients (R 2 s) higher than 0.98, low detection limits (LODs) of 0.3-0.8 ng mL -1 , and logical precision (relative standard deviations (%RSDs, n = 3) of 5.2-6.5%). Also, enrichment factors and extraction recoveries were 110-135 and 44-54%, respectively. These satisfactory results confirmed the potent effectiveness of the proposed microextraction procedure for achievement to clean and proper enrichment of the aforesaid compounds in highly complex real samples., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Use of drug-metal complexes for the treatment of several human diseases has resulted in significant progress in the field of medicinal inorganic chemistry. The current study describes the synthesis and characterization of Cu (II) and Ni (II) complexes of Losartan, an antihypertensive drug. These complexes were evaluated for their cytotoxic activity against four human cancer cell lines; SNB-19, HCT-15, COLO-205 and KB-3-1. Spectroscopic characterization revealed that during complex formation, the metal was bound through the nitrogen atoms of the tetrazole moiety of the losartan molecule. The molecular formulas of copper ([Cu (LS) 2 Cl 2 ].6H 2 O) and nickel ([Ni (LS) 2 Cl 2 ]. H 2 O) complexes were found to be in agreement with the analytical data obtained through elemental analysis. For both the complexes, metal to ligand ratios of 1:2 were calculated. As revealed by FTIR, UV-Visible, and 1 H-NMR studies, both the complexes displayed octahedral geometries. Scanning electron microscopy (SEM) revealed marked changes in the morphology of the complexes, compared to the pure drug. From XRD studies, characteristic crystalline peaks of pure losartan were observed whereas no prominent peaks were observed for its complexes. Complexes were found to be inactive in the cytotoxic activity test performed using SNB-19, HCT-15, COLO-205 and KB-3-1 cell lines.

In the current study, liquid-liquid extraction, using deep eutectic solvent (DES) as a "green" extraction solvent, was coupled with a stepwise injection system for the first time. The suggested approach was applied for the development of spectrofluorimetric method for procainamide determination. The method is based on aspiration of saliva sample and DES (choline chloride with glycerol at a 1:2M ratio) solution into the mixing chamber of a flow system, followed by injection of acetonitrile into the mixed DES-sample solution. The extraction process and final phase separation were then promoted by air-bubbling. After phase separation, the DES phase, containing the extracted procainamide, was transported to a spectrofluorimetric detector. The excitation and emission wavelengths were set at 280nm and 347nm, respectively. The calibration plot was linear in the range of 5×10 -6 to 5×10 -5 molL -1 . The limit of detection, calculated as 3σ of a blank test (n=10), was found to be 1.5×10 -6 molL -1 . The developed method was successfully applied for the determination of procainamide in human saliva samples, and the analytical results agreed rather well with the results obtained by the reference HPLC-UV method., (Copyright © 2017 Elsevier B.V. All rights reserved.)

In this work, two stability-indicating chromatographic methods have been developed and validated for determination of flecainide acetate (an antiarrhythmic drug) in the presence of its degradation products (flecainide impurities; B and D). Flecainide acetate was subjected to a stress stability study including acid, alkali, oxidative, photolytic and thermal degradation. The suggested chromatographic methods included the use of thin layer chromatography (TLC-densitometry) and high-performance liquid chromatography (HPLC). The TLC method employed aluminum TLC plates precoated with silica gel G.F254 as the stationary phase and methanol-ethyl acetate-33% ammonia (3:7:0.3, by volume) as the mobile phase. The chromatograms were scanned at 290 nm and visualized in daylight by the aid of iodine vapor. The developed HPLC method used a RP-C18 column with isocratic elution. Separation was achieved using a mobile phase composed of phosphate buffer pH 3.3-acetonitrile-triethylamine (53:47:0.03, by volume) at a flow rate of 1.0 mL/min and UV detection at 292 nm. Factors affecting the efficiency of HPLC method have been studied carefully to reach the optimum conditions for separation. The developed methods were validated according to the International Conference on Harmonization guidelines and were applied for bulk powder and dosage form. Copyright © 2016 John Wiley & Sons, Ltd., (Copyright © 2016 John Wiley & Sons, Ltd.)

Two new potential impurities of antiarrhythmic drug substance Dronedarone Hydrochloride together with debutyldronedarone were detected by LC-MS analysis during process development. A successful synthetic strategy for the synthesis of these potential impurities was developed facilitating the access to new impurity reference standards. Their synthesis and characterization are discussed in detail. The availability of these impurity standards allowed cost reduction through the increase of process control., (Copyright © 2015 Elsevier B.V. All rights reserved.)

To achieve successful drug delivery via nanoparticles the interactions between the nanoparticle and the chemistry of the surrounding biological environment is of central importance. A thorough understanding of these interactions is necessary in order to better elucidate information regarding drug pathways and mechanisms of action in treatment protocols. As such, it is important to identify the location of the nanoparticle, the state of its functionalization, as well as any changes in the cellular environment. The use of cluster secondary ion mass spectrometry (SIMS) using C60 (+) primary ions makes simultaneous acquisition of this information possible. Here, SIMS has been successfully used to chemically image gold nanoparticles (AuNPs) within a model, single cell system involving macrophage-like RAW 264.7 cells. The macrophage-like properties of this cell line make it extremely well-suited for cell-uptake studies. Both AuNPs and two pharmaceutical compounds, amiodarone and elacridar, were successfully imaged within a cellular system using cluster SIMS. To verify that SIMS can also be used to detect functionalization and nanoparticles simultaneously, fluorophore-functionalized AuNPs were studied as a model system. The fluorescent characteristics of these functionalized nanoparticles enabled the visual confirmation of the presence and location of the particles within the cell.

