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Abstract: New antidiabetic drug combination Metformin Hydrochloride, Dapagliflozin, and Saxagliptin have been recently approved for type II diabetes. This is marketed by Takeda Pharmaceuticals under the brand name Qternmet XR. Although different analytical and bioanalytical methods using different techniques such as liquid chromatography, high-performance liquid chromatography, high-performance thin-layer chromatography, gas chromatography, spectrophotometry, spectrofluorimetric methods coupled with ultraviolet, fluorescence, mass, or tandem-mass spectrometry detection have already been developed for the determination of Metformin Hydrochloride, Dapagliflozin, and Saxagliptin. Sensitive, cost-effective, and more optimized methods are yet required. Therefore, this review summarizes the main analytical and bioanalytical aspects regarding not only simultaneous estimation but also stability-indicating methods, kinetic studies, and impurity analysis for the analysis of proposed drugs in bulk and pharmaceutical dosage forms. Thus, this review gathers, for the first time, important background information on all analytical and bioanalytical methods that have been developed and applied for the determination of Metformin Hydrochloride, Dapagliflozin, and Saxagliptin, which should be considered as a starting point if new techniques are aimed to be implemented for these drugs.

Aim: The present study developed an oral in-situ gel containing the leaf extract of Tarax-acum officinale (T.O.) using an experimental design. Background: Peptic ulcer disease was an epidemic in the 19th and early 20th centuries. The applica-tion of herbal drugs for peptic diseases is an attractive area for research and implementation as compared to the allopathic system in the recent era. From the number of plants, the antioxidant effect of an aqueous extract of Taraxacum officinale, which is the dandelion leaf, was proven by an animal study in previous literature. However, most of the marketed preparations consist of root ex-tract primarily used for the detoxification of the liver in comparison to the leaf extract having a non-specific application. Hence, the aqueous extract of dandelion leaf was taken as the active ingredient in the formulation. Over the past 30 years, greater attention has been given to the development of controlled and sustained drug delivery systems. The development of in situ gel systems has received considerable attention over the past few years due to the number of advantages. Objective: By considering the merits of herbal ingredients with drug delivery technology, herbal in-situ gel was developed using statistical analysis. Methods: Herbal in-situ oral gel was developed using a factorial design. The concentration of sodi-um alginate, xanthan gum, and HPMC K15 M was taken as the factors, and viscosity and drug re-lease (total phenol content) in 10 h were selected as the responses. By evaluating the designed batches, the optimized batch was chosen as the promising formulation for curing the peptic ulcer. Results: The batch consisting of 1.711% w/v sodium alginate, 0.727% w/v xanthan gum, and 0.869% w/v HPMC K15M was selected as the optimized batch, as per the software, and the viscosi-ty and % drug release in 10 h were found to be 299.5 cps and 70.2%, respectively. Other evaluation parameters such as gelling capacity, floating parameters, and stability were also found to be good for the designed batches. Conclutions: The improved in-situ gel was found to be effective for extending the floating of the drug incorporated into the formulation as well as the residence time in the stomach for sustained drug release.

Variable and low oral bioavailability (4-11%) of lumefantrine (LUF), an anti-malarial agent, is characterized by very low solubility in aqueous vehicle. Thus, the present study was intended to formulate lyophilized nanosuspensions of LUF to resolve its solubility issues for the improvement of oral bioavailability. A three level 3

Epigenetic therapeutics are the new generation of chemotherapeutics for treatment of cancer, and histone deacetylase inhibitors have been actively discovered in this category They target the biological processes including the cell cycle, apoptosis, DNA repair, cell cycle control, autophagy, metabolism, senescence and chaperone function. Several families of histone deacetylase (HDAC) inhibitors have been synthesized and evaluated. Their positive effects on the cell cycle have been demonstrated in biological models and in clinical trials. Recently Food and Drug Administration has approved Vorinostat, Romidepsin and Belinostat for oncologic indications of refractory cutaneous and peripheral T cell Lymphoma. These advances have provided the motivation to develop more potent and selective inhibitors and target other pathologic conditions with these drugs. Major ongoing efforts are to develop inhibition as monotherapy, rational combination with chemotherapy and other targeted drugs. Some progress is made into developing isoform specific drugs. In this perspective, the biological functions and potential substrates of histone deacetylase enzymes are reviewed and the characteristics of this inhibitors are discussed in respect with anticancer activity and further therapeutic interest. Keywords: Epigenetic therapeutics, Histone deacetylase inhibitors, Anticancer, Classification, Food and drug administration approved drugs.

Objective: Role of leptin resistance in correlation between obesity and asthma. Methods: High-caloric diet was given for 8 weeks to induce obesity. Ovalbumin followed by aluminum hydroxide was given to induce asthma. The animal was treated with leptin analog (0.4 mg/kg, i.p. for 7 days) and leptin antagonist (3 mg/kg, p.o., for 7 days). Biochemical parameters such as serum leptin, ghrelin, and tumor necrosis factor alpha (TNF-α) and physical parameters such as tidal volume and airflow rate were estimated to confirm the state of asthma and obesity, respectively. Results: It was found that leptin and ghrelin were elevated in obese and obese asthmatic condition, responsible for leptin resistance. Treatment with leptin analog and leptin antagonist significantly increases and decreases serum leptin levels, respectively. There was no significant change in TNF-α and ghrelin level after leptin analog treatment. The result of respiratory parameters improved with leptin analog. From our study, we found beneficial role of leptin analog in obese asthmatic condition. Conclusion: Leptin is an alternative treatment approach to treat obese asthmatic condition.

