Simultaneous estimation of acetylsalicylic acid and clopidogrel bisulfate in pure powder and tablet formulations by high-performance column liquid chromatography and high-performance thin-layer chromatography

This paper describes validated high-performance column liquid chromatographic (HPLC) and high-performance thin-layer chromatographic (HPTLC) methods for simultaneous estimation of acetylsalicylic acid (ASA) and clopidogrel bisulfate (CLP) in pure powder and formulations. The HPLC separation was achieved on a Nucleosil C8 column (150 mm length x 4.6 mm id, 5 microm particle size) using acetonitrile-phosphate buffer, pH 3.0 (55 + 45, v/v) mobile phase at a flow rate of 1.0 mL/min at ambient temperature. The HPTLC separation was achieved on an aluminum-backed layer of silica gel 60F254 using ethyl acetate-methanol-toluene-glacial acetic acid (5.0 + 1.0 + 4.0 + 0.1, v/v/v/v) mobile phase. Quantitation was achieved with UV detection at 235 nm over the concentration range 4-24 microg/mL for both drugs, with mean recoveries of 99.98 +/- 0.28 and 100.16 +/- 0.66% for ASA and CLP, respectively, using the HPLC method. Quantitation was achieved with UV detection at 235 nm over the concentration range of 400-1400 ng/spot for both drugs, with mean recoveries of 99.93 +/- 0.55 and 100.21 +/- 0.83% for ASA and CLP, respectively, using the HPTLC method. These methods are simple, precise, and sensitive, and they are applicable for the simultaneous determination of ASA and CLP in pure powder and formulations.     

Simultaneous determination of atorvastatin calcium and ramipril in capsule dosage forms by high-performance liquid chromatography and high-performance thin layer chromatography

Two simple and accurate methods to determine atorvastatin calcium and ramipril in capsule dosage forms were developed and validated using HPLC and HPTLC. The HPLC separation was achieved on a Phenomenex Luna C18 column (250 x 4.6 mm id, 5 microm) in the isocratic mode using 0.1% phosphoric acid-acetonitrile (38 + 62, v/v), pH 3.5 +/- 0.05, mobile phase at a flow rate of 1 ml/min. The retention times were 6.42 and 2.86 min for atorvastatin calcium and ramipril, respectively. Quantification was achieved with a photodiode array detector set at 210 nm over the concentration range of 0.5-5 microg/mL for each, with mean recoveries (at three concentration levels) of 100.06 +/- 0.49% and 99.95 +/- 0.63% RSD for atorvastatin calcium and ramipril, respectively. The HPTLC separation was achieved on silica gel 60 F254 HPTLC plates using methanol-benzene-glacial acetic acid (19.6 + 80.0 + 0.4, v/v/v) as the mobile phase. The Rf values were 0.40 and 0.20 for atorvastatin calcium and ramipril, respectively. Quantification was achieved with UV densitometry at 210 nm over the concentration range of 50-500 ng/spot for each, with mean recoveries (at three concentration levels) of 99.98 +/- 0.75% and 99.87 +/- 0.83% RSD for atorvastatin calcium and ramipril, respectively. Both methods were validated according to International Conference on Harmonization guidelines and found to be simple, specific, accurate, precise, and robust. The mean assay percentages for atorvastatin calcium and ramipril were 99.90 and 99.55% for HPLC and 99.91 and 99.47% for HPTLC, respectively. The methods were successfully applied for the determination of atorvastatin calcium and ramipril in capsule dosage forms without any interference from common excipients.     

Determination of rosuvastatin and ezetimibe in a combined tablet dosage form using high-performance column liquid chromatography and high-performance thin-layer chromatography