The USP monograph describes an HPLC method for seven impurities in the amiodarone drug substance using a L1 column, 4.6mm×150mm, 5μm packing (PF listed ODS2 GL-Science, Inertsil column) at 30°C with detection at 240nm. The standard contains 0.01mg/mL of amiodarone, and USP specified impurities D and E with a resolution requirement of NLT 3.5 between peaks D and E. Impurities in a 5mg/mL sample are quantitated against the standard. Impurity A peak elutes just before peak D. We observed two problems with the method; the column lot-to-lot variability resulted in unresolved A, D, and E peaks, and peak D in the sample preparation eluted much later than that in the standard solution. Therefore, optimization experiments were conducted on the USP method following the QbD approach with Fusion AE™ software (S-Matrix Corporation). The resulting optimized conditions were within the allowable changes per USP 〈621〉. Lot-to-lot variability was negligible with the Atlantis T3 (Waters Corporation) L1 column. Peak D retention time remained constant from standard to sample. The optimized method was validated in terms of accuracy, precision, linearity, range, LOQ/LOD, specificity, robustness, equivalency to the USP method, and solution stability. The QbD based development helped in generating a design space and operating space with knowledge of all method performance characteristics and limitations and successful method robustness within the operating space., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Two cases of mothers given postpartum short-term administration of amiodarone, with and without bisoprolol, are described along with determinations of amiodarone and (±)-bisoprolol in the breast milk. In one mother given a cumulative total of amiodarone of 8 g over 1 week, concentrations 11 days after the drug had been stopped were initially deemed sufficient to pose a risk to an infant. Over the next 5 days the concentrations steadily dropped with amiodarone and desethylamiodarone concentrations being found to be at a level comprising minimal risk to the infant. Bisoprolol was not found in the expressed breast milk. In the second case the mother was given a single 150 mg dose of amiodarone and breast milk concentrations were measured on postpartum days 4 and 5. Breast milk amiodarone concentrations were very low and of little concern clinically had the mother breast fed her baby. The risk to the baby of ingesting breast milk after amiodarone administration postpartum depends on the duration of amiodarone exposure, with a single dose posing minimal risk. Bisoprolol does not appear to accumulate to any great extent in breast milk., (© 2014, The American College of Clinical Pharmacology.)

Lethal occurrence is exceptional after disopyramide or mianserin poisoning. A case of intentional lethal intoxication with these drugs was reported, as well as a review of the literature. Pre- and postmortem blood concentrations of disopyramide or mianserin were assessed in a woman who died from acute cardiac failure after ingestion. The premortem blood concentration of disopyramide alone was considered lethal, and a toxic premortem concentration of mianserin was observed that may have increased cardiovascular failure induced by disopyramide because the metabolism of both drugs is mediated via cytochrome P450. Moreover, it was shown that the postmortem redistribution of disopyramide was limited, as pre- and postmortem concentrations were 48 and 65 mg/L, respectively. As regards mianserin, redistribution was observed after death with pre- and portmortem concentrations at 0.23 and 0.79 mg/L, respectively. This case illustrates that if postmortem blood concentration of disopyramide is known, the premortem concentration can be deduced., (© 2014 American Academy of Forensic Sciences.)

An ultrasensitive electrochemical sensor based on the electrocatalytic properties of nano graphene (nGN) and multiwalled carbon nanotubes (MWCNTs) for the detection of quinidine (QD) in solubilized systems has been developed. This nano graphene-multiwalled carbon nanotube (nGN-MWCNT) composite film modified sensor shows the higher stability and stronger catalytic activity towards oxidation of quinidine and the over-potential decreased significantly compared with the bare glassy carbon electrode (GCE). The electrochemical characterization of the sensor was done by scanning electron microscopy (SEM) and cyclic voltammetry (CV) with working electrode surface area 0.56 cm(2) and diffusion coefficient 2.15×10(-3)cm(2)s(-1). Under the optimized conditions, the oxidation peak current of QD is found to be proportional to its concentration in the range of 60 ng-50 μg with a detection limit of 0.186 ng. The sensor was successfully employed for the detection of quinidine in bulk drugs and in its commercial pharmaceutical formulation., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Ambient ionization methods for mass spectrometry have enabled the in situ and in vivo analysis of biological tissues and cells. When an etched optical fiber is used to deliver laser energy to a sample in laser ablation electrospray ionization (LAESI) mass spectrometry, the analysis of large single cells becomes possible. However, because in this arrangement the ablation plume expands in three dimensions, only a small portion of it is ionized by the electrospray. Here we show that sample ablation within a capillary helps to confine the radial expansion of the plume. Plume collimation, due to the altered expansion dynamics, leads to greater interaction with the electrospray plume resulting in increased ionization efficiency, reduced limit of detection (by a factor of ~13, reaching 600 amol for verapamil), and extended dynamic range (6 orders of magnitude) compared to conventional LAESI. This enhanced sensitivity enables the analysis of a range of metabolites from small cell populations and single cells in the ambient environment. This technique has the potential to be integrated with flow cytometry for high-throughput metabolite analysis of sorted cells.

