RP-LC and HPTLC Methods for the Determination of Olmesartan Medoxomil and Hydrochlorothiazide in Combined Tablet Dosage Forms

Two new, rapid, precise, accurate and specific chromatographic methods were described for the simultaneous determination of olmesartan medoxomil and hydrochlorothiazide in combined tablet dosage forms. The first method was based on reversed phase liquid chromatography using an Eurosphere 100 RP C18 column (250 × 4.6 mm ID, 5 μm). The mobile phase was methanol–0.05% o-phosphoric acid (60:40 v/v) at a flow rate of 1.0 mL min^?1. Commercially available tablets and laboratory mixtures containing both drugs were assayed and detected using a UV detector at 270 nm. The second method involved silica gel 60 F₂₅₄ high performance thin layer chromatography and densitometric detection at 254 nm using acetonitrile–ethyl acetate–glacial acid (7:3:0.4 v/v/v) as the mobile phase. Calibration curves ranged between 200–600 and 125–375 ng spot^?1 for olmesartan and hydrochlorothiazide, respectively.     

Validated LC Method for Simultaneous Analysis of Tramadol Hydrochloride and Aceclofenac in a Commercial Tablet

This paper describes development and validation of a high-performance liquid chromatographic method for simultaneous analysis of tramadol hydrochloride (TR) and aceclofenac (AC) in a tablet formulation. When the combination formulation was subjected to ICH-recommended stress conditions, adequate separation of TR, AC, and the degradation products formed was achieved on a C₁₈ column with 65:35 (v/v) 0.01 M ammonium acetate buffer, pH 6.5—acetonitrile as mobile phase at a flow rate of 1 mL min^?1. UV detection was performed at 270 nm. The method was validated for specificity, linearity, LOD and LOQ, precision, accuracy, and robustness. The method was specific against placebo interference and also during forced degradation. The linearity of the method was investigated in the concentration ranges 15–60 μg mL^?1 (r = 0.9999) for TR and 40–160 μg mL^?1 (r = 0.9999) for AC. Accuracy was between 98.87 and 99.32% for TR and between 98.81 and 99.49% for AC. Because degradation products were well separated from the parent compounds, the method was stability-indicating.     

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

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

A Validated LC Method for the Quantitation of Cefotaxime in pH-Sensitive Nanoparticles

A sensitive and rapid routine LC method was validated for measuring cefotaxime incorporated in three different pH-sensitive nanoparticles. The drug was chromatographed on a C18 reversed-phase column; the mobile phase used was 0.05 M aqueous ammonium acetate, acetonitrile and tetrahydrofuran (87:11:2, v/v) adjusted to pH 5.5 with acetic acid. The flow rate was 1 mL min^?1 and cefotaxime was quantified at 254 nm, with a sensitivity range of 0.005 AUFS. The validated method was specific, linear (R ² ≥ 0.999), precise and accurate in a concentration range of 0.2–50.0 μg mL^?1. The method was rapid, selective and suitable for evaluation of cefotaxime in pH-sensitive Eudragit nanoparticles.     

LC Determination of Curcumin in Dog Plasma for a Pharmacokinetic Study

A rapid, simple, and sensitive high-performance liquid chromatographic method for quantification of curcumin in dog plasma has been developed and validated. After addition of the internal standard (berberine), plasma was acidified and extracted with ethyl acetate. Analysis was performed on a C₁₈ column. The mobile phase was acetonitrile–5% acetic acid, 52:48 (v/v) and the flow rate 1.0 mL min^?1. The eluent was monitored at 425 nm. Chromatographic separation was achieved in less than 7 min and the calibration plot was linear over the concentration range 2–128 ng mL^?1. Intra- and inter-assay variability were less than 7.3%. The accuracy ranged from 98.7 to 105.0%. The method was successfully applied to a pharmacokinetic study of curcumin in dogs.     