This paper describes validated HPLC and HPTLC methods for the simultaneous determination of rosuvastatin (ROS) and ezetimibe (EZE) in a combined tablet dosage form. The isocratic RP-HPLC analysis was performed on a Chromolith C18 column (100 x 6 mm id) using 0.1% (v/v) orthophosphoric acid solution (pH 3.5)-acetonitrile (63 + 37, v/v) mobile phase at a flow rate of 1 mL/min at ambient temperature. Quantification was carried out using a photodiode array UV detector at 245 nm over the concentration range of 0.5-10 microg/mL for ROS and EZE. The HPTLC separation was carried out on an aluminum-backed sheet of silica gel 60F(254) layers using n-butyl acetate-chloroform-glacial acetic acid (1 + 8 + 1, v/v/v) mobile phase. Quantification was achieved with UV densitometry at 245 nm over a concentration range of 0.1-0.9 micro/spot for ROS and EZE. The analytical methods were validated according to International Conference on Harmonization guidelines. Low RSD values indicated good precision. Both methods were successfully applied for the analysis of the drugs in laboratory-prepared mixtures and commercial tablets. No chromatographic interference from the tablet excipients was found. These methods are simple, precise, and sensitive, and are applicable for simultaneous determination of ROS and EZE in pure powder and tablets.     

Simultaneous estimation of pantoprazole and domperidone in pure powder and a pharmaceutical formulation by high-perfomance liquid chromatography and high-performance thin-layer chromatography methods

This paper describes validated high-performance liquid chromatography (HPLC) and high-performance thin-layer chromatography (HPTLC) methods for the simultaneous estimation of pantoprazole (PANT) and domperidone (DOM) in pure powder and capsule formulations. The HPLC separation was achieved on a Phenomenex C18 column (250 mm id, 4.6 mm, 5 pm) using 0.01 M, 6.5 pH ammonium acetate buffer-methanol-acetonitrile (30 + 40 + 30, v/v/v, pH 7.20) as the mobile phase at a flow rate of 1.0 mL/min at ambient temperature. The HPTLC separation was achieved on an aluminum-backed layer of silica gel 60F254 using ethyl acetate-methanol (60 + 40, v/v) as the mobile phase. Quantification was achieved with ultraviolet (UV) detection at 287 nm over the concentration range 400-4000 and 300-3000 ng/mL with mean recovery of 99.35+/-0.80 and 99.08+/-0.57% for PANT and DOM, respectively (HPLC method). Quantification was achieved with UV detection at 287 nm over the concentration range 80-240 and 60-180 ng/spot with mean recovery of 98.40+/-0.67 and 98.75+/-0.71% for PANT and DOM, respectively (HPTLC method). These methods are simple, precise, and sensitive, and they are applicable for the simultaneous determination of PANT and DOM in pure powder and capsule formulations.     

Simultaneous determination of losartan and hydrochlorothiazide in combined dosage forms by first-derivative spectroscopy and high-performance thin-layer chromatography.

Losartan (LST) is the first orally active nonpeptide angiotensin-II receptor antagonist with an improved safety and tolerability profile. It is prescribed alone or in combination with hydrochlorothiazide (HCTZ) for the treatment of moderate-to-severe hypertension. This paper describes the development of 2 methods that use different techniques, first-derivative spectroscopy and high-performance thin-layer chromatography (HPTLC), to determine LST and HCTZ in the presence of each other. LST and HCTZ in combined preparations were quantitated by using the first-derivative responses at 271.6 nm for LST and 335.0 nm for HCTZ in spectra of their solutions in water. The linearity ranges are 30-70 microg/mL for LST and 7.5-17.5 microg/mL for HCTZ with correlation coefficients of 0.9998 and 0.9997, respectively. In the HPTLC method, a mobile phase of chloroform-methanol-acetone-formic acid (7.5 + 1.5 + 0.5 + 0.03, v/v) and a prewashed Silica Gel G60 F254 TLC plate as the stationary phase were used to resolve LST and HCTZ in a mixture. Two well-separated and sharp peaks for LST and HCTZ were obtained at Rf values of 0.61+/-0.02 and 0.41+/-0.02, respectively. LST and HCTZ were quantitated at 254.0 nm. The linearity ranges obtained for the HPTLC method are 400-1200 and 100-300 ng/spot with corresponding correlation coefficients of 0.9944 and 0.9979, for LST and HCTZ, respectively. Both methods were validated, and the results were compared statistically. They were found to be accurate, specific, and reproducible. The methods were successfully applied to the estimation of LST and HCTZ in combined tablet formulations.     