Objective: Immunofluorescence and Western blot methods were adopted for qualitative and quantitative detections of the effect of different concentrations of berberine, liensinine and neferine on the expression of stable transfection in HERG potassium channel in HEK-293 cells, as well as the effect of different concentrations of berberine on protein expression of Ikr channel in cardiac muscular tissues, in order to investigate the anti-arrhythmic mechanism of berberine, liensinine and neferine., Method: Western blot method was used to detect protein expression of HERG channel in HERG-HEK cells. Immunofluorescence method as well as confocal laser microscope were used to detect the effect of different concentrations of berberine, liensinine and neferine on protein expression of HERG channel. Western blot method was used to detect the effect of different concentrations of berberine on protein expression of Ikr channel in cardiac muscular tissues as well as the effect of berberine, liensinine and neferine on protein expression of stable transfection in HERG potassium channel in HEK-293 cells., Result: Western blot experiment manifested that stable transfection of HEK293 cells containing HERG genes could increase protein expression of HERG channel. Berberine (10, 30 micromol x L(-1)) remarkably inhibited protein expression of HERG channel in HERG-HEK cells (P < 0.01). Berberine (10, 20 mg x kg(-1)) also inhibited protein expression of Ikr channel in rat ventricular tissues (P < 0.05). Liensinine (3, 10, 30 micromol x L(-1)) increased protein expression of HERG channel in HERG-HEK cells (P < 0.05). Neferine showed no effect on protein expression of HERG channel in HERG-HEK cells., Conclusion: The stably transfection of HERG-HEK cells can increase protein expression of HERG channel. Berberine shows inhibitory effect on protein expressions of in vitro HERG-HEK cells and Ikr channel in rat ventricular tissues. Liensinine improves protein expression of HERG channe in HERG-HEK cells. Neferine shows no effect on protein expression of HERG channel.

In order to quantify small molecules at the early stage of drug discovery, we developed a quantitation approach based on mass spectrometry imaging (MSI) using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) without the use of a labeled compound. We describe a method intended to respond to the main challenges encountered in quantification through MALDI imaging dedicated to whole-body or single heterogeneous organ samples (brain, eye, liver). These include the high dependence of the detected signal on the matrix deposition, the MALDI ionization yield of specific target molecules, and lastly, the ion suppression effect on the tissue. To address these challenges, we based our approach on the use of a normalization factor called the TEC (Tissue Extinction Coefficient). This factor takes into account the ion suppression effect that is both tissue- and drug-specific. Through this protocol, the amount of drug per gram of tissue was determined, which in turn, was compared with other analytical techniques such as Liquid Chromatography-Mass spectrometry (LC-MS/MS)., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Amiodarone hydrochloride, a class III antiarrhythmic agent, shows ß blocker-like and potassium channel blocker-like actions on the sinuatrial and atrioventricular nodes. It is given by mouth in the treatment of all forms of atrial, junctional and ventricular arrhythmias. Capsules for paediatric patients are not commercially available and must be prepared in the pharmacy department. The aim of the study was to evaluate the stability of amiodarone hydrochloride at different dosages, 10, 60 and 100 mg, in capsules for paediatric patients stored in three packages under dark conditions and at room temperatures over one year. At different intervals during the storage period, amiodarone hydrochloride concentration was tested using a high-performance liquid chromatography (HPLC) method. Amiodarone hydrochloride content remained greater than 95% of the initial concentration in all capsules at all dosages. The 10, 60 and 100 mg amiodarone paediatric capsules were stable for one year when stored in the three packages at ambient temperature and under dark conditions., (Copyright © 2012 Elsevier Masson SAS. All rights reserved.)

Amiodarone is an antiarrhythmic agent used for various types of tachyarrhythmia, both ventricular and supraventricular (atrial) arrhythmia. A spectrophotometric method for the assay of amiodarone was established. Based on the reduction of potassium ferricyanide in hydrochloric acid medium to potassium ferrocyanide forming a blue colored complex ferric ferrocyanide with Fe (III) ions. The compound was most stable in a mixture of ethylic alcohol and water (2:1, v/v) and it had an absorption maximum at 725 nm. The data were according to the Lambert-Beer Law in the concentration range of 0.5-5.0 microg/sample: correlation and coefficient R = 0.99977, R2 = 0.999541, slope of the line 0.12775, intercept 0.042077. The detection limit (DL) was 0.1032 microg/sample and the quantification limit (QL) 0.344 microg/ sample.

