Reduced graphene oxide as an efficient sorbent in microextraction by packed sorbent: Determination of local anesthetics in human plasma and saliva samples utilizing liquid chromatography-tandem mass spectrometry.

Herein, reduced graphene oxide (RGO) has been utilized as an efficient sorbent in microextraction by packed sorbent (MEPS). The combination of MEPS and liquid chromatography-tandem mass spectrometry has been used to develop a method for the extraction and determination of three local anesthetics (i.e. lidocaine, prilocaine, and ropivacaine) in human plasma and saliva samples. The results showed that the utilization of RGO in MEPS could minimize the matrix effect so that no interfering peaks at the retention times of the analytes or internal standard was observed. The high extraction efficiency of this method was approved by mean recoveries of 97.26-106.83% and 95.21-105.83% for the studied analytes in plasma and saliva samples, respectively. Intra- and inter-day accuracies and precisions for all analytes were in good accordance with the international regulations. The accuracy values (as percentage deviation from the nominal value) of the quality control samples were between -2.1 to 13.9 for lidocaine, -4.2 to 11.0 for prilocaine and between -4.5 to -2.4 for ropivacaine in plasma samples while the values were ranged from -4.6 to 1.6 for lidocaine, from -4.2 to 15.5 for prilocaine and from -3.3 to -2.3 for ropivacaine in human saliva samples. Lower and upper limit of quantification (LLOQ, ULOQ) were set at 5 and 2000 nmol L-1 for all of the studied drugs. The correlation coefficients values were ≥0.995. The limit of detection values were obtained 4 nmol L-1 for lidocaine and prilocaine, and 2 nmol L-1 for ropivacaine.

Continuous purification of active pharmaceutical ingredients utilizing polymer membrane surface wettability.

Continuous processing of pharmaceuticals opens opportunities for continuous separation based on wettability of polymer membranes. Dual use of hydrophobic and hydrophilic membranes realize in-line liquid-liquid extraction in the synthesis of four essential APIs. A secondary membrane with opposite wetting characteristics proves critical to phase separation of aqueous-organic reaction streams.

Development, Validation and Application of a Stability Indicating HPLC Method to Quantify Lidocaine from Polyethylene-co-Vinyl Acetate (EVA) Matrices and Biological Fluids.

An efficient and cost-effective quantification procedure for lidocaine by HPLC has been developed to estimate lidocaine from an EVA matrix, plasma, peritoneal fluid and intra-articular fluid (IAF). This method guarantees the resolution of lidocaine from the degradation products obtained from alkaline and oxidative stress. Chromatographic separation of lidocaine was achieved with a retention time of 7 min using a C18 column with a mobile phase comprising acetonitrile and potassium dihydrogen phosphate buffer (pH 5.5; 0.02 M) in the ratio of 26:74 at a flow rate of 1 mL min-1 with detection at 230 nm. Instability of lidocaine was observed to an oxidizing (0.02% H2O2) and alkaline environments (0.1 M NaOH). The calibration curve was found to be linear within the concentration range of 0.40-50.0 µg/mL. Intra-day and inter-day accuracy ranged between 95.9% and 99.1%, with precision (% RSD) below 6.70%. The limit of quantification and limit of detection were 0.40 µg/mL and 0.025 µg/mL, respectively. The simple extraction method described enabled the quantification of lidocaine from an EVA matrix using dichloromethane as a solvent. The assay and content uniformity of lidocaine within an EVA matrix were 103 ± 3.60% and 100 ± 2.60%, respectively. The ability of this method to quantify lidocaine release from EVA films was also demonstrated. Extraction of lidocaine from plasma, peritoneal fluid and IAF followed by HPLC analysis confirmed the utility of this method for ex vivo and in vivo studies where the calibration plot was found to be linear from 1.60 to 50.0 µg/mL.

Application of graphitic sorbent for online microextraction of drugs in human plasma samples.