The objective of the present study was to prepare a matrix-type transdermal drug delivery system containing metoprolol succinate and to evaluate the effect of essential oils on transdermal permeation of drug. The solvent evaporation method was employed to fabricate transdermal patches of drug using HPMC K15M and Acrycoat L100 as polymers and polyethylene glycol 400 as a plasticizer. Prepared patches were subjected for evaluation of physical as well as physicochemical properties, moisture characteristics, and ex-vivo drug permeation studies. The results of physical characterization showed uniform casting and good appearance of patches. Satisfactory mechanical strength was revealed in the results of tensile strength and folding endurance. Ex-vivo permeation study indicated enhancing effect of essential oils on drug permeation through the skin. The maximum flux observed was 94 μg/cm2.hr achieved by formulation M3 containing lemongrass oil (20%). The results suggested that lemongrass oil was effective at higher concentration (20%) after 14 hrs of permeation while peppermint oil and eucalyptus oil were effective at low concentration (10%). Release kinetic suggested diffusion-controlled release of drug which followed the Korsmeyer-Peppas model. In conclusion, essential oils can be utilized as a safe and effective alternative for the permeation enhancement of drugs through transdermal delivery.

An isotope dilution selective and sensitive high-performance liquid chromatography-tandem mass spectrometry (LC-ESI-MS/MS) method has been developed for the simultaneous determination of hydrochlorothiazide (HCTZ) and ramipril in human plasma through a new concept of periodical polarity switching. Extraction of HCTZ, ramipril and their deuterated analogs as internal standards (ISs) was carried out from 150 μL of human plasma by solid-phase extraction method. Chromatographic separation of analytes was performed on Hypurity C18 (150 mm × 4.6 mm, 5 μ) column under gradient conditions with methanol:0.2% (v/v) formic acid in water as the mobile phase. The method was validated over a concentration range of 0.750-300 ng/mL for HCTZ and 0.125-80.0 ng/mL for ramipril. The mean extraction recovery for analytes and ISs were >(86.0%), consistent across all four QC levels. The challenges to evaluate matrix effect and continuous reproducibility of method during long analytical run was studied and resolved. Processed samples, freeze-thaw, long-term and whole blood stability were evaluated for both the analytes. The method was applied to support a bioequivalence study of 25 mg of HCTZ and 5 mg of ramipril tablet formulation in nine healthy Indian subjects. Assay reproducibility was demonstrated by reanalysis of 42 incurred samples.

In the current scenario, green technologies in chemistry and various fields create attention to every researcher. The green synthesis provides environmentally and eco-friendly raw materials and intermediate products, making a simple process without using harmful and toxic reactants. Amino acids are one of the reactants from which the intermediates or excipients can be made using the straightforward methodology, and these are utilized in drug delivery via a different route of Administration. By employing such medication at a cellular level, we can eliminate the adverse events and toxicity of chemically synthesized excipients and achieve a better therapeutic effect. In the present paper, Arginine was reviewed as the starting material from which the greener excipients can be synthesized, and these materials were employed for drug delivery systems. However, some of the synthetic processes employed chemicals, and also usage of the synthesized green material in the formulation of different dosage forms may require proper guidelines, but still, the research is going on for the application of green excipients for therapeutic purposes. Moreover, Arginine, as such, is an amino acid. It can also be utilized in different formulations as an active ingredient, which was also emphasized in the review paper from past research and patents. This review aims to be comprehensive and have general interest for chemists as it involves the various routes by which excipients were synthesized and the study, including the properties of different excipients and their application.

A specific, rapid, sensitive and selective ultra-performance liquid chromatography - tandem mass spectrometry has been developed for the simultaneous determination of midodrine and desglymidodrine in human plasma. The analytes and its deuterated analogs were quantitatively extracted from 100μL of human plasma by solid phase extraction technique. Separation of analytes was achieved on the Waters Acquity UPLC BEH C18 (50×2.1mm, 1.7μm) column using acetonitrile-4.0mM ammonium formate, pH 2.5(90:10, v/v) as mobile phase. The protonated analytes were quantified by selected reaction monitoring in the positive ionization mode by triple quadrupole mass spectrometer. The calibration plots were linear over the concentration range of 0.050-50.0ng/mL. The intra-batch and inter-batch precision (%CV) across quality control levels was4.0 and the% mean relative recovery was ≥96%. Various other parameters like stability in different conditions; matrix effect and reproducibility of the method were performed in accordance with the guidelines specified by the USFDA for bioanalytical method development and validation. The developed method was successfully administered to the pharmacokinetics study of 5 mg midodrine tablet in 12 healthy subjects. Reproducibility of assay was proved by reanalysis of 48 incurred samples.

The application of l -arginine, the nonessential amino acid, as in pharmaceutical and cosmetics is raised as it shows the number of advantages for internal and external, applications. But its hydrophilic nature and lower partition co-efficient value limit its penetration through the skin. From the different approaches for topical drug delivery systems, solid lipid nanoparticles are a promising vehicle to carry the drug molecule at the appropriate site of absorption and also ensures the fast therapeutic action. With ability to incorporate lipophilic as well as a hydrophilic molecule. The present investigation aimed to implement the Quality by design concept for the development of SLNs incorporated with arginine molecule for its application on skin in the form of cosmoceutical formulation. The developed formulation will enhance the blood circulation of skin cells and hence beneficial for skin as cosmoceutical dosage form. To enhance the loading of drug molecule in a lipolphilic environment like solid lipid, insitu ion-pairing was carried out using an optimum concentration of oleic acid. The optimization was done by QbD approach considering concentration of poloxamer 188, oleic acid and sonication amplitude as the critical processing parameters. Average particle size and % drug loading were taken as critical product quality attributes using 23 full factorial design. The characterization of freeze dried solid lipid nanoparticles are investigated by transmission electron microscopy, x-ray diffraction study, differential scanning calorimetry and fourier transform infrared spectroscopy. Finally, an in-vitro and ex-vivo drug diffusion stuies were carried out to check the permeation of arginine. The optimized SLN formulation were successfully developed having lower particle size and higher % drug loading with effective skin penetration parameters to achieve the therapeutic benefits of Arginine.

