Mixture Of Phosphate Buffer Research Articles

A STABILITY INDICATING ION-PAIR LC METHOD FOR THE DETERMINATION OF ASENAPINE IN PHARMACEUTICALS

In this study, a new, simple and specific stability indicating ion-pair LC method was developed and fully validated for the determination of asenapine in tablets. The analysis was performed on an Agilent Eclipse XDB-C8 column (4.6 x 150 mm, 3.5 μm particles) at 30˚C. A mixture of phosphate buffer (pH 3, 20 mM) containing 10 mM 1-heptane sulfonic acid and acetonitrile, (60:40, v/v) at a flow rate of 1 mL min-1 was used as mobile phase. Detection was performed by a diode array detector at 220 nm. The developed method was validated according to related ICH guideline and US Pharmacopeia and it was suitable in terms of accuracy, precision, specificity, robustness and stability. The method was linear in the concentration range of 0.5-100 μg mL-1. Limit of detection and limit of quantification values were calculated as 0.0836 μg mL-1 and 0.2788 μg mL-1, respectively. This ion-pair LC method was applied successfully for the determination of asenapine in its sublingual tablets.

Wpływ bazy kosmetycznej na uwalnianie kwasu usninowego

Usnic acid is a type of lichen acid which exhibits a broad spectrum of biological activity, functioning as an antimicrobial, an anti-inflammatory and a UV-filter. It is used as an active ingredient in herbal medicines and cosmetics products. Its influence as a cosmetics base, including through properties such as emulsion type or degree of internal phase droplet size, on the release of usnic acid, was studied. Two types (O/W, W/O) of usnic acid – loaded, stable emulsions with varying degrees of dispersion of the dispersed phase were obtained. The concentration of usnic acid in all of the formulations studied was 0.05% w/w. The formulations’ physicochemical properties, including stability, viscosity, pH and the droplet size of the dispersed phase, were evaluated. The kinesis of the usnic acid’s release was performed in a thermostatic diffusion chamber, at a temperature of T = 32°C, using dialysis membranes of regenerated cellulose. A mixture of phosphate buffer (PBS) and ethanol (EtOH), in 90/10, PBS/EtOH, volume ratio, was used as an acceptor solution. The concentration of released usnic acid was analyzed by UV-Vis spectrophotometer. The results showed that the O/W emulsions, as compared to W/O emulsions, are a much more effective means of administering usnic acid to the skin. After 24 hours, approx. 45% of the initial acid concentration penetrated into the acceptor solution from O/W emulsion, while from the W/O emulsion the released amount was about 18%.

Enzymatic synthesis of 5-amino quinoline oligomers and evaluation of their free radical scavenging activity

The compound 5-aminoquinoline (AQ) was treated with horseradish peroxidase (HRP) and H2O2 in a reaction medium containing a dioxane:phosphate buffer mixture (70:30). Products (OAQ) with a degree of polymerization ranging from 4 to 12 were obtained. 1H NMR, FTIR, UV–vis, SEM, TG–DTA, DSC, XRD analysis, and conductivity measurements were conducted for characterization of OAQ. Compared with a monomer, OAQ showed higher thermal stability. After heating to 1000 °C, 46% of the initial weight of the OAQ remained. The antioxidant efficiencies of the monomer and oligomer were evaluated by the typical spectrophotometric method such as the bleaching of 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals. OAQ showed improved DPPH activity compared with the AQ monomer.

Simultaneous Determination of Ursodeoxycholic Acid and Chenodeoxycholic Acid in Pharmaceutical Dosage Form by HPLC-UV Detection.