In the present work a new sorbent based on graphitized carbon (CarbonX(®) COA) was evaluated in microextraction by packed sorbent (MEPS) for extraction of lidocaine and ropivacaine from human plasma samples. The new graphitic sorbent showed high recoveries of lidocaine and ropivacaine compared to C18 sorbent. In the present study the G-MEPS (syringe packed with graphitic sorbent) was connect online with liquid chromatography tandem mass spectrometry (LC-MS/MS). In order to obtain a fast and reliable method different factors affecting MEPS performance were investigated. The extraction efficiency of the graphitic sorbent was compared with silica-based sorbents used in MEPS. The G-MEPS was also evaluated for reuse (50-100 times). The recoveries of lidocaine and ropivacaine from plasma samples were 79% and 82%; respectively. The method was validated according to FDA (Food and Drug Administration) guideline for bioanalytical method validation. Linearity was assessed in the range 5-2000nmol/L, with coefficient of determination r(2)>0,995 (n=3) for lidocaine and r(2)>0.997 (n=3) for ropivacaine. The lower limit of quantification (LLOQ) was 5nmol/L and the limit of detection (LOD) was 1nmol/L for studied analytes in plasma samples. For both analytes considered in this study the accuracy values in plasma samples were ranged from 86% to 113%. The Inter-day precisions, expressed as relative standard deviation (%RSD), at three different concentrations (QC-samples) ranged from 8% to 9% for lidocaine, and from 4% to 11% for ropivacaine.

Liquid-phase microextraction based on two immiscible organic solvents followed by gas chromatography with mass spectrometry as an efficient method for the preconcentration and determination of cocaine, ketamine, and lidocaine in human urine samples.

A new type of liquid-phase microextraction based on two immiscible organic solvents was optimized and validated for the quantification of lidocaine, ketamine, and cocaine in human urine samples. A hollow-fiber based microextraction technique followed by gas chromatography coupled with mass spectrometry detection was used to reduce matrix interferences and improve limits of detection. The analytes were extracted from aqueous sample with pH 11.0, into a thin layer of organic solvent (n-dodecane) sustained in the pores of a hollow fiber, and then into a second organic acceptor (acetonitrile) located inside the lumen of the hollow fiber. With the application of optimized values, good linearity was obtained in the range of 1-500 µg/L for lidocaine and ketamine and 2-500 µg/L for cocaine with the determination coefficient values (r(2) ) >0.9943. The preconcentration factors and limits of detection (S/N > 3) were 250-350 and 0.01-0.05 µg/L, respectively. Intra and interassay precision values were <7.3 and 9.3%, respectively. The method was successfully applied for the determination and quantification of target analytes in human urine samples.

Molecularly imprinted polymer in microextraction by packed sorbent for the simultaneous determination of local anesthetics: lidocaine, ropivacaine, mepivacaine and bupivacaine in plasma and urine samples.

This study presents the use of molecularly imprinted polymer (MIP) as packing material for microextraction by packed syringe (MEPS) to achieve higher extraction selectivity. Pentycaine was used as template for MIP. Development and validation of the determination of lidocaine, ropivacaine, mepivacaine and bupivacaine in human plasma and urine samples utilizing MIP-MEPS and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were carried out. The MEPS MIP-cartridge could be used for 100 extractions before it was discarded. The extraction recovery ranged from 60 to 80%. The correlation coefficients values were >0.999 for all assays using lidocaine, ropivacaine, mepivacaine and bupivacaine in the calibration range 5-2000 nmol/L. The accuracy of the studied compounds, given as a percentage variation from the nominal concentration values, ranged from -4.9 to 8.4% using plasma and urine samples. The between-batch precision, given as the relative standard deviation, at three different concentrations (quality control samples) was ranged from -4.7 to 14.0% and from 1.8 to 12.7% in plasma and urine, respectively. The lower limit of quantification and limit of detection of the studied substances were 5.0 and 1.0 nm, respectively.

Screening and determination of drugs in human saliva utilizing microextraction by packed sorbent and liquid chromatography-tandem mass spectrometry.