The stability of silver nanoparticles is controlled mainly by two major factors, namely, aggregation and oxidation. In the present study, silver nanoparticles were synthesized by using different series of reducing agents like a strong reducing agent (sodium borohydride), a mild reducing agent (tri-sodium citrate), and a weak reducing agent (glucose) with different capping agents, namely, polyvinyl pyrrolidone (PVP K 30), starch, and sodium carboxyl methyl cellulose (NaCMC). The synthesized silver nanoparticles were characterized by UV-Visible absorption spectroscopy, dynamic light scattering (DLS), atomic force microscopy (AFM), and anti-microbial activity. The particle size of silver nanoparticles varies in the following order: sodium borohydride

Background: Balofloxacin is a third generation fluoroquinolone with a broad antibacterial spectrum ranging from gram-positive bacteria to gram-negative bacteria. It is used in treatment of uncomplicated urinary tract infections. No stability indicating analytical method has been reported for BFX. Also stress degradation studies of Balofloxacin were not found in literature. Objective: To develop and validate a stability indicating RP-HPLC method for estimation of Balofloxacin in presence of its hydrolytic degra­dation products. Materials and Method: The chromatographic separation was performed using C 18 , Grace Smart column (250 x 4.6 mm), 5 μm as the stationary phase and Water: Acetonitrile: Tri ethylamine (72:28:1 v/v/v), pH adjusted to 3.0 using ortho-phosphoric acid as mobile phase with detec­tion wavelength 294 nm. The developed method was validated according to ICH Q2R1 guideline. Balofloxacin was subjected to degradation in acidic, alkaline and neutral conditions. Results and Discussion: The linearity was established over concentration range of 20-100 μg/ml with correlation coefficient r 2 = 0.9979. Recovery of drug was achieved in the range of 99.19–101.65%. Limit of Detection and Limit of Quantitation was found to be 4.13 and 12.51 μg/ml. Balofloxacin was found to be stable under alkaline and neutral conditions, while it degraded under acidic hydrolytic condition. The retention times for Balofloxacin and its acid degradation product were found to be 7.0 ± 0.1 and 5.7 ± 0.1 minutes, respectively. Application: The developed RP-HPLC method was applied for estimation of Balofloxacin in its tablet dosage forms and results were found to be in good agreement with the labeled claim. The method was also applied for degradation kinetic study of Balofloxacin in acidic medium. Conclusion: The developed RP-HPLC method was found to be accurate, precise, specific and sensitive. It can be applied for routine analysis (assay) of tablets containing Balofloxa­cin. The degradation of Balofloxacin in all conditions was found to be first order and highest degradation was found in 2.0 N HCl at 75°C. Key words: Stability indicating HPLC method, Balofloxacin (BFX), Degradation products, Degradation kinetic study, Design expert software–9.

Background: Dipeptidyl Peptidase 4 (DPP 4) enzyme cleaves an incretin-based glucoregulatory hormone Glucagon Like Peptide -1 from N-terminal where penultimate amino acid is either alanine or proline. Several DPP 4 inhibitors, “gliptins”, are approved for the management of Type 2 Diabetes or are under clinical trial. In the present study, combined pharmacophore and docking-based virtual screening protocol were used for the identification of new hits from the Specs Database, which would inhibit DPP 4. Methods: The entire computational studies were performed using the Discovery Studio v. 4.1 software package, Pipeline Pilot v. 9.2 (Accelrys Inc.) and FRED v. 2.2.5 (OpenEye Scientific Software). Common feature pharmacophore model was generated from known DPP 4 inhibitors and validated by Receiver Operating curve analysis and GH-scoring method. Database search of Specs commercial database was performed using validated pharmacophore. Hits obtained from pharmacophore search were further docked into the binding site of DPP 4. Based on the analysis of docked poses of hits, 10 compounds were selected for in- vitro DPP 4 enzyme inhibition assay. Results: Based on docking studies, virtual hits were predicted to form interaction with essential amino acid residues of DPP 4 and have an almost similar binding orientation as that of the reference molecule. Three compounds having Specs database ID- AN-465/42837213, AP-064/42049348 and AN- 465/43369427 were found to inhibit DPP 4 enzyme moderately. Conclusion: The present study demonstrates a successful utilization of in-silico tools in the identification of new DPP 4 inhibitor, which can serve as a starting point for the development of novel DPP 4 inhibitors.

Microemulsions (ME) have been proved to increase the cutaneous absorption of both lipophilic and hydrophilic medicaments when compared to conventional vehicles (emulsions, pure oils, aqueous solutions). Hence the aim of present investigation is to prepare, characterize and optimize microemulsion of Itraconazole (ITZ). Itraconazole is an anti fungal agent, most widely used in the ringworm infection. It is classified as class III drug as per BCS classification. It indicates lower permeability through skin. Therefore objective of the research is to improve permeability of Itraconazole through skin. Microemulsion was prepared using eucalyptus oil, tween 20 and methanol as oil phase, surfactant and co-surfactant respectively. Pseudo ternary phase diagrams were constructed to find out optimum ratio of oil: Smix (surfactant: Co-Surfactant): water. A 32 full factorial design was applied for the optimization of prepared microemulsion. Microemulsion was evaluated for globule size, zeta potential, in-vitro diffusion study etc. Results of globule size measurements and zeta potential indicated ME7 had high stability then other formulation of microemulsion. For the optimization transdermal flux and %Q6 was selected as dependant variables. Results of optimization study also revealed ME 7 as optimized microemulsion for high permeability to the skin. Further ME7 was compared to marketed Itraconazole preparation (ITASPORE) and evaluated using similarity factor F2. Results of F2 value was not near to 100 indicated there is no similarity in diffusion profiles of ME7 and ITASPORE. Hence, indirectly it suggest there was increased in permeability of drug by preparing microemulsion.