A selective capillary electrophoresis method for determination of enantiomeric purity of RS86017, a new antiarrhythmic agent with two chiral centers, was developed and validated using sulfobutyl ether-β-cyclodextrin as chiral selector. The concentration of the chiral selector and organic modifier, pH of background electrolyte (BGE), capillary temperature, and applied voltage were systematically optimized by using orthogonal design and concentration of chiral selector was further optimized. The optimal conditions included 25mM phosphate buffer at pH 8.0, containing 28mg/mL sulfobutyl ether-β-cyclodextrin and 20% acetonitrile as running buffer, an applied voltage of 22kV, and a temperature of 20°C. The detection wavelength was 206nm. The obtained method was capable of separating RS86017 from its potential chiral impurities, the S,R-enantiomer, the R,R-diastereomer and the S,S-diastereomer with a short analysis time of 10min. The separation was validated with respect to its selectivity, repeatability, linearity, precision, accuracy, limits of detection (LOD), limits of quantitation (LOQ) and robustness testing. The LODs and LOQs were 0.8μg/mL and 2.5μg/mL for all isomers of RS86017, respectively. Finally, the method was used to investigate the chiral purity of RS86017 in bulk samples., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Voltammetric behaviour and assay of nicorandil were investigated using square-wave and cyclic voltammetry. Voltammograms in Britton-Robinson (BR) buffer exhibited one well-defined and two merged reduction peaks. The influence of different buffers, electrolytes, pH, scan rates, and concentration of the drug on cathodic peak current was studied. On the basis of electrochemical behaviour of nicorandil, a direct square-wave voltammetric procedure for quantitation of nicorandil has been developed and validated. The proposed square-wave voltammetric method allows quantitation over the range 12.5-62.5 µg mL(-1) with correlation coefficient of 0.992. The limit of quantification and limit of detection were 21.95 µg mL(-1) and 6.58 µg mL(-1) , respectively. Precision and accuracy were also checked and found within limits. The developed square-wave method has also been successfully applied for the determination of nicorandil in pharmaceutical formulations., (Copyright © 2010 John Wiley & Sons, Ltd.)

Verapamyl substance and market verapamyl were investigated with circular dichroism. It was shown that circular dichroism provided rapid and highly efficient determination of the optic isomers and could be recommended as a method for control of drug quality.

Objective: To investigate the stability of tramadol hydrochloride 100mg associated with alizapride 50mg in 100ml of 5% dextrose solution stored at 5+/-3 degrees C., Methods: Solutions of 5% dextrose 100ml in polyolefin bags (n=5) containing approximately tramadol hydrochloride 100mg associated with alizapride 50mg were prepared under aseptic conditions and stored about 32 days at 5+/-3 degrees C. The tramadol hydrochloride and alizapride concentrations were measured by high-performance liquid chromatography (HPLC). Visual inspection was performed and pH was measured periodically during the storage. Stability of the solutions was defined as the common regression line 95% lower confidence limit of the concentration remaining superior to 90% of the initial concentration as recommended by the Food and Drug Administration (FDA)., Results: No color change or precipitation in the solutions was observed. Tramadol hydrochloride 100mg associated with alizapride 50mg in 100ml of 5% dextrose infusions was stable when stored at 5+/-3 degrees C during 32 days. Throughout this period, the lower confidence limit of the estimated regression line of concentration-time profile remained above 90% of the initial concentration. There was no significant change in pH during storage., Conclusion: Under the conditions of this study, tramadol hydrochloride 100mg associated with alizapride 50mg in 100ml of 5% dextrose infusions stored up to 32 days at 5+/-3 degrees C remain stable and may be prepared in advance by a Centralized Intravenous Additive Service (CIVAS) to improve safety and management., (Copyright 2010 Elsevier Masson SAS. All rights reserved.)

Two simple, accurate, and precise HPTLC methods have been established for the determination of mexiletine hydrochloride, an antiarrhythmic agent, in Mexicord capsules. Analyses were performed in horizontal chambers on RP C18F254s and normal-phase amino (NH2) HPTLC precoated plates with the mobile phases tetrahydrofuran-citrate buffer, pH 4.45 (3 + 7, v/v) and chloroform-tetrahydrofuran-hexane-ethylamine (3 + 2 + 5 + 0.1, v/v/v/v), respectively. The plates were developed for a distance of 40 mm in both cases. Densitometric measurements were achieved in the UV mode at 217 nm based on peak areas with semilinear calibration curves (R2 > or = 0.97) in the concentration range 0.5-8.0 microg/spot for the NH2 and C18 HPTLC methods. The elaborated chromatographic methods were validated in accordance with International Conference on Harmonization guidelines in terms of linearity, accuracy (99.64% for NH2 and 99.53% for C18), precision (intraday RSD 1.16 and 2.71%, respectively), sensitivity (LOD 0.1 microg/spot for both systems), and specificity.

A sensitive HPLC method combined with a column-switching system and tris(bipyridine)ruthenium(III) electrogenerated chemiluminescence (ECL) detection has been developed for the quantitative determination of mexiletine (MEX). MEX was derivatized by divinylsulfone (DVS) and then measured. The optimum conditions for the derivatization reaction were 10 µL of sample solutions, 40 mM DVS (pH 8.0), a reaction temperature of 50°C, and a reaction time of 15 min. The derivatized samples were cleaned up by an on-line pretreatment column. Also, after column-switching to the analytical column, the derivatized MEX was separated and detected. The calibration curves of MEX in human control serum showed good linear regression (r = 0.9996) from 0.008 to 6.56 µg ml(-1). The detection limit of MEX was 0.008 µg ml(-1) (S/N = 3). At a concentration of 2.0 µg ml(-1) MEX, the relative standard deviation (n = 5) was 0.98%. In this method the concentration of MEX in human control serum was readily measured, and this method was successfully applied to the time courses of the concentration of MEX in rabbit plasma after intravenous administration. The proposed method involved a simple and minimum sample-preparation procedure and a short run time (<20 min). Therefore it can be applied to routine therapeutic monitoring and pharmacokinetic studies of MEX.