Simultaneous assay of olanzapine and fluoxetine in tablets by column high-performance liquid chromatography and high-performance thin-layer chromatography

This paper describes validated high-performance liquid chromatographic (LC) and high-performance thin-layer chromatographic (TLC) methods for the simultaneous estimation of olanzapine and fluoxetine in pure powder and tablet formulations. The LC separation was achieved on a Lichrospher 100 RP-180, C18 column (250 mm, 4.0 mm id, 5 microm) using 0.05 M potassium dihydrogen phosphate buffer (pH 5.6 adjusted with o-phosphoric acid)-acetonitrile (50 + 50, v/v) as the mobile phase at a flow rate of 1 mL/min and ambient temperature. The TLC separation was achieved on aluminum sheets coated with silica gel 60F254 using methanol-toluene (40 + 20, v/v) as the mobile phase. Quantitation was achieved by measuring ultraviolet absorption at 233 nm over the concentration range of 10-70 and 40-280 microg/mL with mean recovery of 99.54 +/- 0.89 and 99.73 +/- 0.58% for olanzapine and fluoxetine, respectively, by the LC method. Quantitation was achieved by measuring ultraviolet absorption at 233 nm over the concentration range of 100-800 and 400-3200 ng/spot with mean recovery of 101.53 +/- 0.06 and 101.45 +/- 0.35% for olanzapine and fluoxetine, respectively, by the TLC method with densitometry. These methods are simple, precise, and sensitive, and they are applicable for simultaneous determination of olanzapine and fluoxetine in tablet formulations.     

Development of a simple and sensitive high-performance liquid chromatography method for determination of glucosamine in pharmaceutical formulations

A column high-performance liquid chromatography (HPLC) method was developed for the determination of glucosamine in dosage forms. Glucosamine was derivatized by addition of a solution containing orthophthaldialdehyde. The HPLC separation was achieved on a Spherimage 80 ODS2 column (250 x 4 mm id, 5 microm particle size) using an isocratic mobile phase containing phosphate buffer-methanol (90 + 10, v/v, pH 6.50) and methanol-tetrahydrofuran (97 + 3, v/v) in proportions of 85 + 15 at a flow rate of 1 mL/min, followed by fluorescence detection. The method was validated for specificity, linearity, accuracy, precision, limit of detection (LOD), and limit of quantitation (LOQ). The detector response for glucosamine HCI was linear over the concentration range of 0.1-20 microg/mL with a correlation coefficient of 0.9980. The accuracy was between 99.4 and 100.8%. The LOD and the LOQ were 0.009 and 0.027 microg/mL, respectively. The method was applied to determination of glucosamine in solid dosage forms.     

Modification and validation of a high-performance liquid chromatography method for quantification of Huperzine A in Huperzia crispata

The present study describes a rapid and sensitive HPLC method for the quantification of huperzine A (HupA) in Huperzia crispata (Huperziaceae). The sample extraction and preparation involved a simple, time-saving, single-solvent extraction, with each sample being analyzed within 12 min. The mobile phase was ammonium acetate (0.1 M, pH 6.0)--methanol (64 + 36, v/v) at a flow rate of 1.0 mL/min. Detection was at 308 nm. The calibration curve was linear from 0.049 to 7.84 microg (R2 = 0.9997), with intraday and interday precision RSD of less than 2%. The extraction recovery rate was over 98.49%. Quantification of HupA was performed using this modified method, and the content of HupA was 1.86 times higher in the whole plant of H. crispata (218.17 +/- 1.55 microg/g) than in that of H. serrata (117.03 +/- 2.97 microg/g). In the whole plant of H. crispata, HupA mainly accumulated in the actively growing shoot tips, the apical bud, and the 10 youngest leaves, reaching 455.23 +/- 2.97 microg/g. The content of HupA in the samples from sunshine-sheltered sites was 3.45 times higher than in that from sunshine-abundant sites. The satisfactory results indicate that this modified method can be applied in the quality control of large-scale Huperziaceae plant extracts and that changes should be made in the cultivation of H. crispata so as to maximize the production of HupA.     