This study presents a new method for collecting and handling saliva samples using an automated analytical microsyringe and microextraction by packed syringe (MEPS). The screening and determination of lidocaine in human saliva samples utilizing MEPS and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were carried out. An exact volume of saliva could be collected. The MEPS C8 -cartridge could be used for 50 extractions before it was discarded. The extraction recovery was about 60%. The pharmacokinetic curve of lidocaine in saliva using MEPS-LC-MS/MS is reported.

Automated analysis of lidocaine and its metabolite in plasma by in-tube solid-phase microextraction coupled with LC-UV for pharmacokinetic study.

A sensitive, selective, and reproducible in-tube solid-phase microextraction and liquid chromatographic (in-tube SPME/LC-UV) method for determination of lidocaine and its metabolite monoethylglycinexylidide (MEGX) in human plasma has been developed, validated, and further applied to pharmacokinetic study in pregnant women with gestational diabetes mellitus (GDM) subjected to epidural anesthesia. Important factors in the optimization of in-tube SPME performance are discussed, including the draw/eject sample volume, draw/eject cycle number, draw/eject flow rate, sample pH, and influence of plasma proteins. The limits of quantification of the in-tube SPME/LC method were 50 ng/mL for both metabolite and lidocaine. The interday and intraday precision had coefficients of variation lower than 8%, and accuracy ranged from 95 to 117%. The response of the in-tube SPME/LC method for analytes was linear over a dynamic range from 50 to 5000 ng/mL, with correlation coefficients higher than 0.9976. The developed in-tube SPME/LC method was successfully used to analyze lidocaine and its metabolite in plasma samples from pregnant women with GDM subjected to epidural anesthesia for pharmacokinetic study.

Development of an on-column enrichment technique based on C18-functionalized magnetic silica nanoparticles for the determination of lidocaine in rat plasma by high performance liquid chromatography.

In this study, a novel on-column enrichment technique filled with C(18)-functionalized magnetic silica nanoparticles was successfully developed for the determination of lidocaine in rat plasma by high performance liquid chromatography (HPLC). The synthesized Fe(3)O(4)@SiO(2)-C(18) nanoparticles were locally packed into the capillary by the application of magnets. Lidocaine in the sample solutions pumped into the capillary tube could be easily adsorbed by Fe(3)O(4)@SiO(2)-C(18) through hydrophobic interaction by the interior C(18) groups, and eluted by acetonitrile solution. Different extraction conditions were investigated. Method validations including linear range, quantification limit, detection limit, precision, accuracy and recovery were also studied. The results showed that the proposed method based on on-column enrichment by Fe(3)O(4)@SiO(2)-C(18) was a novel, little solvent and efficient approach for the determination of lidocaine in the complex plasma samples.

HPLC-DAD stability indicating determination of nitrofurazone and lidocaine hydrochloride in their combined topical dosage form.

In this work, a simple, rapid, and selective high-performance liquid chromatography (HPLC) method with diode array detection was developed for the simultaneous determination of nitrofurazone (NZ) and lidocaine hydrochloride (LD). The chromatographic separation was achieved by using Zorbax Eclipse XDB-C(18) (4.6 x 150 mm, 5 mum p.s.) analytical column and a mobile phase composed of 0.025 M disodium hydrogen phosphate-methanol-triethylamine (70:30:0.1, v/v/v) (pH 4.0) at a flow rate of 1 mL/min. The detector was set at wavelengths 374 and 220 nm for NZ and LD, respectively, and quantification of the analytes was based on measuring their peak areas. The retention times for NZ and LD were approximately 4.5 and 5.7 min, respectively. The reliability and analytical performance of the proposed HPLC procedure were statistically validated with respect to system suitability, linearity, ranges, precision, accuracy, selectivity, robustness, and detection and quantification limits. The linear dynamic ranges were 0.5-25 and 2.5-100 mug/mL for NZ and LD, respectively, with correlation coefficients > 0.999. The stability-indicating aspects of the proposed method were demonstrated by the resolution of the two analytes from the related substance and potential impurity (2,6-dimethylaniline) as well as from forced-degradation products. The validated HPLC method was successfully extended to the analysis of the combined topical dosage form (soluble dressing) where no interfering peaks were encountered from the dosage form matrix or the inactive ingredients.