A simple, sensitive and rapid flow injection analysis (FIA) method with resonance light scattering (RLS) was described for the determination of propafenone (PPF). The method was based on the ion-association reaction of 12-tungstophosphoric acid (TP) with propafenone. In pH 1.0 acidic medium, TP reacted with PPF to form an ion-associate complex, which resulted in a significant enhancement of RLS intensity. The maximum scattering peak was located at 340 nm, the RLS intensity was proportional to the concentration of PPF in the range 0.003-9.0 microg/mL, and the detection limit (3sigma) of 1.0 ng/mL was obtained at a sampling rate of 60 samples/h. The feasible reaction conditions and FIA parameters for the system were optimized. The method proposed in this paper shows satisfactory reproducibility with a relative standard deviation (RSD) of 2.1% for 10 successive determinations of 2.0 microg/mL PPF. The present method had been successfully applied to the determination of PPF in serum samples and pharmaceutical samples. The results obtained were in agreement with the method used in the Chinese Pharmacopoeia.

For the risk assessment of drug candidates, the identification and quantification of their metabolites is required. The majority of analytical techniques is based on calibration standards for quantification of the metabolites. As these often are not readily available, the use of inductively coupled plasma mass spectrometry (ICPMS) is an attractive alternative for drugs containing heteroatoms. In this work, the online coupling of electrochemistry (EC), liquid chromatography (LC), and ICPMS is presented. The antiarrhythmic agent amiodarone, which contains two iodine atoms, is oxidized in an electrochemical flow-through cell under N-dealkylation and deiodination. The metabolites that are generated at different EC potentials are identified by electrospray ionization (ESI) mass spectrometry, compared to those from rat liver microsomal incubations and quantified by ICPMS. Phase-optimized LC, a recent approach for high-performance isocratic separations, is used to avoid the ICPMS calibration problems known to occur with gradient separations. The potential of the complementary use of ESI-MS and ICPMS for the qualitative and quantitative analysis of drug metabolites becomes apparent in this work.

The first automated method for the determination of mexiletine hydrochloride - an antiarrhythmic agent - is reported. The method is based on the reaction of the analyte with o-phthalaldehyde (OPA) in the presence of sulfite in basic medium using a sequential injection (SI) manifold. The reaction product was monitored spectrofluorimetrically (lambda(ex)=350 nm/lambda(em)=446 nm). A simple and effective on-line dilution approach was adopted in order to expand the linearity and apply the method to assay, dosage uniformity and dissolution tests with minimum sample preparation. Chemical (pH, amount concentrations of OPA and sulfite) and instrumental variables (temperature, flow rate, injection volumes, etc.) that affected the determination were studied. The developed assay was validated in terms of linearity, range, limits of detection (LOD=3.4 mg L(-1)) and quantitation (LOQ=10 mg L(-1)), accuracy, precision (R.S.D.<3.4%) and selectivity. The method was applied successfully to the quality control of a mexiletine-containing formulation. The results were in good agreement with the US pharmacopoeia HPLC method.

The use of metabolites of antiarrhythmic drugs (ethmozine, ethacizine, and bonnecor) as haptens in the synthesis of conjugated antigens allowed us to induce the formation of antibodies with different specificity for certain metabolites. A new enzyme immunoassay was developed for the detection of phenothiazine and dibenzazepine derivatives (ethmozine, ethacizine, and bonnecor). Nanogram and subnanogram quantities of these substances may be detected in biological fluids.

Four simple, rapid, sensitive, and selective analytical procedures were developed for determination of mexiletine hydrochloride (MX) and/or its related substance: 2,6-dimethylphenol (DMP). The latter is a synthetic impurity for which a maximum pharmacopeial limit is defined. The first method depends on derivative-ratio spectrophotometry, for which the first-derivative signals of the ratio spectra at 259 nm (Deltalambda = 3 nm) are selected for the determination of MX. The second method is based on the spectrofluorometric measurement of MX in alkaline solution in the presence of 15 mM sodium dodecyl sulfate micellar medium at 292 nm (lambdaEx 260 nm). The third method is based on liquid chromatographic (LC) separation of MX and DMP on an RP-C8 column with a mobile phase consisting of 50 mM Na2HPO4-acetonitrile (60 + 40, adjusted to pH 2.4), and quantification of the analytes is achieved with UV detection at 212 nm based on peak area. The fourth method uses the coupling reaction of DMP with 2,6-dibromo-quinone-4-chlorimide (DBQC) in borate buffer to form an intensely colored product that was spectrophotometrically measured using first-derivative amplitudes at 670 nm (Deltalambda = 6 nm) for the determination of DMP. Different variables affecting each method were carefully investigated and optimized. The reliability and analytical performance of the proposed methods, including linearity, range, precision, accuracy, and detection and quantitation limits, were statistically validated. The first 3 methods were successfully applied for the stability-indicating determination of MX in laboratory-prepared mixtures with DMP, as well as for the determination of MX in capsules. Also, the LC and the DBQC spectrophotometric methods permitted the selective determination of DMP in the presence of a large excess of the parent drug at or near the pharmacopeial limit (0.1-1%).