A high-performance liquid chromatographic method for determination of mitragynine in serum and its application to a pharmacokinetic study in rats

A simple HPLC technique for determining mitragynine levels in serum was developed. The separation system consisted of a C18 column heated to 35 degrees C, a methanol-water (80:20, v/v) mobile phase, a flow rate of 0.8 mL/min and detection in the ultraviolet at 225 nm. Mitragynine, with a retention time of 10.09 min, was well resolved from any interferences in human serum and the internal standard peak. The calibration curve was linear from 0.1 to 10 microg/mL (r = 0.9995). Extraction of mitragy-nine from alkalinized serum using diethyl ether gave a high recovery (>or=85%). The intra- and inter-day precisions of the method were 4.29-5.88%RSD and 7.06-8.45%RSD, respectively. The accuracy ranged from -9.54 to +0.67%DEV. The limit of detection was 0.03 microg/mL and the lower limit of quantification was 0.1 microg/mL. Mitragynine in the stock solution was stable during 30 days of storage at 4 degrees C. This method was successfully applied to determine the pharmacokinetic characteristics of mitragynine levels in the serum of rats after it was administered orally.     

Simultaneous determination of caffeine, theobromine, and theophylline by high-performance liquid chromatography

This work relates the development of an analytical methodology to simultaneously determine three methylxanthines (caffeine, theobromine, and theophylline) in beverages and urine samples based on reversed-phase high-performance liquid chromatography. Separation is made with a Bondesil C18 column using methanol-water-acetic acid or ethanol-water-acetic acid (20:75:5, v/v/v) as the mobile phase at 0.7 mL/min. Identification is made by absorbance detection at 273 nm. Under optimized conditions, the detection limit of the HPLC method is 0.1 pg/mL for all three methylxanthines. This method is applied to urine and to 25 different beverage samples, which included coffee, tea, chocolate, and coconut water. The concentration ranges determined in the beverages and urine are: < 0.1 pg/mL to 350 microg/mL and 3.21 microg/mL to 71.2 microg/mL for caffeine; < 0.1 pg/mL to 32 microg mL and < 0.1 pg/mL to 13.2 microg/mL for theobromine; < 0.1 pg/mL to 47 microg/mL and < 0.1 pg/mL to 66.3 microg/mL for theophylline. The method proposed in this study is rapid and suitable for the simultaneous quantitation of methylxanthines in beverages and human urine samples and requires no extraction step or derivatization.     

Development and validation of reversed-phase column high-performance liquid chromatographic and first-derivative UV spectrophotometric methods for estimation of voriconazole in oral suspension powder

This research paper describes validated reversed-phase high-performance column liquid chromatographic (RP-HPLC) and first-derivative UV spectrophotometric methods for the estimation of voriconazole (VOR) in oral suspension powder. The RP-HPLC separation was achieved on Phenomenex C18 column (250 x 4.6 mm id, 5 microm particle size) using water-acetonitrile (40 + 60, v/v; pH adjusted to 4.5 +/- 0.02 with acetic acid) as the mobile phase at a flow rate of 1.4 mL/min and ambient temperature. Quantification was achieved with photodiode array detection at 255 nm over the concentration range of 0.1-1 microg/mL with mean recovery of 99.49 +/- 0.83% for VOR by the RP-HPLC method. Quantification was achieved with UV detection at 266 nm over the concentration range of 8-20 microg/mL with mean recovery of 99.74 +/- 0.664% for VOR by the first-derivative UV spectrophotometric method. These methods are simple, precise, and sensitive, and they are applicable for the determination of VOR in oral suspension powder.     

Simultaneous quantitation of rosuvastatin and gemfibrozil in human plasma by high-performance liquid chromatography and its application to a pharmacokinetic study