Simultaneous Determination of Ibuprofen and Diphenhydramine Citrate in Tablets by Validated LC

A stability-indicating LC method was developed for the simultaneous determination of ibuprofen and diphenhydramine citrate in pharmaceutical dosage forms. The chromatographic separation was achieved on an Inertsil ODS 3V, 150 × 4.6 mm, 5 μm, column. The mobile phase contained a mixture of 50 mM potassium dihydrogen phosphate buffer:acetonitrile:triethylamine:glacial acetic acid (55:45:0.2:0.2, v/v/v/v). This method allowed the determination of 2.85–9.14 mg mL^?1 of ibuprofen and 0.54–1.73 mg mL^?1 of diphenhydramine citrate, in a diluent consisting of pH 7.2, 50 mM potassium dihydrogen phosphate buffer:acetonitrile (40:60, v/v). The flow rate was 1.2 mL min^?1 and the detection wavelength was 260 nm. The limit of detection for ibuprofen and diphenhydramine citrate was 1.72 and 0.54 μg mL^?1 and the limit of quantification was 5.73 and 1.64 μg mL^?1, respectively. This method was validated for accuracy, precision and linearity. The method was also found to be stability indicating.     

A Weak Cation-Exchange Monolithic SPE Column for Extraction and Analysis of Caffeine and Theophylline in Human Urine

A weak cation-exchange monolithic column has been prepared in stainless steel tubing and used as the solid-phase extraction material in quantitative analysis of caffeine and theophylline in urine. Column switching, with water as mobile phase, was used for on-line cleaning and screening of human urine samples. Reversed-phase high-performance liquid chromatography was then performed on a C₁₈ column with methanol–water 30:70 (v/v) as mobile phase at a flow rate of 0.5 mL min^?1. Ultraviolet detection was performed at 274 nm. Good linear relationships were obtained between response and concentrations of caffeine and theophylline in the range 0.1–50 μg mL^?1. Absolute recovery ranged from 77.4 to 82.3% and inter-day and intra-day relative standard deviations were less than 5%. The method was suitable for analysis of caffeine and theophylline in human urine, because it eliminated tedious pretreatment and enabled rapid, economic, repeatable, and effective assay of traces of the drugs in biological samples.     

RP-LC and HPTLC Methods for the Determination of Olmesartan Medoxomil and Hydrochlorothiazide in Combined Tablet Dosage Forms

Two new, rapid, precise, accurate and specific chromatographic methods were described for the simultaneous determination of olmesartan medoxomil and hydrochlorothiazide in combined tablet dosage forms. The first method was based on reversed phase liquid chromatography using an Eurosphere 100 RP C18 column (250 × 4.6 mm ID, 5 μm). The mobile phase was methanol–0.05% o-phosphoric acid (60:40 v/v) at a flow rate of 1.0 mL min⁻¹. Commercially available tablets and laboratory mixtures containing both drugs were assayed and detected using a UV detector at 270 nm. The second method involved silica gel 60 F₂₅₄ high performance thin layer chromatography and densitometric detection at 254 nm using acetonitrile–ethyl acetate–glacial acid (7:3:0.4 v/v/v) as the mobile phase. Calibration curves ranged between 200–600 and 125–375 ng spot⁻¹ for olmesartan and hydrochlorothiazide, respectively.

     

Sensitive Quantification of Carboxylic Acid Metabolite of Clopidogrel in Human Plasma by LC with UV Detection

A rapid LC method with UV detection was developed for the quantification of carboxylic acid metabolite of clopidogrel in human plasma. Following a simple protein precipitation using a mixture of methanolic solution of ZnSO₄, the analyte and commercially available internal standard were separated using a mobile phase of water–acetonitril (85:15, v/v) adjusted to pH 3.5 on a Chromolith C18 column at a flow rate of 2.5 mL min^?1 with a total retention time of 4 min. Linearity was verified over the range of 20–3,000 ng mL^?1 where the LOQ was 20 ng mL^?1. This method was applied in a pharmacokinetic study.     