We have developed a lidless micropillar array electrospray ionization chip (microPESI) combined with mass spectrometry (MS) for analysis of drugs and biomolecules. The microPESI chip, made of silicon, contains a sample introduction spot for a liquid sample, an array of micropillars (diameter, height, and distance between pillars in the range of 15-200, 20-40, and 2-80 microm, respectively), and a sharpened tip for direct electrospray formation. The microchips were fabricated using deep reactive ion etching (DRIE) which results in accurate dimensional control. The chip, providing a reliable open-channel filling structure based on capillary forces and a electrospray emitter tip for ionization, allows an easy operation and reliable, non-clogging liquid transfer. The microPESI chip can be used for a fast analysis using single sampling or for continuous infusion measurements using a syringe pump for sample introduction. The microPESI-MS shows high sensitivity, with limit of detection 30 pmol/L (60 amol or 28 fg) for verapamil measured with tandem mass spectrometry (MS/MS) and using a sample volume of 2.5 microL. The system shows also good quantitative linearity (r2 > 0.99) with linear dynamic range of at least six orders of magnitude and good ion current stability (standard deviation <5%) in 1-h continuous flow measurement. The microPESI-MS is shown to be a very potential method for direct analysis of drugs and biomolecules., (Copyright 2007 John Wiley & Sons, Ltd.)

Extracts of hawthorn (Crataegus oxycantha) have become popular herbal supplements for their well-recognized cardiotonic effects. Many commercial preparations have been used successfully in the treatment of congestive heart failure, although the active principles within these extracts have yet to be conclusively identified. Several hawthorn preparations were studied and found to have negative chronotropic effects in a cultured neonatal murine cardiomyocyte assay using unpaced cells. As compared to conventional cardiac drugs (i.e., epinephrine, milrinone, ouabain, or propranolol), hawthorn extract has a unique activity profile. Hawthorn extract appears to be anti-arrhythmic and capable of inducing rhythmicity in quiescent cardiomyocytes. Hawthorn extract does not cause beta-adrenergic receptor blockade at concentrations which cause negative chronotropic effects. Commercial hawthorn preparations, extracts prepared from dried leaves and those made from dried berries have similar chronotropic activities. When crude extracts are separated using size-exclusion chromatography, several fractions retain multiple cardiac activities. Assays with chromatographic fractions reveal that multiple dissimilar cardioactive components may exist within the extract, making the identification of individual active constituents more challenging.

Four methods were developed for the determination of intact disopyramide phosphate in the presence of its degradation product. In the first and second methods, third-derivative spectrophotometry and first derivative of the ratio spectra were used. For the third-derivative spectrophotometric method, the peak amplitude was measured at 272 nm, while for the derivative ratio spectrophotometric method, disopyramide phosphate was determined by measuring the peak amplitude at 248 and 273 nm. Both methods were used for the determination of disopyramide phosphate in the concentration range 12.5-87.5 microg/mL, with corresponding mean recovery 100.8 +/- 0.7% for the first method and 99.9 +/- 0.7% and 99.6 +/- 0.7% for the second method at 248 and 273 nm, respectively. In the third method, an ion selective electrode (ISE) was fabricated using phosphotungstic acid as an anionic exchanger, PVC as the polymer matrix, and dibutylsebacate as a plasticizer. The ISE was used for the determination of disopyramide phosphate in pure powder form in the concentration range 10(-2)-10(-5) M. The slope was found to be 58.5 (mV/decade), and the average recovery was 99.9 +/- 1.6%. The fourth method depended on the quantitative densitometric determination of the drug in concentration range of 0.25-2.5 microg/spot using silica gel 60 F245 plates and ethyl acetate-chloroform-ammonium hydroxide (85 + 10 + 5, v/v/v) as the mobile phase, with corresponding mean accuracy of 100.3 +/- 1.1%. The 4 proposed methods were found to be specific for disopyramide phosphate in presence of up to 80% of its degradation product for the spectrophotometric methods, 90% of its degradation for the densitometric method, and 40% for the ISE method. The 4 proposed procedures were successfully applied for the determination of disopyramide phosphate in Norpace capsules. Statistical comparison between the results obtained by these methods and the official method of the drug was done, and no significant differences were found.

Capillary electrophoresis with electrochemical detection has been employed for the determination of honokiol and magnolol in Cortex Magnoliae Officinalis (i.e. Magnolia Bark) for the first time. Effects of several important factors such as the concentration and the acidity of the running buffer, separation voltage, injection time, and detection potential were investigated to acquire the optimum conditions. The detection electrode was a 300 microm diameter carbon disc electrode at a working potential of +0.90 V (versus saturated calomel electrode (SCE)). The two analytes can be well separated within 6 min in a 40 cm length fused silica capillary at a separation voltage of 18 kV in a 50mM borate buffer (pH 9.2). The relation between peak current and analyte concentration was linear over about three orders of magnitude with the detection limits (S/N=3) of 0.38 and 0.51 microM for honokiol and magnolol, respectively. The proposed method has been successfully applied to monitor the two bioactive constituents in the real plant samples with satisfactory assay results.

A robust, inexpensive, and fully validated CE method for the simultaneous determination of the enantiomers of propafenone (PPF), 5-hydroxy-propafenone (5OH-PPF) and N-despropyl-propafenone (NOR-PPF) in serum and in in vitro media is described. It is based upon liquid-liquid extraction at alkaline pH followed by analysis of the reconstituted extract by CE in presence of a pH 2.0 running buffer composed of 100 mM sodium phosphate, 19% methanol, and 0.6% highly sulfated beta-CD. For each compound, the S-enantiomers are shown to migrate ahead of their antipodes, and the overall run time is about 30 min. Enantiomer levels between 25 and 1000 ng/mL provide linear calibration graphs, and the LOD for all enantiomers is between 10 and 12 ng/mL. The assay is shown to be suitable for the determination of the enantiomers of PPF and its metabolites in in vitro incubations comprising human liver microsomes or single CYP450 enzymes (SUPERSOMES). Incubations with CYP2D6 SUPERSOMES revealed, for the first time, the simultaneous formation of the enantiomers of 5OH-PPF and NOR-PPF with that enzyme. CE data can be used for the evaluation of the enzymatic N-dealkylation and hydroxylation rates.