[Determination of monoethylglycinexylidide concentration in serum using solid phase extraction and capillary gas chromatography-mass spectrometry].

A novel method for the determination of monoethylglycinexylidide (MEGX) (lidocaine metabolin) in serum using solid phase extraction (SPE) and capillary gas chromatography-mass spectrometry (GC-MS) was established. The serum sample was extracted with a CN-SPE column. An HP-5MS capillary column (15 m x 0.25 mm x 0.1 microm) was used. The initial temperature of the column was set at 100 degrees C, held for 1 min, then raised to 200 degrees C at 40 degrees C/min, and held at 200 degrees C for 0.5 min. The sample size was 2 microL, and the split ratio was set at 1 : 1. The carrier gas was high purity helium with a flow rate of 1.0 mL/min. The monitoring ions for the determination were m/z 58 for MEGX and m/z 86 for procaine (internal standard). The calibration curve of MEGX had good linear relationship in the range of 1.562 - 25 ng/mL ( r = 0.998 1). The limit of detection was 0.5 ng/mL. The extraction recovery ranged from 80.1% to 85.7%. The method advanced the quantitative analysis of MEGX in serum by combining rapid and efficient SPE with specific and sensitive quantitation by GC-MS.

Determination of difenidol hydrochloride by capillary electrophoresis with electrochemiluminescence detection.

A novel and sensitive method for the determination of difenidol hydrochloride has been established using capillary electrophoresis coupled with end-column electrogenerated chemiluminescence (ECL) detection, based on the ECL reaction of tris(2,2'-bypyridine)ruthenium(II) (Ru(bpy)(3)(2+)) with the tertiary amino groups of the difenidol analyte. Parameters that affect separation and detection were optimized. Calibration curve was linear over the range from 1 x 10(-6)M to 6 x 10(-5)M with a detection limit of 1 x 10(-7)M (S/N=3). Separation of difenidol hydrochloride from clomifene citrate and lidocaine was achieved using the proposed method. This method was successfully utilized to the assay of the active ingredients of the "difenidol hydrochloride" tablets and to the investigation on the interaction of difenidol hydrochloride with hemoglobin. The number of binding sites and the binding constant were estimated as (11.2 and 2.5) x 10(3)M(-1), respectively.

Decreasing analysis time in capillary electrophoresis: validation and comparison of quantitative performances in several approaches.

Capillary electrophoresis (CE) can be used for the rapid determination of pharmaceuticals, particularly in routine quality control analysis. This paper focuses on several approaches aimed at decreasing the analysis time with commercially available instrumentation by (i) application of a high electric field through a reduced capillary, (ii) use of a dynamically coated capillary to increase the electroosmotic flow, (iii) short-end injection (SEI) technique, and (iv) application of multiple sample injections. Moreover, SEIs were combined with the three other approaches. A pharmaceutical formulation containing lidocaine as an active component was selected, and the methods were validated according to the ICH guidelines. The seven approaches investigated fulfilled different statistical requirements and demonstrated their linearity and trueness, with good recoveries and confidence limits always inferior to 1.5%. Furthermore, relative standard deviation (RSD) values for repeatability and intermediate precision were inferior to 1.1 and 1.8%, respectively. These results confirmed that each approach is of utmost interest to increase the analyte throughput in CE.

Retention of ionizable compounds on HPLC. 8. Influence of mobile-phase pH change on the chromatographic retention of acids and bases during gradient elution.

The relationships between retention and mobile-phase pH in gradient elution are studied for acids and bases. The apparent pH shift caused by the increasing amount of acetonitrile and methanol has been determined starting from a wide range of pH values. It is shown that good relationships between the retention of ionizable compounds and the pH of the aqueous buffer can be established if the same type of buffer (ammonium acetate in this work) is used for all pH points. Equations are proposed to fit the gradient retention data to the pH of the aqueous buffer. The proposed equation gives an account of the relative variation of the pKa of the compound in the reference to the variation of the pH of the buffer as both parameters change during gradient elution.