Extraction and Determination of β-Sitosterol from Salicornia herbacea L. Using Monolithic Cartridge

A short ionic liquids-based monolithic cartridge was prepared and used as the selective extraction sorbent. Characteristic and evaluation are investigated by field emission scanning electron microscopy (FE-SEM), and a new approach was developed for the extraction and determination of β-sitosterol from Salicornia herbacea L. using the ionic liquids-based monolithic cartridge. Chromatographic analysis was conducted on a C₁₈ column with UV detection at 210 nm, and an eluting solution consisting of acetonitrile–water (60/40, v/v) was used as the mobile phase at a flow rate of 0.8 mL min^?1. The linearity was confirmed in the concentration range of 0.50–100.00 μg mL^?1, with RSDs within 4.20%, and a recovery of β-sitosterol ranging from 97.20 to 102.93%. This method effectively removed the impurities without any tedious pretreatment, and it provided a fast, economic and effective way to assay trace drugs from natural plants.     

Development and Validation of an RP-HPLC Method for Determination of Valganciclovir in Human Serum and Tablets

This paper describes the validation of an isocratic high-performance liquid chromatographic method for the assay of valganciclovir in raw materials, tablets and human serum samples. Valganciclovir and fluvastatin (internal standard) were well separated using a reversed phase column and a mobile phase consisting of a mixture of acetonitrile:methanol:KH₂PO₄ (0.02 M) (40:20:40; v/v/v) (at pH 5.0). The mobile phase was pumped at 1.0 mL min⁻¹ flow rate and valganciclovir was detected by diode-array detection at 255 nm. The retention times for valganciclovir and fluvastatin were 3.41 and 5.60 min, respectively. A linear response (r > 0.999) was observed in the range of 10–30,000 ng mL⁻¹ in mobile phase and serum. The limit of detection and limit of quantification were found as 2.95 and 9.82 ng mL⁻¹ in mobile phase and 1.73 and 5.77 ng mL⁻¹ in human serum samples, respectively. Validation parameters as precision, accuracy, selectivity, reproducibility and system suitability tests were also determined. The method can be used for valganciclovir assay of tablets and human serum samples as the method separates valganciclovir from tablet excipients and endogenous substances.

     

Boronate-Containing Copolymer Grafted on Eupergit C as Matrix for Affinity Chromatography: Isotherms and Kinetics Study

A stability-indicating HPLC method has been developed and subsequently validated for the simultaneous determination of domperidone and pantoprazole in commercial tablets. The proposed HPLC method utilizes Phenomenex^® Gemini C₁₈ column (150 mm × 4.6 mm i.d., 5 μm) and mobile phase consisting of methanol-acetonitrile-20 mM dipotassium hydrogen phosphate and phosphoric acid buffer pH 7.0 (20:33:47, v/v/v) at a flow rate of 1.19 mL min^?1. Quantitation was achieved with UV detection at 285 nm based on peak area with linear calibration curves at concentration ranges 0.5–5.0 μg mL^?1 for domperidone and 1.0–10 μg mL^?1 for pantoprazole (R ² > 0.999 for both drugs). The method was validated in terms of accuracy, precision, linearity, limits of detection, limits of quantitation and robustness. This method has been successively applied to pharmaceutical formulation and no interference from the tablet excipients was found. Domperidone, pantoprazole and their combination drug product were exposed to acid, base and neutral hydrolysis, oxidation, dry heat and photolytic stress conditions and the stressed samples were analyzed by the proposed method. As the proposed method could effectively separate the drug from its degradation products, it can be employed as stability-indicating method for the determination of instability of these drugs in bulk and commercial products.     