A HPTLC method for the simultaneous estimation of aliskiren, amlodipine, and hydrochlorothiazide in the combination dosage form is proposed. Silica gel 60 F254 precoated plates were used as the stationary phase, whereas Quality by design approach was applied for the efficient separation of the drugs through which a simple mobile phase composition Ethyl acetate:Methanol:Ammonia (7.5:2.8:0.22, v/v/v) was finalized. The RF values optimized were 0.43 ± 0.01, 0.29 ± 0.01, and 0.65 ± 0.01 for aliskiren, amlodipine, and hydrochlorothiazide, respectively. The densitometric analysis was carried out at 229 nm. The method was linear in the range of 1500–5250 ng spot−1 for aliskiren, 50–175 ng spot−1 for amlodipine, and 125–432.5 ng spot−1 for hydrochlorothiazide. The method was validated as per the International Conference on Harmonization guidelines and the RSD (%) was within the acceptable criterion indicating the method is accurate and precise. The LOD and LOQ were found to be 133.38 ng spot−1 and 404.19 ng spot−...

The objective of the present work was to carry out systematic evaluation to eliminate matrix effect owing to plasma phospholipids as observed during sample preparation and to develop a cross-talk-free sensitive, selective and rapid bioanalytical method for the simultaneous determination of zolmitriptan (ZT) and N-desmethyl zolmitriptan (DZT) in human plasma by liquid chromatography–tandem mass spectrometry using naratriptan as internal standard (IS). The analytes and IS were quantitatively extracted from 200 μL human plasma by solid phase extraction. No cross-talk was found between ZT and DZT having identical product ions. Quantitation was performed on a triple quadrupole mass spectrometer employing electrospray ionization technique, operating in multiple reaction monitoring and positive ion mode. The total chromatographic run time was 2.5 min. The method was fully validated for sensitivity, selectivity, specificity, linearity, accuracy, precision, recovery, matrix effect, dilution integrity and stability studies. The method was validated over a dynamic concentration range of 0.1–15 ng/mL for ZT and DZT. The method was successfully applied to a bioequivalence study of 2.5 mg ZT tablet formulation in 18 healthy Indian male subjects under fasting conditions. Assay reproducibility was assessed by reanalysis of 62 incurred samples. Copyright © 2015 John Wiley & Sons, Ltd.

Background: The presence of potentially active nutrients and their multifunctional properties make prickly pear a perfect candidate for the production of phytopharmaceutical products. Among the numerous Opuntia species, bioactive compounds have been isolated and characterized primarily from Opuntia ficus-indica, Opuntia polycantha, Opuntia stricta, Opuntia dilleni for various medicinal properties. Objective: Based on the traditional use of prickly pear for enhancement of immune function, the objective of the present study to evaluate the effect of prickly pear on mast cell degranulation function. Materials and Methods: The Opuntia fruit juice (OFJ) (10-200 μl/ml) were studied for the effect on sensitized rat peritoneal mast cell degranulation induced by immunological (egg albumin), and nonimmunological (compound 48/80) stimuli and compared with that of the reference standard, sodium cromoglycate and ketotifen (10 μg/ml). Results and Conclusion: The OFJ exhibited significantly (P < 0.001) concentration dependent inhibition of mast cell degranulation. The IC 50 value of OFJ was found 12.24 and 18 μl/ml for immunological and nonimmunological induced mast cell degranulation, respectively. The betacyanin is an active principle compound in prickly pear that may responsible for mast cell stabilizing action.

Background: Opuntia elatio Mill is a xerophytic plant with potentially active nutrients. It is traditionally appreciated for its pharmacological properties; however, the scientific information on this plant is insufficient. Objective: The present study evaluates the antinociceptive and anti-inflammatory action of prickly pear. Materials and Methods: Writhing and tail-immersion tests were carried out to evaluate analgesic action, while the carrageenan-induced paw edema and neutrophil adhesion tests were conducted in Albino wistar rats to assess anti-inflammatory action. Results: ED 50 values of the fruit juice in writhing, tail immersion, and paw edema test were 0.919, 2.77, and 9.282 ml/kg, respectively. The fruits of Opuntia produced analgesic and anti-inflammatory action in a dose-dependent manner. Conclusion: The results establish the folklore use of prickly pear may be due to the presence of betacyanin and/or other phenolic compounds.

A sensitive and specific ultra-performance liquid chromatography tandem mass spectrometric (UPLC–MS/MS) method has been developed, validated and applied for the assay of Nebivolol and S-amlodipine in human plasma. Sample extraction was carried out through hybrid extraction method from 250 μL of human plasma sample. Linearity of the method was (r ≥ 0.9996) was found to be dynamic for both the analytes over concentration range of 25.0–4000 pg/mL. Chromatographic separation was achieved on UPLC column {Waters Acquity UPLC BEH C18 (50 mm × 2.1 mm, 1.7 micrometer)} with the mobile phase composition of 0.1% (v/v) formic acid in 5 mM Ammonium formate in water-acetonitrile (20:80, %v/v). Analytes Stability was assured under different requisite conditions in human plasma, reconstitution solution and diluents. Inter and intra-day assay precision and relative error (accuracy) were within ±5% for both analytes. The method was applied and reproduced to support a pharmacokinetic study of 5 mg Nebivolol (NEB) and 2.5 mg S-amlodipine (LAM) tablet on 9 healthy subjects.