A simple, sensitive and specific high-performance liquid chromatography method is described for simultaneous determination of rosuvastatin (RST) and gemfibrozil (GFZ) in human plasma using celecoxib as an internal standard (IS). The assay procedure involved extraction of RST, GFZ and IS from plasma into acetonitrile. Following separation and evaporation of the organic layer the residue was reconstituted in the mobile phase and injected onto an X-Terra C(18) column (4.6 x 150 mm, 5.0 microm). The chromatographic run time was less than 20 min using flow gradient (0.0-1.60 mL/min) with a mobile phase consisting of 0.01 M ammonium acetate:acetonitrile:methanol (50:40:10, v/v/v) and UV detection at 275 nm. Nominal retention times of RST, GFZ and IS were 6.7, 13.9 and 16.4 min, respectively. Absolute recovery of both analytes and IS was greater than 90%. The lower limit of quantification (LLOQ) of RST and GFZ was 0.03 and 0.30 microg/mL, respectively. Linearity was excellent (r(2) = 0.999) in the 0.03-10 microg/mL and 0.3-100 microg/mL ranges for RST and GFZ, respectively. The inter- and intra-day precisions in the measurement of RST quality control (QC) samples 0.03, 0.09, 2.50 and 8.00 microg/mL were in the range 2.37-9.78% relative standard deviation (RSD) and 0.92-10.08% RSD, respectively. Similarly, the inter- and intra-day precisions in the measurement of GFZ quality control (QC) samples 0.30, 0.90, 25.0 and 80.0 microg/mL were in the ranges 2.79-6.27 and 0.96-9.69% RSD, respectively. Accuracies in the measurement of QC samples for RST and GFZ were in the range 85.43-107.23 and 84.98-102.35% respectively, of the nominal values. RST and GFZ were stable in the array of stability studies viz., bench-top, auto-sampler and freeze-thaw cycles. Stability of RST and GFZ was established for 1 month at -80C. The application of the assay in an oral pharmacokinetic study in rats co-administered with RST and GFZ is described.     

Simultaneous determination of albiflorin and paeoniflorin in rat urine by solid-phase extraction and high-performance liquid chromatography following oral administration of Si-Wu decoction

A high performance liquid chromatographic method (HPLC), together with solid phase extraction (SPE), was developed for simultaneous determination of albiflorin and paeoniflorin in rat urine after oral administration of Si-Wu decoction. The samples were pretreated with solid phase extraction using Extract-Cleantrade mark cartridges. Analysis of the extract was performed on a reversed-phase C18 column and a mobile phase made up of acetonitrile and 0.03% formic acid (17:83, v/v). UV detection was set at 230 nm. The assay was linear over the range 2.625-52.50 mg/mL for albiflorin and 3.875-77.50 microg/mL for paeoniflorin. The average percentage recoveries of three spiked urines were 97.01 +/- 3.32 and 102.32 +/- 6.97 for albiflorin and paeoniflorin, respectively. The intra-day precision (RSD) ranged from 0.21 to 1.79% at concentrations of 4.20, 10.50, 26.25 and 39.375 microg/mL of albiflorin and 0.12 to 2.92% at concentrations of 3.875, 10.85, 23.25 and 58.125 microg/mL of paeoniflorin, and inter-day precision (RSD) was from 1.02 to 1.86% for albiflorin and 0.94 to 3.30% for paeoniflorin, at the same four concentrations. This method was applied in order to analyze albiflorin and paeoniflorin in rat urine following oral administration of traditional Chinese medicinal preparation of Si-Wu decoction.     

A rapid and simple high-performance liquid chromatography method for the determination of human plasma levofloxacin concentration and its application to bioequivalence studies

A high-performance liquid chromatography method with fluorescence detection (HPLC-FLD) for the determination of levofloxacin in human plasma is described. Neutralized with phosphate buffer (pH 7.0), the sample (0.1 mL) was extracted with dichlormethane (1 mL). After voltex-mixing and centrifuged at 3000g for 6 min at 4 degrees C, the upper aqueous layer was aspirated using a micro vacuum pump and the organic layer was directly transferred to a clean test tube without pipetting. The organic solvent was evaporated and the residues were reconstituted with the mobile phase. Levofloxacin and terazosin (internal standard, IS) were chromatographically separated on a C(18) column with a mobile phase containing phosphate buffer (pH 3.0, 10 mm), acetonitrile and triethylamine (76:24:0.076, v/v/v) at a flow rate of 1 mL/min. The analytes were detected using fluorescence detection at an excitation and emission wavelength of 295 and 440 nm, respectively. The linear range of the calibration curves was 0.0521-5.213 microg/mL for levofloxacin with a lower limit of quantitation (0.0521 microg/mL). The retention times of levofloxacin and terazosin were 2.5 and 3.1 min, respectively. Within- and between-run precision was less than 12 and 11%, respectively. Accuracy ranged from -6.3 to 4.5%. The recovery ranged from 86 to 89% at the concentrations of 0.0521, 0.5213 and 5.213 microg/mL. The present HPLC-FLD method is sensitive, efficient and reliable. The method described herein has been successfully used for the pharmacokinetic and bioequivalence studies of a levofloxacin formulation product after oral administration to healthy Chinese volunteers.     