A reversed-phase HPLC method was developed for the simultaneous determination of ursodeoxycholic acid (UDCA) and the epimeric isomer, chenodeoxycholic acid (CDCA), in their synthetic mixtures and in tablet dosage form. The proposed HPLC method uses a C18 column and mobile phase consisting of an acetonitrile-phosphate buffer mixture (pH 2.3, 100 mM; 50 + 50, v/v) at a flow rate of 2.0 mL/min with UV detection at 210 nm. The method was validated according to the International Conference on Harmonization guidelines; and linearity, range, accuracy, precision, robustness, and system suitability were studied. The LOD and LOQ were also calculated and found to be 1.23 and 3.73 μg/mL for UDCA and 0.83 and 2.52 μg/mL for CDCA, respectively. The method was adapted for UHPLC, in which baseline separation was achieved in <2.5 min. The assay results of Ursomix tablets by the developed method were statistically compared with those obtained by the reference method using t- and F-tests, and no significant differences were observed.

Determination of phenolic acids and rutin in Heliotropium thermophilum by high-performance liquid chromatography with photodiode array detection

ABSTRACTA novel and rapid method was validated for the determination of phenolics in plants by high-performance liquid chromatography coupled to photodiode array detection. The optimized method was employed for the quantification of phenolic acids and rutin present in Heliotropium thermophilum. Eight samples collected from two locations were analyzed. Additionally, temperature dependent changes in the phenolic composition of H. thermophilum samples was evaluated. A series of standards were used for identification that included gallic acid, protocatechuic acid, 4-hydroxybenzoic acid, chlorogenic acid, vanillic acid, caffeic acid, syringic acid, p-coumaric acid, rutin, ferulic acid, benzoic acid, 2-hydroxycinnamic acid, abscisic acid, and trans-cinnamic acid. Chromatographic separation was performed on C8 column at 40°C with gradient elution. A phosphate buffer (20 mM, pH: 2.5) and acetonitrile mixture was used as the mobile phase with detection at 280 nm wavelength and a flow rate of 1.5 mL/min. The developed method was shown to be sensitive, accurate, and reproducible for the determination of phenolic acids and rutin in plants.

RP-HPLC-PDAAnalyses of Tapentadol: Application of Experimental Design

A simple, rapid and sensitive RP-HPLC-PDA method was developed and validated as per ICH guidelines for the analysis of tapentadol in bulk as well as in tablet formulation. The RP-HPLC isocratic method was developed using a column LC-GC Qualisil BDS C-18 (250 mm × 4.6 mm × 5 μm) and methanol - potassium dihydrogen phosphate buffer (pH 7.0) mixture (80:20, v/v) as a mobile phase. The flow rate was retained at 1 ml min-1 and the response was detected at 274 nm. Robustness of the developed method was evaluated using two experimental designs (central composite design and response surface methodology). The effect of factors such as mobile phase composition, flow rate and pH of mobile phase were studied.

Comparative Study of the Resolution Efficiency of High-performance Liquid Chromatographic and Chemometrics-assisted UV Spectrophotometric Methods: Application on Pharmaceutical Mixtures

High-performance liquid chromatographic and two chemometrics-assisted UV spectrophotometric methods were proposed for the resolution of pharmaceutical binary mixtures of lidocaine hydrochloride(LH) and fluocortolone pivalate, (FCP) and their ternary mixture with chlorquinaldol (CQ) without any chemical pretreatment. The chromatographic separation was achieved on normal-phase C8 analytical column with a mobile phase consisting of a mixture of phosphate buffer : acetonitrile (40:60, v/v) buffered with 0.5% triethylamine, (adjusted to pH 3.3 ± 0.2 with o- phosphoric acid) with flow rate 1 mL/min. Quantitation was achieved with UV detection at 220 nm. Also, the resolution has been accomplished by using numerical spectrophotometric methods as principal component regression (PCR) and partial least squares (PLS-1) applied to the UV spectra of the mixture. The proposed methods were validated and successfully applied for the analysis of dosage forms and laboratory-prepared mixtures containing the components combination. The developed analytical methods did not show any interference of the excipients when applied to pharmaceutical dosage forms. The results of HPLC were compared with PLS-1 and PCR methods and a good agreement was found.

Determination of Degradation Products of Cyclobenzaprine Hydrochloride, Lidocaine and Piroxicam in a Semi-Topical Formulation: MS-MS Confirmation of Unknown Impurities.

