Degradation kinetics of telithromycin determined by HPLC method

The degradation kinetics of the antibiotic telithromycin using a stability-indicating high-performance liquid chromatography (HPLC) method is demonstrated. The photodegradation is performed by UVC lamp-254 nm (15W), installed in a chamber internally coated with mirrors, where telithromycin solutions prepared from coated tablets are placed in quartz cells. To promote oxidation, the reaction between the telithromycin solution and 3% hydrogen peroxide solution is carried out. The kinetics parameters of order of reaction and the rate constants of the degradation are determined for both conditions. The degradation process of telithromycin can be described by first-order kinetics under both experimental conditions used in this study. The results reveal the photo and oxidation lability of the drug and confirm the reliability of HPLC method for telithromycin in the presence of its degradation products.     

Chromatographic determination of itopride hydrochloride in the presence of its degradation products

Two sensitive and reproducible methods are described for the quantitative determination of itopride hydrochloride (IH) in the presence of its degradation products. The first method is based on HPLC separation on a reversed phase Kromasil column [C18 (5-microm, 25 cm x 4.6 mm, ID)] at ambient temperature using a mobile phase consisting of methanol and water (70:30, v/v) adjusted to pH 4.0 with orthophosphoric acid with UV detection at 258 nm. The flow rate was 1.0 mL per min with an average operating pressure of 180 kg/cm2. The second method is based on HPTLC separation on silica gel 60 F254 using toluene:methanol:chloroform:10% ammonia (5.0:3.0:6.0:0.1, v/v/v/v) as mobile phase at 270 nm. The analysis of variance (ANOVA) and Student's t-test were applied to correlate the results of IH determination in dosage form by means of HPLC and HPTLC methods. The drug was subjected to acid and alkali hydrolysis, oxidation, dry heat, wet heat treatment, UV, and photodegradation. The proposed HPLC method was utilized to investigate the kinetics of the acidic, alkaline, and oxidative degradation processes at different temperatures and the apparent pseudo-first-order rate constant, half-life, and activation energy were calculated. In addition the pH-rate profile of degradation of IH in constant ionic strength buffer solutions in the pH range 2-11 was studied.     

High Pressure Liquid Chromatographic Determination of Promethazine Hydrochloride in the Presence of its Thermal and Photolytic Degradation Products: A Stability Indicating Assay

Abstract

A stability indicating method has been developed for the quantitation of promethazine hydrochloride in the presence of its photolytic and thermal degradation products. Following a basic extraction with acetonitrile, promethazine is separated from its internal standard, promazine, and vehicle components by direct high performance liquid chromatography using ultraviolet detection (249 nm) and a stainless steel column 25 cm in length, 0.46 cm i.d. packed with octa-decyl silica 5μ in diameter. A linear relationship was obtained between peak height ratio (promethazine/promazine) and promethazine hydrochloride in water over the range 30–600 g/ml. The percent coefficient of variation of the assay is 0.8% and the recovery of promethazine hydrochloride from aqueous solutions is 99.7%. The photolytic degradation of promethazine hydrochloride does not follow simple first order kinetics. Potassium iodide and p-benzoquinone had a significant effect on the degradation rate of promethazine during the first 30 minutes of the photolytic degradation reaction. However, after one hour there is no apparent quenching effect on the photolytic degradation rate of promethazine hydrochloride in the presence of these quenchers.     

Stability-indicating method for determination of oxyphenonium bromide and its degradation product by high-performance liquid chromatography

A high-performance liquid chromatographic (HPLC) method was developed for determination of oxyphenonium bromide (OX) and its degradation product. The method was based on the HPLC separation of OX from its degradation product, using a cyanopropyl column at ambient temperature with mobile phase of acetonitrile-25 mM potassium dihydrogen phosphate, pH 3.4 (50 + 50, v/v). UV detection at 222 nm was used for quantitation based on peak area. The method was applied to the determination of OX and its degradation product in tablets. The proposed method was also used to investigate the kinetics of the acidic and alkaline degradation of OX at different temperatures, and the apparent pseudo first-order rate constant, half-life, and activation energy were calculated. The pH-rate profile of the degradation of OX in Britton-Robinson buffer solutions within the pH range 2-12 was studied.     