A simple, sensitive, precise, and stability-indicating high-performance thin-layer chromatographic method has been developed and validated for the quantitative determination of pitavastatin calcium (PTV) in a pharmaceutical dosage form in the presence of its degradation products. The stability of PTV was investigated under different stress conditions, including hydrolytic, oxidative, photolytic, and thermal, as recommended by the International Conference on Harmonization guidelines. PTV was separated on aluminum-backed silica gel 60 F254 high-performance thin-layer chromatography (HPTLC) plates as a stationary phase with ethyl acetate–methanol–toluene–glacial acetic acid (4:1:5:0.1, v/v/v/v) as a mobile phase. Regression analysis data for the calibration plots were indicative of good linear relationships between responses and concentration over the range 25–150 ng per band. The method was validated for linearity, precision, accuracy, robustness, specificity, and sensitivity. The drug was subjected to stre...

Two simple and accurate methods to determine pitavastatin calcium (PIT) and ezetimibe (EZE), ina binary mixture, were developed and validated using LC and densitometric-TLC. The LC separation was achieved on a Phenomenex Luna C18 column (250 × 4.6 mm id, 5 μm), in the isocratic mode using 0.1% orthophosphoric acid–acetonitrile–triethylamine (19.8 + 80 + 0.2, v/v/v), pH 3.00 ± 0.05, at a flow rate of 1.4 mL/min. The retention times were about 6.98 and 2.36 min for PIT and EZE, respectively. Quantification was achieved with a photodiode array detector at 235 nm over the concentration range of 0.5–5 μg/mL for each drug, with meanrecoveries of 99.35 ± 0.19% and 99.51 ± 0.23% for PIT and EZE, respectively. The TLCseparation was achieved on silica gel 60 F254 HPTLC plates using toluene–ethyl acetate–glacial acetic acid (59.8 + 40.0 + 0.2, v/v/v), as the mobile phase. The Rf values were about 0.38 and 0.58 for PIT and EZE, respectively. Quantification was achieved withUV detection at 235 nm over the concentration range of 50–600 ng/spot for each drug, with mean recoveries of 99.49 ± 0.15% and 99.58 ± 0.10% for PIT and EZE, respectively. Both methods were validated, and based on statistical results were found to be specific, accurate, precise, and robust. The methods were successfully applied for the determination of PIT and EZE in abinary mixture without any interferences from common excipients.

A novel, precise, sensitive and accurate ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method has been developed for the simultaneous determination of a novel drug combination, candesartan (CAN) and chlorthalidone (CHL), in human plasma. Chromatographic separation was achieved on Waters Acquity UPLC BEH C18 (50 × 2.1 mm, 1.7 μm). Mobile phase consisting of 1 mm ammonium acetate in water-acetonitrile (20:80 v/v) was used. The total chromatographic runtime was 1.9 min with retention times for CAN and CHL at 0.7 and 1.1 min respectively. Ionization and detection of analytes and internal standards was performed on a triple quadrupole mass spectrometer, operating in the multiple reaction monitoring and negative ionization mode. Quantitation was done to monitor protonated precursor → product ion transition of m/z 439.2 → 309.0 for CAN, 337.0 → 189.8 for CHL and 443.2 → 312.1 for candesartan D4 and 341.0 → 189.8 for chlorthalidone D4. The method was validated over a wide dynamic concentration range of 2.0-540.0 ng/mL for candesartan and 1.0-180.0 ng/mL for chlorthalidone. The validated method was successfully applied for the assay of CAN and CHL in healthy volunteers.

Introduction: The fruits of Opuntia elatior Mill. are known as prickly pear and folkloric use as hematinic, anti-inflammatory and antiasthmatic action. Previously, the fruit juice of prickly pear was evaluated in reversed anemia induced by HgCl2 in a dose dependant manner and present study revealed about its effect in acute hemolytic anemia. Aim: To evaluate the hematinic activity of fruits of Opuntia elatior Mill. Materials and Methods: The hematinic activity of an orally administered fruit juice was studied on phenylhydrazine (PHZ)-induced anemic rats. The hematological parameters such as hemoglobin (Hb) content, red blood cell (RBC), packed cell volume (PCV), and reticulocyte count were analyzed as indices of anemia. Results: PHZ altered the hematological parameters by hemolysis characterized by a decrease in Hb content, total RBC counts and PCV (P < 0.001) on day 3. The Hb content (g%) was significantly increased (P < 0.05) at day 7 in 10 and 15 ml/kg fruit juice treated rats, which was a good improvement compared to the standard. Conclusion: The speedy and progressive recovery of anemic rats responding to treatment of the O. elatior Mill. fruits may be due to increased erythropoiesis and/or antioxidant property of betacyanin.

Betalains are of great taxonomic significance in higher plants and occur only in 10 families of the order Caryophyllales (Centrospermae). They are water-soluble nitrogenous pigments. They can be divided into two major structural groups, the red to red-violet betacyanins and the yellow betaxanthins. Betalains are widely used as natural red food colorant as well as antioxidant potentials. Several methods have been published for the determination of betalain in fruits of Opuntia species. The purpose of the current review is to provide a systematic survey of the analytical techniques for the determination of betalain from fruits of Opuntia species.