Association of cyclobenzaprine hydrochloride, piroxicam and lidocaine in a topical formulation is one of the newest innovations in the pharmaceutical formulary field. In this study, a reversed-phase liquid chromatographic method was developed for the establishment of the impurities of cyclobenzaprine hydrochloride, lidocaine and piroxicam in the semisolid topical formulation. In this study, we not only determined 2,6-dimethylaniline, 2-pyrydilamine but also specified impurities of cyclobenzaprine hydrochloride (dibenzosuberenone, amitriptyline, carbinole, cyclobenzaprine N-oxide and anthrachinone). The target compounds were determined using a mobile phase that consisted of a mixture of phosphate buffer (0.025 M; pH 6.2)-acetonitrile-methanol (60 : 13 : 27, v/v/v). A minimum of three supplementary possible degradation products were determined. Using mass spectrometry, the unspecified impurities were identified and the use of correlation matrices permitted the association with the possible source compounds. The chromatographic conditions were qualified and validated according to ICH guideline requirements to confirm specificity, linearity, accuracy and precision.

Preparation of molecularly imprinted polymers for strychnine by precipitation polymerization and multistep swelling and polymerization and their application for the selective extraction of strychnine from nux-vomica extract powder.

Monodisperse molecularly imprinted polymers for strychnine were prepared by precipitation polymerization and multistep swelling and polymerization, respectively. In precipitation polymerization, methacrylic acid and divinylbenzene were used as a functional monomer and crosslinker, respectively, while in multistep swelling and polymerization, methacrylic acid and ethylene glycol dimethacrylate were used as a functional monomer and crosslinker, respectively. The retention and molecular recognition properties of the molecularly imprinted polymers prepared by both methods for strychnine were evaluated using a mixture of sodium phosphate buffer and acetonitrile as a mobile phase by liquid chromatography. In addition to shape recognition, ionic and hydrophobic interactions could affect the retention of strychnine in low acetonitrile content. Furthermore, molecularly imprinted polymers prepared by both methods could selectively recognize strychnine among solutes tested. The retention factors and imprinting factors of strychnine on the molecularly imprinted polymer prepared by precipitation polymerization were 220 and 58, respectively, using 20 mM sodium phosphate buffer (pH 6.0)/acetonitrile (50:50, v/v) as a mobile phase, and those on the molecularly imprinted polymer prepared by multistep swelling and polymerization were 73 and 4.5. These results indicate that precipitation polymerization is suitable for the preparation of a molecularly imprinted polymer for strychnine. Furthermore, the molecularly imprinted polymer could be successfully applied for selective extraction of strychnine in nux-vomica extract powder.

Simultaneous estimation of levodopa, carbidopa and 3-oxymethyldopa in rat plasma using HPLC-ECD.

The aim of study was to develop a suitable analytical method for simultaneous estimation of levodopa, carbidopa and 3-O-methyl dopa in rat plasma. Chromatographic separation of plasma samples was achieved using a reverse-phase C18 column. The mobile phase used consisted of a mixture of methanol and phosphate buffer (10 mM, pH 3.50) in the ratio of 90:10 v/v. All analytes were estimated by electrochemical detection at +800 mV. The developed method has been validated as per the standard guidelines. Precision study results were found to be satisfactory, with percentage relative standard deviation for repeatability and intermediate precision <3.96 and 6.56%, respectively, for all analytes detected in rat plasma. The developed method in rat plasma was found to be simple, rapid, accurate, precise and specific. The proposed method has been successfully applied for analysis of rat plasma samples obtained during an oral pharmacokinetic study of sustained release pellets of levodopa and carbidopa in rats. Copyright © 2016 John Wiley & Sons, Ltd.