The paper presents the results of an analytical evaluation and a study of some physicochemical properties of a potential antiarrhythmic agent marked as 6f. The melting point, spectral properties, solubility and chromatographic behaviour of the drug on the thin layer were investigated. For the content determination, spectrophotometry in the ultraviolet region of the spectrum at the wavelength of the second absorption maximum of the substance and high-performance liquid chromatography were used.

A first order derivative UV-spectrophotometric method for the determination of diltiazem hydrochloride and mexiletine hydrochloride has been developed and validated. In the assay, the first- and second-order measurements with the use of the "peak-zero" and "peak-peak" techniques were applied. The linear correlation (r < 0.9999) between the amplitude of the peak and the concentration of the examined drugs in the range of 3.0-8.0 microg mL(-1) for diltiazem and 50-100 microg mL(-1) for mexiletine was obtained. The proposed method was successfully applied for accurate (mean recovery about 100%), precise (RSD about 1%) and selective determination of the studied drug in the pure and dosage forms.

Two simple, sensitive and economical spectrophotometric methods have been developed for the determination of amiodarone hydrochloride in pure form and commercial dosage form. These methods (A and B) are based on the reaction of amiodarone base as n-electron donor with p-chloranilic acid and 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) as pi-acceptors to give highly colored complex species which absorb maximally at 535 and 570 nm, respectively. Beer's law is obeyed in the concentration ranges 10.0 - 360.0 and 2.0 - 65.0 microg ml(-1) for methods A and B, respectively. Application of the proposed methods to commercial pharmaceutical tablets are presented.

The death of a 76-year-old man with heart disease as a result of the injection of an excessive dose of lidocaine is presented. The patient was given 5 ml of 10% lidocaine hydrochloride (500 mg) intravenously instead of 2.5 ml of 2% lidocaine hydrochloride (50mg) in order to treat repeated paroxysmal ventricular arrhythmia. Immediately following the injection the patient had tonic clonic seizures and complete cardiopulmonary arrest followed. Although resuscitation attempts once successfully restarted his pulse and spontaneous respiration, the patient died on the eighth day after the injection. Toxicological examinations were carried out on the tissues obtained at the time of autopsy and which had been fixed in formalin solution for 40 days, and lidocaine was detected in each tissue examined. The concentrations were (ng/g or ml): parietal lobe, 308.0; occipital lobe, 208.7; temporal lobe, 318.0; frontal lobe, 223.2; cerebellum 200.9; pons 285.7; liver, 109.5; kidney 52.2; skeletal muscle 127.0; and formalin solution 8.4. In an experiment on rats we determined the concentration changes of lidocaine in formalin fixed tissues. The concentrations of lidocaine in these tissues significantly decreased to 1/3-1/4 from the original. This data shows that the cause of death was poisoning by lidocaine overdose.

A few days after her admittance to a hospital for a suicide attempt with benzodiazepines, a 15-year-old girl was found dead in bed. At autopsy, no specific anatomo-pathologic cause of death was identified. Systematic toxicological analysis (HPLC-DAD, GC-NPD, and GC-MS) of postmortem blood and urine revealed the presence of high concentrations of flecainide and its two major metabolites. Flecainide is a class IC anti-arrhythmic drug causing a decreased intracardiac conduction velocity in all parts of the heart. To identify and quantitate flecainide together with its metabolites in blood, urine, and other toxicologically relevant matrices, a new method was developed using high-performance liquid chromatography with diode-array detection. All compounds were separated on a Hypersil BDS phenyl column using water, methanol, and 1.5M ammonium acetate in a gradient system. Chromatographic analysis was preceded by an optimized solid-phase extraction procedure on RP-C18 extraction columns. The flecainide concentrations in blood and urine were 18.73 and 28.3 mg/L, respectively, and the metabolites were detected only in urine at the following concentrations: 9.4 mg/L for meta-O-dealkylated flecainide and 8.59 mg/L for meta-O-dealkylated flecainide lactam. Based on these results, it was concluded that the suicide was consistent with an overdose of this anti-arrhythmic drug.

This review reports the latest achievements in chromatographic enantioseparations of various classes of cardiovascular drugs and selected applications of these methods in pharmaceutical and clinical analysis. The use of these drugs as test compounds for new chiral stationary phases and different parameters of chromatographic processes is also presented., (Copyright 2002 Elsevier Science B.V.)