Determination of nifuroxazide and drotaverine hydrochloride in pharmaceutical preparations by three independent analytical methods

Three new, different, simple, sensitive, and accurate methods were developed for quantitative determination of nifuroxazide (I) and drotaverine hydrochloride (II) in a binary mixture. The first method was spectrophotometry, which allowed determination of I in the presence of II using a zero-order spectrum with an analytically useful maximum at 364.5 nm that obeyed Beer's law over a concentration range of 2-10 microg/mL with mean percentage recovery of 100.08 +/- 0.61. Determination of II in presence of I was obtained by second derivative spectrophotometry at 243.6 nm, which obeyed Beer's law over a concentration range of 2-10 microg/mL with mean recovery of 99.82 +/- 1.46%. The second method was spectrodensitometry, with which both drugs were separated on a silica gel plate using chloroform-acetone-methanol-glacial acetic acid (6 + 3 + 0.9 + 0.1) as the mobile phase and ultraviolet (UV) detection at 365 nm over a concentration range of 0.2-1 microg/band for both drugs, with mean recoveries of 99.99 +/- 0.15 and 100.00 +/- 0.34% for I and II, respectively. The third method was reversed-phase liquid chromatography using acetonitrile-water (40 + 60, v/v; adjusted to pH 2.55 with orthophosphoric acid) as the mobile phase and pentoxifylline as the internal standard at a flow rate of 1 mU/min with UV detection at 285 nm at ambient temperature over a concentration range of 2-10 microg/mL for both drugs, with mean recoveries of 100.24 +/- 1.51 and 100.08 +/- 0.78% for I and II, respectively. The proposed methods were checked using laboratory-prepared mixtures and were successfully applied for the analysis of pharmaceutical formulations containing the above drugs with no interference from other dosage form additives. The validity of the suggested procedures was further assessed by applying the standard addition technique which was found to be satisfactory, and the percentage recoveries obtained were in accordance with those given by the EVA Pharma reference spectrophotometric method.     

Estimation of trans-resveratrol in herbal extracts and dosage forms by high-performance thin-layer chromatography

A simple, sensitive and precise high-performance thin-layer chromatographic (HPTLC) method of analysis of trans-resveratrol in Polygonum cuspidatum root extracts and in dosage forms was developed and validated. The separation was carried out on a TLC aluminium plates precoated with silica gel 60F-254 as the stationary phase, eluted with chloroform-ethylacetate-formic acid (2.5 : 1 : 0.1) as mobile phase. Densitometric analysis of trans-resveratrol was carried out in the absorbance mode at 313 nm. This system was found to give compact spot for trans-resveratrol (Rf value of 0.40+/-0.03). A good linear regression relationship between peak areas and the concentrations was obtained over the range of 0.5-3.0 microg/spot with correlation coefficient 0.9989. The limit of detection and quantification was found to be 9 and 27 ng/spot. The method was validated for precision and recovery. The spike recoveries were within 99.85 to 100.70%. The RSD values of the precision in the range 0.37-1.84%. The proposed developed HPTLC method can be applied for identification and quantitative determination of trans-resveratrol in herbal extracts and dosage forms.     

Simultaneous determination of potassium clavulanate and cefixime in synthetic mixtures by high-performance liquid chromatography

A simple, precise, and sensitive high-performance liquid chromatographic method was developed and validated for the simultaneous determination of potassium clavulanate and cefixime in synthetic mixture form. The analytes were separated on a C18 column by using 0.03 M disodium hydrogen phosphate buffer (pH 6.5)-methanol (84 + 16, v/v) as the mobile phase with detection at 220 nm. The method exhibited high sensitivity and good linearity in the concentration ranges of 12.5-62.5 and 20-100 microg/mL for potassium clavulanate and cefixime, respectively. The total run time for the 2 components was <8 min, and the average recovery was >101.5% with a relative standard deviation of <1.0%. The proposed method was validated according to guidelines of the International Conference on Harmonization by evaluation of linearity, recovery, selectivity, robustness, limits of detection and quantitation, and within- and between-day precision. The results obtained for the synthetic mixture show that the method is highly precise and accurate for the simultaneous determination of potassium clavulanate and cefixime.     