Hyperlipidemia is considered one of the key factors for cardiovascular diseases. Based on earlier work on a series of 5-alkyl-4-aryl-3-mercapto-(4H)-1,2,4-triazoles, for further lead modification, a series of 4-(substituted)amino-5-substituted-3-mercapto-(4H)-1,2,4-triazoles was designed. Target compounds were synthesized by the well known Hoggarth synthesis of substituted 1,2,4-triazoles. Synthesized compounds were screened for lipid lowering activity using the "Poloxamer 407 induced hyperlipidemia in rats" model at a dose of 100 mg/kg p.o. Compounds were found to alter serum lipid levels significantly. Most of the compounds significantly reduced serum cholesterol and triglyceride levels. Some of the compounds were found to reduce triglycerides and elevate high density lipoprotein (HDL) levels more than the standard drug atorvastatin (CAS 134523-03-8). Compounds with chloro substitution on aryl rings were found more active in reducing serum lipid levels than other substitutions.

A quantitative high-performance thin-layer chromatography (HPTLC) method for determination of darunavir ethanolate (DRV) in tablets has been established and validated. DRV from the formulations was separated and identified on silica gel 60 F254 HPTLC plates with toluene-ethyl acetate-methanol 7.0:2.0:1.0 (ν/ν) as mobile phase. The plates were developed to a distance of 8 cm. Quantification was performed at λ = 267 nm. Well-resolved bands were obtained for DRV. The method was validated for specificity, precision, robustness, and accuracy. The calibration plot for DRV standard was linear in the range 250–1750 ng per band with r = 0.9994, slope = 0.4253, and intercept = 44.81. The limits of detection and quantification were 15.28 and 45.84 ng per band, respectively. The method is selective, sensitive, and specific, with potential application in pharmaceutical analysis.

Objective: A simple, precise and accurate isocratic reversed phase (RP) column high-performance liquid chromatographic (HPLC) method has been developed for simultaneous analysis of eprosartan (EPR) and hydrochlorothiazide (HCT) in tablet formulations. Materials and Methods: Isocratic RP-HPLC separation was achieved on phenomenex C18 column (250 x 4.6 mm i.d., 5 μm particle size) using mobile phase composed of 0.5% formic acid-methanol-acetonitrile [(80 : 25 : 20 v/v/v) pH, 2.80 ± 0.04] at a flow rate of 1.0 ml/min. The retention time for EPR and HCT was 7.69 ± 0.10 and 4.24 ± 0.09 minutes, respectively. The detection was performed at 272 nm. Results: The method was linear in the concentration range of 60-600 μg/ml for EPR and 2.5-25 μg/ml for HCT with a correlation coefficient of 0.9992 and 0.9997, respectively. The repeatability for six samples was 0.53 and 0.61 % RSD for EPR and HCT, respectively. The accuracy (recovery) was found to be in the range of 99.46 to 100.61% for EPR and 99.06 to 100.93% for HCT, respectively. Conclusions: The method was validated and successfully used for determination of the drugs in tablets. Key words: Eprosartan, high-performance liquid chromatographic, hydrochlorothiazide, tablets.

Cholesteryl ester transfer protein (CETP), an enzyme which catalyses the transfer of cholesteryl ester from HDL to VLDL, is a promising target for discovery of novel antihyperlipidemic agents due to its pivotal role in HDL metabolism and reverse cholesterol transport. Quantitative structure activity relationship study of a series of CETP inhibitors was carried out using genetic function approximation to study various structural requirements for CETP inhibition. Various lipophilic, electronic, geometric and spatial descriptors were correlated with CETP inhibitory activity. Developed models were found predictive as indicated by their goodr2predvalues and satisfactory internal and external cross-validation results. Study reveals that lipophilicity (ClogP), with parabolic relationship, contributed significantly to the activity along with some electronic, geometric and quantum mechanical descriptors. The present study can be applied to future lead optimization of CETP inhibitors.

A simple, precise, and accurate isocratic RP-HPLC method was developed and validated for determination of eprosartan in bulk drug and tablets. Isocratic RP-HPLC separation was achieved on a Phenomenex C18 column (250 4.6 mm id, 5 m particle size) using the mobile phase 0.5 formic acidmethanolacetonitrile (80 25 20, v/v/v, pH 2.80) at a flow rate of 1.0 mL/min. The retention time of eprosartan was 7.64 0.05 min. The detection was performed at 232 nm. The method was validated for linearity, precision, accuracy, robustness, solution stability, and specificity. The method was linear in the concentration range of 10400 g/mL with a correlation coefficient of 0.9999. The repeatability for six samples was 0.253 RSD; the intraday and interday precision were 0.210.57 and 0.330.71 RSD, respectively. The accuracy (recovery) was found to be in the range of 99.86100.92. The drug was subjected to the stress conditions hydrolysis, oxidation, photolysis, and heat. Degradation products produced as a result of the stress conditions did not interfere with detection of eprosartan; therefore, the proposed method can be considered stability-indicating.

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 ⨯ 4.6 mm id, 5 µm) in the isocratic mode using 0.1% phosphoric acidacetonitrile (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 µg/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 methanolbenzeneglacial 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 50500 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.

A rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS-MS) method has been developed and validated for the determination of lamotrigine in human plasma using multiplexing technique (two HPLC units connected to one MS-MS). Lamotrigine was extracted from human plasma by solid-phase extraction technique using Oasis Hydrophilic Lipophilic Balance (HLB) or N-vinylpyrrolidone and divinylbenzene cartridge. A structural analog, 3,5-diamino-6-phenyl-1,2,4-triazine, was used as an internal standard (IS). A BetaBasic C(8) column was used for the chromatographic separation of analytes. The mass transition [M+H](+) ions used for detection were m/z 256.0 --> 211.0 for lamotrigine and m/z 188.0 --> 143.0 for IS. The method involved a simple multiplexing, rapid solid-phase extraction without evaporation and reconstitution. The proposed method has been validated for a linear range of 0.025 to 10.000 microg/mL with a correlation coefficient > or = 0.9991. The limit of quantification for lamotrigine was 0.025 microg/mL, and limit of detection was 50.000 pg/mL. The intra-run and inter-run precision and accuracy were within 10.0% for intra-HPLC runs and inter-HPLC runs. The overall recoveries for lamotrigine and IS were 97.9% and 92.5%, respectively. Total MS run time was 1.4 min per sample. The validated method has been successfully used to analyze human plasma samples for applications in pharmacokinetic, bioavailability, bioequivalence, or in vitro in vivo correlation studies.