We report a method for measuring the concentration of flecainide in hair. An animal study, in which flecainide (1, 5, and 10 mg/kg/day) was orally administered for 1, 2, and 3 weeks to pigmented rats, showed that flecainide concentration in rat hairs newly regrown after administration significantly correlated with both the daily dose and the dosing period. The part of hair containing flecainide continued to grow upward, retaining the drug within the hair structure that had been formed at the time of drug exposure. Flecainide was also determined in human scalp hairs collected from patients treated with flecainide. The drug content of white hairs was much less than that black hairs collected from the same rats and subjects, suggesting the determinant effect of hair pigment on flecainide accumulation in hair. These findings suggest that the analysis of flecainide in hair may be useful for assessing exposure to drug qualitatively., (Copyright 2001 Wiley-Liss, Inc. and the American Pharmaceutical Association)

A sensitive and specific ELISA for an antiarrhythmic drug, pilsicainide, was developed, which is capable of measuring as low as 1.6 ng/ml. Anti-pilsicainide antibody was obtained by immunizing rabbits with pilsicainide conjugated with bovine serum albumin using diazotized N-(3-amino-2,6-dimethylphenyl)-8-pyrrolizidinylacetamide (3-aminopilsicainide). Enzyme labeling of pilsicainide with beta-D-galactosidase was similarly performed using a diazotized 3-aminopilsicainide. Cross-reactivity data showed that the antibody well recognizes both the aromatic ring and the pyrrolizidine moieties, and thus specific enough to the structure of pilsicainide. The values for the pilsicainide concentrations detected using this assay were comparable with those detected using HPLC. There was a good correlation between the values determined by the two methods. Moreover, the ELISA was about 30-fold more sensitive in detecting pilsicainide at lower concentrations. Using this assay, drug levels were easily measured in the serum of rabbits after oral administration of pilsicainide at a single dose of 1 mg/kg. The ELISA should be a valuable tool in therapeutic drug monitoring (TDM) and pharmacokinetic studies of pilsicainide.

Using the high performance liquid chromatography/atmospheric pressure chemical ionization tandem mass spectrometry (HPLC/APCI-MS/MS) technique, together with established trends from the literature, the structures of metabolites and impurities of amiodarone, an anti-arrhythmic drug, have been assigned. By comparing analyses of products of incubation with rat liver microsomes with controls in which glucose 6-phosphate dehydrogenase was omitted, metabolites could be distinguished from impurities. Structures for the two proposed metabolites and four impurities are proposed.

A fatal case attributed to flecainide acetate (Tambocor), a class Ic antiarrythmic drug, is presented. Flecainide was detected by GC/MS in gastric contents, blood and liver as well. The urine analysis revealed the presence of its dealkylated metabolite. Body fluids and tissue concentrations determined by GC/ECD were 7.7 mg/kg in femoral blood, 0.26 mg/kg in bile, 18 mg/kg in liver, 0.17 mg/kg in cerebrospinal fluid, 0.22 mg/kg in brain cortex and 28.9 mg/kg in urine. The total amount of flecainide in gastric contents was about 43 mg. Even taking into account the postmortem redistribution of flecainide, its blood level still remains in the toxic range.

The in vitro metabolism of RWJ-34130, an antiarrhythmic agent, was conducted using rat hepatic 9000 x g supernatant (S9) and microsomes in an NADPH-generating system, and the rat liver perfusion. The 100 and 20 microg ml(-1) concentrations of RWJ-34130 aqueous solution were used for microsomal incubation and liver perfusion, respectively. Unchanged RWJ-34130 (approximately 77-78% of the sample in both S9 and microsomes) plus a major metabolite, RWJ-34130 sulfoxide (20% of the sample in both S9 and microsomes) were profiled, isolated and identified from both hepatic S9 and microsomal incubates (60 min) using HPLC and mass spectrometry (MS), and by comparison to a synthetic RWJ-34130 sulfoxide, which was synthesized by reacting RWJ-34130 with MCPBA (meta-chloroperoxy benzoic acid). No unchanged RWJ-34130 was detected in the 3 h liver perfusate, however, 1-phenyl-2-oxo-pyrrolidine was profiled, isolated and identified as a major hydrolyzed metabolite of liver perfusate. RWJ-34130 is not extensively metabolized in vitro in rat hepatic S9 and microsomes. All HPLC metabolic profiles of hepatic S9 and microsomal samples (30 min, 60 min) were qualitatively and nearly quantitatively identical.

Purpose: To study the partitioning of model acids ((RS)-warfarin and salicylic acid), and bases (lidocaine, (RS)-propranolol and diazepam), with immobilized artificial membrane (IAM)-HPLC, as compared to partitioning in the standardized phosphatidylcholine liposome/buffer system., Methods: The pH-dependent apparent partition coefficients D were calculated from capacity factors (k'IAM) obtained by IAM-HPLC, using a 11-carboxylundecylphosphocholine column. For lipophilic compounds k'IAM, values were determined with organic modifiers and extrapolation to 100% water phase (k'IAMw) was optimized. Temperature dependence was explored (23 to 45 degrees C), and Gibbs free energy (deltaG), partial molar enthalpy (deltaH) and change in entropy (deltaS) were calculated. Equilibrium dialysis was used for the partitioning studies with the liposome/buffer system., Results: For extrapolation of k'IAMw, linear plots were obtained both with the respective dielectric constants and the mole fractions of the organic modifier. All tested compounds showed a similar pH-D diagram in both systems; however, significant differences were reproducibly found in the pH range of 5 to 8. In all cases, deltaG and deltaH were negative, whereas deltaS values were negative for acids and positive for bases., Conclusions: In both partitioning systems, D values decreased significantly with the change from the neutral to the charged ionization state of the solute. The differences found under physiological conditions, i.e. around pH 7.4, were attributed to nonspecific interactions of the drug with the silica surface of the IAM column.