SPE and LC–ESI–MS for Quantitative Analysis of Mitiglinide in Human Plasma in a Bioequivalence Study

Liquid chromatography–electrospray ionization mass spectrometry has been used for rapid, selective, and sensitive quantitative analysis of mitiglinide in human plasma. Sample pretreatment involved solid-phase extraction from plasma with gliclazide as internal standard. Separation was performed on a C₁₈ column (150 × 2.0 mm) with 71:29 (v/v) acetonitrile–water (containing 0.1% formic acid and 0.2 mmol L^?1 ammonium acetate) as mobile phase at a flow rate of 0.2 mL min^?1. The method was validated then successfully applied to a clinical bioequivalence study of mitiglinide in 20 healthy volunteers after oral administration.     

Chiral Separation of Tenatoprazole and Several Related Benzimidazoles by Normal Phase LC Using Amylose-Based Stationary Phase

Simple and efficient analytical LC methods using amylose-based stationary phase Chiralpak AS-H were developed for direct enantioseparation of tenatoprazole and several related benzimidazoles. The chromatographic experiments were performed in the normal phase mode with n-hexane–ethanol–triethylamine (TEA) as mobile phase. The effects of the mobile phase additive, concentration of organic modifiers and column temperature were studied for the enantioseparation. The thermodynamic parameters were also calculated from the van’t Hoff plots. It was found that the enantioseparations were all enthalpy driven. The enantiomers of all compounds were resolved (R _s  > 3.3) within 14 min using n-hexane–ethanol–TEA (20:80:0.1%, v/v/v) as mobile phase with a flow rate of 0.4 mL min^?1 at 40 °C. The optimized method was validated for determination of the enantiomers of tenatoprazole in terms of linearity, precision and accuracy according to ICH guidelines and applied to the assay of tenantoprazole bulk drugs. The proposed method was shown to be accurate and suitable for the quantitative determination of tenatoprazole enantiomers.     

Development and Validation of a Reversed-Phase HPLC Method for the Determination of Deflazacort in Pharmaceutical Dosage Forms

A reversed-phase high-performance liquid chromatographic method has been developed and validated for quantitative determination of deflazacort in tablets and in compounded capsules. Isocratic chromatography was performed on a C₁₈ column with acetonitrile–water 80:20 (v/v) as mobile phase at a flow rate of 1.0 mL min^?1. UV detection was at 240 nm. The linearity of the method was good (r > 0.999), as also were intra-day and inter-day precision (RSD <2%) and accuracy (recovery >98%). The method was also validated for specificity and robustness. The results showed the proposed method is suitable for its intended use.     

Simultaneous Determination of Dextromethorphan Hydrobromide, Pyrilamine Maleate and Sodium Benzoate in a Cough Cold Syrup by LC

A liquid chromatographic method for the simultaneous determination of dextromethorphan hydrobromide, pyrilamine maleate and sodium benzoate in cough cold syrup has been developed. The method was based on replacing heptane sulfonate by sodium chloride as ion pairing agent. The addition of sodium chloride to the mobile phase has changed the retention behaviour of the basic drugs. The separation of these compounds was achieved in less than 8 min with an isocratic mobile phase consisting of acetonitrile/0.1 M dihydrogenphosphate buffer containing 0.1 M sodium chloride (29:71 v/v) at pH 2.5 and using a Kromasil C18 column. The analysis was performed at a flow rate of 1 mL min^?1 and at a detection wavelength of 220 nm. The selectivity, linearity of calibration, accuracy, within and between days precision, limit of detection and quantification, recovery were examined as parts of the method validation. Calibration curves were linear in the range 1–140 μg mL^?1 with a regression coefficient (R ²) better than 0.999. The results of the method repeatability (intra-day) and reproducibility (inter-day) were all less than 2% (n = 6). The lowest detectable concentration of dextromethorphan hydrobromide and pyrilamine maleate varied between 0.10 and 0.12 μg mL^?1. The proposed liquid chromatographic method was satisfactorily applied for the routine quality control of dextromethorphan hydrobromide, pyrilamine maleate and sodium benzoate in cough cold syrup formulations.     