Refractive index matching to develop transparent polyaphrons: Characterization of immobilized proteins

Refractive index matching was used to create optically transparent polyaphrons to enable proteins adsorbed to the aphron surface to be characterized. Due to the significant light scattering created by polyaphrons, refractive index matching allowed for representative circular dichroism (CD) spectra and acceptable structural characterization. The method utilized n-hexane as the solvent phase, a mixture of glycerol and phosphate buffer (30% [w/v]) as the aqueous phase, and the non-ionic surfactants, Laureth-4 and Kolliphor P-188. Deconvolution of CD spectra revealed that the immobilized protein adapted its native conformation, showing that the adsorbed protein interacted only with the bound water layer (“soapy shell”) of the aphron. Isothermal calorimetry further demonstrated that non-ionic surfactant interactions were virtually non-existent, even at the high concentrations used (5% [w/v]), proving that non-ionic surfactants can preserve protein conformation.

Development and Validation of a Novel Stability-Indicating RP-HPLC Method for the Simultaneous Determination of Related Substances of Ketoprofen and Omeprazole in Combined Capsule Dosage Form.

A novel, simple, sensitive, selective and reproducible stability-indicating high performance liquid chromatographic method was developed for the quantitative determination of degradation products and process-related impurities of ketoprofen (KET) and omeprazole (OMZ) in combined oral solid dosage form. Chromatographic separation was achieved on a Phenomenex Luna C18 (2) column (150 × 4.6 mm, 5 μm) under gradient elution by using a binary mixture of potassium dihydrogen phosphate buffer and acetonitrile at a flow rate of 0.8 mL/min. Chromatogram was monitored at 233 nm for KET impurities and at 305 nm for OMZ impurities using a dual wavelength UV detector. Resolution for KET and OMZ and 14 impurities was found to be >1.5 for any pair of components. Typical retention behaviors of impurities at various pH values were depicted graphically. To prove the stability-indicating power of the method, the drug product was subjected to hydrolytic, oxidative, photolytic, humidity and thermal stress conditions as per ICH. The developed method was validated according to the current ICH guidelines for specificity, limit of detection, limit of quantification, linearity, accuracy, precision, ruggedness and robustness.

Quantitative Estimation of Four Sartans in Presence of Hydrochlorothiazide in Pharmaceutical Preparations by High Performance Liquid Chromatography

A simple, reliable, reproducible, economical and rapid isocratic RP-HPLC method was developed for the simultaneously separated and quantification of four angiotensin II-receptor antagonists namely Telmisartan (TELM), Losartan (LOSA), Olmesartan (OLME) and Valsartan (VALS) along with thiazide diuretic mostly Hydrochlorothiazide (HCTZ). All the above said drugs were separated by using Welchrom C18 column having internal diameter of 4.6 X 250 mm with 5 µm particle size as stationary phase with mobile phase consisting a mixture of acetonitrile and phosphate buffer (pH-3.3, 50:50 v/v). The mobile phase was pumped at a flow rate of 1 ml/min. The wave length of the UV detection was done at 238 nm. All the sartans were separated within 6 minutes. The calibration curves were linear (r2 = 0.9998) in all cases. The relative standard deviation was less than 2% and average recovery was above 99.95%. All four sartans were routinely assayed without interference. This method was statistically validated in different parameters like linearity, precision, specificity, accuracy, robustness and ruggedness. The optimized method proved to be accurate, specific and robust for the quality control of four angiotensin-II receptor blockers alone or in combination with HCTZ in bulk drug and tablet dosage forms.

Simultaneous Estimation of Emtricitabine and Tenofovir Disoproxil Fumarate in Tablet Dosage Form by Reverse Phase High-performance Liquid Chromatography

A simple, rapid reversed-phase high performance liquid chromatographic method had been developed and validated for estimation of emtricitabine and tenofovir disoproxil fumarate in tablet dosage form. The estimation was carried out on Phenomenex- Luna C18 (25 cm x 4.60 mm, particle size 5 µm) column with a mixture of 10mM phosphate buffer (pH 6.8): Acetonitrile; 40: 60 (v/v) as mobile phase. UV detection was performed at 260 nm. The method was validated for linearity, accuracy, precision, specificity and sensitivity as per ICH norms. The developed and validated method was successfully used for the quantitative analysis of commercially available dosage form. The retention time was 2.81 and 7.42 min for emtricitabine and tenofovir disoproxil fumarate respectively and total run time was 10 min. at a flow rate of 1.0 mL/ min. The calibration curve was linear over the concentration range of 40.00 - 240.00 µg/ mL for emtricitabine and 60.00 - 360.00 µg/ mL for tenofovir disoproxil fumarate. The LOD and LOQ values were found to be 1.54 and 4.54 µg/ mL for emtricitabine and 4.60 and 13.65 µg/ mL for tenofovir disoproxil fumarate respectively. The high percentage of recovery and low percentage coefficient of variance confirm the suitability of the method for the simultaneous estimation of emtricitabine and tenofovir disoproxil fumarate in tablet dosage form.