A simple, sensitive, accurate, and precise high-performance thinlayer chromatographic method has been established for simultaneous analysis of atorvastatin calcium, ramipril, and aspirin in a capsule dosage form. The compounds were separated on silica gel with methanol-benzene-ethyl acetate-glacial acetic acid 0.36:5.6:4.0: 0.04 (v/v) as mobile phase. Ultra-violet detection was performed at 210 nm. The RF values were approximately 0.38, 0.06, and 0.86 for atorvastatin calcium, ramipril, and aspirin, respectively. The method was validated for linearity, accuracy, precision, limits of detection and quantification, and robustness. The linearity ranges were 100–600 ng per band for atorvastatin calcium, 50–300 ng per band for ramipril, and 500–3000 ng per band for aspirin. Mean recoveries were 99.97 ± 0.07%, 100.01 ± 0.10%, and 100.02 ± 0.06 % for atorvastatin calcium, ramipril, and aspirin, respectively. Limits of detection (LOD) were 4.88 ng for atorvastatin calcium, 2.91 ng for ramipril, and 18.63 ng for aspirin. Salicylic acid (RF ~ 0.72) was found as an impurity in the capsule dosage form. The proposed method can be used for analysis of these drugs in combined dosage forms.

Summary A new, simple, accurate, and precise high-performance thin-layer chromatographic (HPTLC) method has been established for simultaneous analysis of eprosartan and hydrochlorothiazide in tablet formulations. Standard and sample solutions of eprosartan and hydrochlorothiazide were applied to precoated silica gel G 60 F254 HPTLC plates and the plates were developed with benzene-methanol-formic acid 7:3:0.1 (υ/υ) as mobile phase. Detection and evaluation of chromatograms was performed densitometrically at 272 nm. The retention factors of eprosartan and hydrochlorothiazide were 0.76 and 0.57, respectively. The linear range was 4.8–43.2 μg per spot for eprosartan and 0.15–1.35 μg per spot for hydrochlorothiazide; the correlation coefficients, r, were 0.998 and 0.999, respectively. The method was validated and successfully used for analysis of the drugs in tablets.

Two simple and accurate methods for the determination of pitavastatin calcium (PIT) in tablet dosage forms were developed and validated using column liquid chromatography (LC) and UV spectrophotometry. The LC separation was achieved on a Phenomenex Luna C18 column (250 mm, 4.6 mm id, 5 m) in the isocratic mode using acetonitrilewatertriethylamine (80 + 19.8 + 0.2, v/v/v), adjusted to pH 3.5 0.05 with orthophosphoric acid, as the mobile phase at a flow rate of 1.5 mL/min. The retention time was 5.70 min. Both the methods were performed at 238 nm, the wavelength of maximum absorbance of PIT in methanol. In the LC method, quantification was achieved with a photodiode array detector over the concentration range of 0.12.5 g/mL with a mean recovery of 100.26 0.75. In the UV method, quantification was achieved over the concentration range of 220 g/mL with mean recovery of 99.65 1.24. Both methods were validated, and the results were compared statistically. They were found to be simple, specific, accurate, and precise. The methods were successfully applied for the determination of PIT in tablet dosage form without any interference from common excipients.

A simple, sensitive, precise and accurate reversed phase liquid chromatographic method has been developed for the simultaneous estimation of atorvastatin (AT) calcium, ramipril (RA) and aspirin (AS) from capsule dosage form. The method was developed using a Phenomenex Luna C18 (250 mm, 4.6 mm i.d., 5 µm) column with a mobile phase consisting of 0.1%, orthophosphoric acid buffer:acetonitrile:methanol (45:50:5 v/v/v), pH 3.3, at a flow rate of 1 mL min−1. Detection was carried out with ultra-violet detection at 210 nm. The retention times were about 12.19, 2.35, and 3.95 min for AT calcium, RA and AS, respectively. The developed method was validated for linearity, accuracy, precision, limit of detection, limit of quantitation and robustness. The linearity ranges were 1–6 µg mL−1 for AT calcium, 0.5–3 µg mL−1 for RA and 7.5–45 µg mL−1 for AS with mean recoveries of 100.59 ± 0.68, 100.62 ± 0.83 and 100.49 ± 0.73% for AT calcium, RA and AS, respectively. Limit of detection obtained were 29.85 ng mL−1 for AT calcium, 4.71 ng mL−1 for RA and 85.13 ng mL−1 for AS. Impurity of salicylic acid was found in capsule dosage form at the retention time of about 4.84 min. The proposed method can be used for the estimation of these drugs in combined dosage forms.

A quantitative high-performance thin-layer chromatographic method for determination of pitavastatin calcium in pharmaceutical preparations has been established and validated. Pitavastatin calcium from the formulations was separated and identified on silica gel 60F 254 HPTLC plates with toluene-methanol-glacial acetic acid 7.6:2.36:0.04 ( v/v ) as mobile phase. The plates were developed to a distance of 8 cm and well resolved bands were obtained for pitavastatin calcium. Quantification was performed at 238 nm. The method was validated for specificity, precision, robustness, and accuracy. The calibration plot for pitavastatin calcium standard was linear in the range 40–250 ng per band with r = 0.9992, slope = 8.2415, and intercept = 203.2359. The limits of detection and quantification were 6.6 and 20.0 ng per band, respectively. The method is selective, sensitive, and specific with potential application in pharmaceutical analysis.