Rapid determination of cinchocaine in skin by high-performance liquid chromatography

A simple, rapid, accurate, precise and specific analytical method has been developed, validated and applied for determination of cinchocaine in guinea pig and albino rabbit dorsal skins, after in vivo application of cinchocaine formulations. Extraction was performed using a solvent mixture of ethanol and 0.1 M hydrochloric acid (90:10; v/v). Samples were chromatographed on Spheri-5, RP(18) column with a particle size of 5 microm and 220 mm x 4.6 mm i.d. The mobile phase was a mixture of acetonitrile and triethylamine phosphate buffer (pH 2.8; 0.04 M) (60:40, v/v). UV detection was carried out at 247 nm and the run time was 6 min with typical retention time of cinchocaine of 3.63 +/- 0.02 min. Specificity was demonstrated, showing that the cinchocaine peak was free of interference from skin endogenous components. The detector response was found to be linear in the concentration range 0.96-56.00 microg/mL with a coefficient of correlation r = 0.99996. The relative standard deviations of within- and between-day analyses were all below 5%. The drug extraction procedure was validated. Satisfactory recoveries with relative standard deviation values below 5% were obtained, indicating efficient quantitative reproducible extraction procedure.     

Determination of the analgesic components of Spasmomigraine tablet by liquid chromatography with ultraviolet detection

A procedure was developed for the determination of the analgesic components of Spasmomigraine tablets, which are ergotamine (I), propyphenazone (II), caffeine (III), camylofin (IV), and mecloxamine (V). They were subjected to high-performance liquid chromatography on a column (300 x 3.9 mm, 10 rlm particle size) packed with micro-Bondapak C18. Separations were achieved with the mobile phase methanol-water-triethylamine (60 + 40 + 0.1, v/v/v) flowing at a rate of 1.5 mL/min, and quantitative determination was performed at 254 nm at ambient temperature for I-III; acetonitrile-25 mM KH2PO4-acetic acid (45 + 55 + 0.2, v/v/v), flowing at a rate of 1.5 mL/min and detection at 234 nm at ambient temperature, was used for IV and V. Methyl paraben was used as an internal standard. The detection limits were 0.35 (I), 5.0 (11), 1.5 (111), 3.0 (IV), and 2.0 microg/mL (V). The method was accurate (mean recovery 98+/-2%, n = 4) and precise (coefficient of variation <5%, n = 5). The proposed method is rapid and sensitive and, therefore, suitable for the routine control of these ingredients in multicomponent dosage forms.     

Validation of liquid chromatography and liquid chromatography/tandem mass spectrometry methods for the determination of etoricoxib in pharmaceutical formulations

Reversed-phase liquid chromatography (LC) and LC/tandem mass spectrometry (LC/MS/MS) methods were developed and validated for the determination of etoricoxib in pharmaceutical dosage forms. The LC method was performed by reversed-phase chromatography on a Synergi fusion C18 column (150 x 4.6 mm id) maintained at ambient temperature. The mobile phase consisted of 0.01 M phosphoric acid, pH 3.0-acetonitrile (62 + 38, v/v) at a flow rate of 1.0 mL/min, and photodiode array detection at 234 nm was used. The chromatographic separation was obtained within 7.0 min, and calibration curves were linear in the concentration range of 0.02-150 microg/mL. The LC/MS/MS method was performed on a Luna C18 column (50 x 3.0 mm id). The mobile phase consisted of acetonitrile-water (95 + 5)-0.1% acetic acid (90 + 10, v/v). Detection was performed by positive electrospray ionization in the multiple reaction monitoring mode, monitoring the transitions 359.3 > 280.0 and 332.0 > 95.0 for etoricoxib and piroxicam (internal standard), respectively. The chromatographic separation was obtained within 2.0 min, and calibration curves were linear in the concentration range of 1-5000 ng/mL. Validation parameters, such as specificity, linearity, precision, accuracy, and robustness, were evaluated, which gave results within the acceptable range for both methods. Moreover, the proposed methods were successfully applied for routine quality control analysis of pharmaceutical products and showed significant correlation (r = 0.9999) of the results.