Simultaneous Chemo/Enantioseparation and Assay of R-(+)-Rabeprazole and Related Impurities in Pharmaceutical Formulations

Simultaneous chiral and chemoseparation of R-(+)-rabeprazole and related (enantio)impurities was achieved on a cellulose tris-(3,5-dichlorophenylcarbamate) stationary phase chemically bonded to silica gel (Chiralpak IC). A gradient elution was applied in the reverse-phase separation mode. The mobile phase consisted of a mixture of acetonitrile and aqueous phosphate buffer at pH 7. The other operational parameters were flow rate of 1 mL min−1, column temperature of 35 °C and ultraviolet (UV) detection at 282 nm. Quantification limits for R-(+)-rabeprazole and the related impurities ranged in the interval of 0.02–0.03 %. Linear response intervals of 0.02–0.66 % were obtained with UV detection. Validation of the proposed method was achieved according to current regulations in force. For better understanding of the R-(+)-rabeprazole impurity profile, (+)-EMS/MS and MS/MS detection were also used.

Development and Validation of an HPLC Method for Determination of Amifostine and/or Its Metabolite (WR-1065) In Human Plasma Using OPA Derivatization and UV Detection.

A rapid, sensitive and reproducible HPLC method was developed and validated for the analysis of amifostine (AMF) and/or its metabolite, WR-1065 in human plasma. The method involves the alkylation of free sulfydryl group with iodoacetic acid followed by derivatization of the drug and its metabolite with o-phthaldialdehyde (OPA) and UVdetection at 340 nm. The derivatized AMF and WR-1065 were eluted in less than 11 min, and in the case of the metabolite with no interferences from the endogenous plasma peaks. Cystein was used as the internal standard. Analysis was carried out on a Eurosphere Performance (RP-18e, 100 × 4.6 mm) analytical column. The mobile phase was a mixture of methanol and phosphate buffer 0.03 M pH = 2.7 at a ratio of 40: 60v/v, respectively, with a flow rate of 1.5 mLmin(-1). Limit of detection was 0.5 µgmL(-1). The method involved a simple extraction procedure for AMF and/or its metabolite and analytical recovery was 90 ± 0.9%.The calibration curve was linear over the concentration range of 1-200 µgmL(-1). The coefficients of variation for intra-day and inter-day assays were less than 10%.

Effect of the p-tert-butylcalix[6]arene modifier on the electrochemical properties of the modified carbon paste electrode

The electrochemical properties of partially blocked electrodes obtained by mixing p-tert-butylcalix[6]arene with the paste of carbon-paste electrodes were studied. The differential capacitance, surface charge, and background current on the modified electrodes decreased relative to the same characteristics on the unmodified electrode for all supporting electrolytes under study (sodium chloride, sodium sulfate, hydrochloric acid, sulfuric acid, phosphoric acid, and phosphate buffer mixtures). The effect of p-tert-butylcalix[6]arene on Faraday currents was studied in the case of the anodic oxidation of vinpocetine and sodium ethylenediaminetetraacetate (EDTA). The signal to background ratio increased on the modified electrodes. At 3 wt % modifier, the signal-to-background ratio for the modified electrode was 2.8- and 3.3-fold higher for vinpocetine and sodium EDTA, respectively.