This paper describes a validated high-performance thin-layer chromatography (HPTLC) method for simultaneous estimation of rabeprazole (RA) and domperidone (DO) in pure powder and in capsule formulations. An HPTLC method separation is achieved on an aluminum sheet of silica gel 60F(254) using ethyl acetate-methanol-benzene-acetonitrile (30:20:30:20 v/v) as mobile phase. Quantitation is achieved with UV detection at 287 nm over a concentration range of 400-1200 ng/spot and 600-1800 ng/spot with mean recovery of 99.82 +/- 0.74 and 99.43 +/- 0.68 for RA and DO, respectively, in the HPTLC method. This method is simple, precise, and sensitive, and it is applicable for the simultaneous determination of RA and DO in pure powder and in capsule formulation.

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 m) 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 methanoltoluene (40 + 20, v/v) as the mobile phase. Quantitation was achieved by measuring ultraviolet absorption at 233 nm over the concentration range of 1070 and 40280 g/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 100800 and 4003200 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.

A sensitive, selective, precise, and stability‐indicating high performance thin layer chromatography (HPTLC) method for analysis of satranidazole both as a bulk drug and in formulations was developed and validated. The method employed TLC aluminium plates precoated with silica gel 60F‐254 as the stationary phase. The solvent system consisted of toluene/acetonitrile (60:40, v/v). Densitometric analysis of satranidazole was carried out in the absorbance mode at 314 nm. This system was found to give compact spots for satranidazole (R f value of 0.53±0.02, for six replicates). Satranidazole was subjected to acid and alkaline hydrolysis, oxidation, and photo degradation. The drug undergoes degradation under acidic and basic conditions, oxidation, and photo degradation. Also, the degraded products were well resolved from the pure drug with significantly different R f values. The method was validated for linearity, precision, limit of detection (LOD), limit of quantitation (LOQ), and accuracy. Linearity...

A rapid, simple, and sensitive HPLC and a densitometric HPTLC method for the determination of omeprazole and domperidone in capsule formulations were developed and validated. For HPLC, the separation of components was achieved on a Phenomenex Rp‐C18 column. Isocratic elution with a mobile phase consisting of 0.01 M pH 6.5, ammonium acetate buffer: methanol:acetonitrile (40:30:30 v/v, pH 7.44±0.02), at a flow rate 1.0 mL/min was employed. Rabeprazole was used as the internal standard. In densitometric HPTLC, separation was achieved on aluminum sheets of silica gel 60F254 using ethyl acetate:methanol:benzene (40:20:40 v/v) as mobile phase. Linear concentration range of HPLC and HPTLC methods were between 400–2000 ng/mL and 120–360 ng/spot for both the drugs, respectively. In HPLC, the detection limit was 131.27 ng/mL for omeprazole and 131.20 ng/mL for domperidone, the mean analytical recovery in determination of omeprazole and domperidone capsules was 99.14±1.81 for omeprazole and 99.63±1.68 for d...

A simple, sensitive, and precise high performance liquid chromatographic method for the analysis of pantoprazole, rabeprazole, esomeprazole, domperidone and itopride, with ultraviolet detection at 210 nm, has been developed, validated, and used for the determination of compounds in commercial pharmaceutical products. The compounds were well separated on a Hypersil BDS C18 reversed-phase column by use of a mobile phase consisting of 0.05 M, 4.70 pH, potassium dihydrogen phosphate buffer - acetonitrile (720:280 v/v) at a flow rate of 1.0 mL min−1. The linearity ranges were 400–4,000 ng mL−1 for pantoprazole, 200–2,000 ng mL−1 for rabeprazole, 400–4,000 ng mL−1 for esomeprazole, 300–3,000 ng mL−1 for domperidone and 500–5,000 ng mL−1 for itopride. Limits of detection (LOD) obtained were: pantoprazole 147.51 ng mL−1, rabeprazole 65.65 ng mL−1, esomeprazole 131.27 ng mL−1, domperidone 98.33 ng mL−1 and itopride 162.35 ng mL−1. The study showed that reversed-phase liquid chromatography is sensitive and selective for the determination of pantoprazole, rabeprazole, esomeprazole, domperidone and itopride using single mobile phase.

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 μm) 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 acetatemethanol (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.

Objective: The present study was aimed to identify the physicochemical data of shodhit guggul. Guggul is a gum-resin exudate from the plant Commiphora weightii (Arn.) Bhandari , belonging to Burseraceae family. In Ayurveda, guggul is always purified. This purification is known as Shodhan. Shodhan is a process by which guggul is made non-toxic, effective, suitable and fit for therapeutic purposes. Methods: The seven different shodhan dravya were used to prepare shodhit guggul. They were evaluated by performing physicochemical parameters including five different extractive value; total ash, acid insoluble, water soluble and sulphated ash value; pH, and loss on drying. Results: Analytical results of raw guggul showed total ash, acid insoluble ash, water soluble ash and sulphated ash value to 5.36±0.04%, 0.96±0.03%, 4.51±0.03 % and 8.40±0.04% respectively. These all values of each shodhit guggul were different. The extractive value of raw guggul was comparable with standard value while the extractive value of each shodhit guggul was totally different. The pH value of 1% w/v and 10% w/v aqueous solution of raw guggul was 6.44±0.18 though pH of each shodhit guggul was changed. The loss on drying of raw guggul was found to be 1.88±0.02%w/v, however, this value was different for each shodhit guggul. Conclusion: The present study revealed that the different shodhan process with specific shodhan dravya affects the physicochemical parameters. The analysis and comparison of the data showed the difference in the properties of seven shodhit guggul with respect to raw Guggul.

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 μg/mL for LST and 7.5–17.5 μg/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.