Separation of lidocaine and its metabolites by capillary electrophoresis using volatile aqueous and nonaqueous electrolyte systems.

The separation of the basic drug lidocaine and six of its metabolites has been investigated both by using volatile aqueous electrolyte system, at low pH and by employing non-aqueous electrolyte systems. In aqueous systems, the best separation of the compounds under the investigated conditions was achieved by using the electrolyte 60 mM trifluoroacetic acid (TFA)/triethylamine (TEA) at pH 2.5 containing 15% methanol. With this electrolyte, all seven compounds were well separated with high efficiency and migration time repeatability. The separations with bare fused-silica capillaries and polyacrylamide-coated capillaries were compared with higher separation efficiency with the latter. On the other hand, near baseline separation of all the seven compounds was also obtained by employing the non-aqueous electrolyte, 40 mM ammonium acetate in methanol and TFA (99:1, v/v), with comparable migration time repeatability but lower separation efficiency relative to the aqueous system.

Coupling device for desorption of drugs from solid-phase extraction-pipette tips and on-line gas chromatographic analysis.

Solid-phase extraction-pipette tips were used for micro solid-phase extraction of lidocaine and diazepam. Off-line desorption was done after in-vial collection for reference purposes, whereas with on-line desorption the eluate was directly introduced in the gas chromatograph. With both methods the total eluate (100 microl) was introduced into the GC system, which was equipped with a programmed-temperature vaporiser (PTV) for large volume injection. For on-line desorption a laboratory-made coupling device was developed to connect the pipette tips with the injector of the PTV. The coupling device was applied successfully since no leakage occurred at the connection of the coupling device and the pipette tip. No significant differences in recovery of lidocaine and diazepam and in presence of impurities were observed between chromatograms obtained with either off-line or on-line desorption. Preliminary experiments with standard solutions showed recoveries of about 75% for a concentration level of 1 microg/ml. The system seems particularly suitable for high-throughput analysis.

[The differential diagnosis of fatal outcomes in patient treatment with lidocaine]. / Differentsial'naia diagnostika letal'nykh iskhodov pri lechenii bol'nykh lidokainom.

A method for isolation, identification, and measurement of lidocaine is proposed for investigations of biological specimens. The presence of lidocaine is confirmed by common alkaloid reactions, chromatography in a thin layer of adsorbent, and color test. Lidocaine concentration is measured by extraction photoelectrocolorimetry. A formula is proposed, permitting a tentative assessment of the injected lidocaine dose. The threshold identification value is 0.2 mg%, threshold detection value 0.5 mg%.

Chromatographic (TLC) analysis of diltiazem in pharmaceutical form in presence of other antiarrhythmics.

Diltiazem was identified by TLC method on silica gel by ascending technique, in presence of five another antiarrhythmic drugs (disopyramide, flecainide, lidocaine, lorcainide, procainamide). Suitable conditions for separation of diltiazem were established using ethanol-benzene-dioxane-ammonia (2:20:16:3) as the mobile phase. When using acidified iodoplatinate solution or Dragendorff reagent as indicators, the amount of diltiazem as small as 100 ng can be identified. The TLC method is simple and sensitive and it was used to identify diltiazem in tablets.

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.

[Amino acid derivatives of 2,6-xylidine and p-aminobenzoic acid. II. Synthesis of derivatives of N-(p-aminobenzoylaminoalkylacyl)-2,6-xylidines and study of their resistance to proteolytic factors]. / Aminokwasowe pochodne 2,6-ksylidyny i kwasu p-aminobenzoesowego. II. Otrzymywanie pochodnych N-(p-aminobenzoiloaminoalkiloacylo)-2,6-ksylidyn i badanie ich odpornosci na dzialanie czynników proteolitycznych.

Syntheses of N-(p-aminobenzoylaminoalkylacyl)-2,6-xylidines are described. The carbodiimide method and the method of mixed anhydrides were employed for the syntheses. Physicochemical properties, yields, data of elementary analysis and IR spectra of eighteen novel compounds are given. Resistance of some obtained compounds to actions of the gastric digestive juice and the intestinal one was tested.