A Simple Method for Determination of Homologue Imidazolium Cations in Ionic Liquids Using IC with Direct Conductivity Detection

An investigation was carried out into the fast determination of five homologue imidazolium cations in ionic liquids by ion chromatography using a cation-exchange column and direct conductivity detection. Ethylenediamine, complex organic acid (citric acid, oxalic acid and tartaric acid) and organic modifiers (acetonitrile) were used as mobile phase. The influences of the eluent types, eluent concentration, eluent pH and column temperature on separation of the cations were discussed. Simultaneous separation and determination of the five homologue imidazolium cations in ionic liquids were achieved under an optimum condition. The optimized mobile phase was consisted of 0.25 mmol L^?1 ethylenediamine + 0.5 mmol L^?1 citric acid + 3% acetonitrile (v/v) (pH 4.1), set at a flow rate of 1.0 mL min^?1. The column temperature was 40 °C and detection limits were obtained in the range of 1.1–45.6 mg L^?1. The relative standard deviations of the chromatographic peak areas for the cations were <3.0% (n = 5). This method was successfully applied to separate imidazolium cations in ionic liquids produced by organic synthesis. The recoveries of spiked components were 92.5–101.9%.     

Quantitative Analysis of Busulfan in Human Plasma by LC-MS–MS

High-performance liquid chromatography with tandem mass spectrometry has been used for rapid, specific, and sensitive analysis of busulfan in human plasma. Busulfan-d₈ was used as internal standard. Analysis was performed on a C₁₈ column (50 mm × 2.1 mm, 3.5-µm particles) with water–methanol 80:20 (v/v) as mobile phase at a flow-rate of 0.30 mL min^?1. Detection was by tandem triple–quadrupole mass spectrometry with turbo ion-spray ionization. Linear calibration plots were obtained over the concentration range 1.096–1,096 ng mL^?1. The assay is ideally suited to monitoring of busulfan and determination of its pharmacokinetic data.     

Estimation of Perospirone in Human Plasma by LC–MS–MS and Its Application to Pharmacokinetics Study

A sensitive liquid chromatography-tandem-mass spectrometry method was developed and validated for the determination of perospirone in human plasma, using quetiapine as internal standard. Plasma samples were extracted from 1 mL of plasma using n-hexane. Chromatographic separation was performed on an Agilent Zorbax SB C₁₈ column with a mobile phase of 5 mM ammonium acetate solution-methanol (12:88, v/v, adjusted to pH 3.8 with glacial acetic acid) at a flow rate of 0.2 mL min^?1. The chromatographic separation was achieved in less than 4.6 min. The linearity was established over the concentration range of 0.05–20 ng mL^?1. Both of the intra- and inter-batch standard deviation was less than 9.8%. The method was successfully applied to study the pharmacokinetic parameters of perospirone hydrochloride tablets in healthy Chinese volunteers.     

Simultaneous Determination of Clopidogrel and Its Carboxylic Acid Metabolite (SR26334) in Human Plasma by LC–ESI–MS–MS: Application to the Therapeutic Drug Monitoring of Clopidogrel

A sensitive and specific liquid chromatography-tandem-mass spectrometry method was developed and validated for the simultaneous determination of clopidogrel and its carboxylic acid metabolite (SR26334) in human plasma using nateglinide and pioglitazone as internal standards. Analytes were extracted from 0.50 mL of plasma using diethyl ether–n-hexane (4:1, v/v). Chromatographic separation was performed on a Teknokroma C₁₈ column with a mobile phase of methanol–water (containing 0.1% formic acid) (80:20, v/v) at a flow rate of 0.20 mL min^?1 within 5.6 min. Linearity was established over the concentration range of 0.005–5 ng mL^?1 for clopidogrel and 20–2,500 ng mL^?1 for SR26334. Intra- and inter-batch standard deviations were less than 9.2% and the accuracy of this assay was found to fall within an acceptable range ≤10.0%. The method was successfully applied to the therapeutic drug monitoring of clopidogrel.