Development of a New UPLC Method for Simultaneous Determination of Valsartan, Amlodipine Besylate, and Hydrochlorothiazide Pharmaceutical Products

A new ultra-performance liquid chromatography method for the simultaneous determination of valsartan, amlodipine besylate, and hydrochlorothiazide in pharmaceutical formulations has been developed. Chromatographic separation was achieved within 1 min on a sub-2 μm RP18 column using an isocratic mode mobile phase consisting of a mixture of methanol and phosphate buffer (0.05 M, pH 3 ± 0.05) in the ratio 70:30 (v/v). The flow rate was set at 0.3 mL min−1 with a detection wavelength of 239 nm. The method was validated over the concentration ranges from 6.5 to 15.0 μg mL−1 for hydrochlorothiazide, from 5.0 to 12 μg mL−1 for amlodipine besylate and from 76.5 to 178.5 μg mL−1 for valsartan. Calibration plots were linear for valsartan, amlodipine besylate, and hydrochlorothiazide with a correlation coefficient greater than 0.99. Relative variation coefficients for repeatability and reproducibility were less than 3.0%. Recoveries from standard addition essay of individual component in pharmaceutical formulations ...

An efficient hydrophilic interaction liquid chromatographic method for the simultaneous determination of metformin and pioglitazone using high-purity silica column.

Hydrophilic interaction liquid chromatography (HILIC) provides a feasible approach to effectively separate polar compounds in complex matrices. Herein, a simple, reproducible and efficient HILIC method was developed for the simultaneous determination of pioglitazone. HCl (PIO) and metformin HCl (MET) in rabbit plasma. High-purity silica column was used for rapid and efficient separation of these co-administered drugs. The chromatographic parameters were optimized for best separation. The proposed HILIC system provides high separation efficiency with good peak shape compared to reversed phase (RP) chromatography. Additionally, a simple isocratic elution mode with a mobile phase composed of a mixture of methanol and 10mM phosphate buffer (pH 3.0) (94:6, v/v) was used and the effluent was monitored at 230nm. The method was validated in accordance with the requirements of US-FDA guidelines and was found to behave efficiently for the intended purpose. The correlation coefficient of 0.9992 was obtained in the concentration ranges of 0.5-100μgmL(-1). The limits of detection (S/N=3) and quantification (S/N=10) were 0.16 and 0.5ngmL(-1), respectively. The retention times were 3.4 and 5.0min for PIO and MET, respectively. Plasma levels were successfully determined in rabbit with satisfactory precision and accuracy. In addition, the stability tests in rabbit plasma proved reliable stability under the experimental conditions. The developed HILIC method was applied successfully to study the pharmacokinetic behaviors of the studied analytes in rabbit plasma after a single oral dose containing PIO and MET.

A novel approach for enantioseparation as applied to (RS)‐etodolac from pharmaceutical formulations: LC MS and density functional theory support for confirmation of diastereomers so separated

In the present studies formation of diastereomers of (RS)-etodolac was confirmed using LC-MS when [M + H](+) or [M](+) were recorded for the diastereomers. The lowest energy optimized structures of two diastereomers were drawn, which confirmed the three-dimensional geometry of the diastereomers. This supports the optimized analytical separation conditions. In addition, separation of diastereomers was successful using a C18 column and a binary mixture of methanol and triethyl ammonium phosphate buffer of pH 4.5 (80:20, v/v) as mobile phase at a flow rate of 1 mL min(-1) and UV detection at 223 nm. The separation method was validated as per International Conference on Harmonization guidelines. (RS)-Etodolac was isolated from commercial tablets and purified and characterized to be used as racemic standard. Three pairs of diastereomers were synthesized using enantiomerically pure amines, namely, (R)-(+)-α-methyl benzyl amine, (S)-(-)-α,4-dimethylbenzylamine and (R)-(-)-1-cyclohexylethylamine. Derivatization reactions were carried out under conditions of stirring at room temperature (30 °C for 2 h) as well as under microwave irradiation (MWI), and the two types of diastereomers were compared. Reaction conditions for derivatization were optimized with respect to mole ratio of chiral derivatizing agent and (RS)-etodolac and MWI time. No racemization was observed throughout the study.