A new HPLC–MS/MS method for investigating degradation kinetics of 6‴-feruloylspinosin and identifying its metabolites by rat intestinal bacterial flora in vitro

6?-Feruloylspinosin, a C-glycoside flavonoid, is the active ingredient responsible for the anxiolytic and sedative effects of Zizyphi Spinosiae Semen. However, the degradation mechanism of 6?-feruloylspinosin in vivo is largely unknown. In this study, we developed a simple and rapid method for determining the degradation of 6?-feruloylspinosinin in the bacterial culture using high-performance liquid chromatography (HPLC)–tandem mass spectrometry (MS). The precisions were less than 2.93%, and the accuracy (RE%) ranged from ?4.07 to 3.09%. The HPLC–MS/MS method was successfully applied to study the degradation kinetics of 6?-feruloylspinosin and identify its metabolic derivatives. The results indicated that the degradation kinetics and the rate constants were correlated with the concentration of sample solutions and the degradation process could be classified as apparent first-order kinetic reaction. Obviously, the degradation rate of 6?-feruloylspinosin was rather rapid as 80% of the compound could be decomposed in only 8–12?hr. Furthermore, two metabolites were identified from the serial hydrolysis of 6?-feruloylspinosin.     

Derivatization of secondary amines with 2-naphthalene-sulfonyl chloride for high-performance liquid chromatographic analysis of spectinomycin

A normal-phase high-performance liquid chromatographic (HPLC) method has been developed for the assay of spectinomycin hydrochloride and spectinomycin sulfate for detection at 254 nm. The method involves pre-column derivatization of secondary amines of spectinomycin with 2-naphthalenesulfonyl chloride (NSCl) using a catalyst. Lincomycin, 1-methylpyrrole, 2-acetyl-1-methylpyrrole, and 2-acetyl-pyrrole act as catalysts for sulfonylation of spectinomycin. Without a catalyst, the derivatization reaction forms a considerable amount of actinospectinoic acid, a degradation compound of spectinomycin, and peak area:weight ratio of the derivative is approximately 15% lower than those with the catalyst. Following derivatization the sample is extracted and chromatographed on a normal-phase silica column with detection at 254 nm. The method is applicable for the analysis of both the hydrochloride and sulfate salt forms of spectinomycin. All the known degradation compounds of spectinomycin such as actinamine, actinospectinoic acid and the biosynthesis intermediates, dihydrospectinomycin diastereoisomers, are completely separated with this method. Mass spectrometric data confirms that spectinomycin is derivatized with NSCl at the secondary amines located at positions 6 and 8 of the ring structure. The standard curves for the HPLC assay of spectinomycin hydrochloride and sulfate are linear with correlation coefficients of 0.9997 and 0.9999, respectively over the range of 0.05 mg/ml to 0.3 mg/ml. The relative standard deviations (R.S.D.) of the HPLC assay methods for spectinomycin hydrochloride and sulfate are 0.67% and 0.86%, respectively. Spectinomycin hydrochloride and sulfate bulk drugs were assayed by the HPLC method and compared to gas-liquid chromatography and microbiological assay results. The HPLC method was used to assay spectinomycin in a veterinary formulation, Linco-Spectin soluble powder. The sensitivity of the HPLC assay was determined to be approximately 4 ng sample load on the column, which suggests applicability in serum and residue level studies.     

Stability-indicating high-performance liquid chromatographic assay of buspirone HCl

The United States Pharmacopoeia high-performance liquid chromatographic (HPLC) assay method of buspirone is not able to discriminate buspirone from its degradation products. The purpose of this work is to develop a sensitive, selective, and validated stability-indicating HPLC assay for the analysis of a buspirone hydrochloride in a bulk drug. Buspirone HCI and its potential impurities and degradation products are analyzed on an Ultrasphere C18 column heated to 40 degrees C using a gradient program that contains monobasic potassium phosphate buffer solution (pH 6.9) and acetonitrile-methanol mixture (13:17) of 35% for 5 minutes, then increased to 54% in 5.5 minutes. The samples are monitored using a photo-diode array detector and integrated at 244 and 210 nm. The stress testing of buspirone HCI shows that buspirone acid hydrochloride is the major degradation product. The developed method shows a separation of buspirone degradation product and its potential impurities in one run. The stability of buspirone HCI is studied under accelerated conditions in order to provide a rapid indication of differences that might result from a change in the manufacturing process or source of the sample. The forced degradation conditions include the effect of heat, moisture, light, acid-base hydrolysis, sonication, and oxidation. The compatibility of buspirone HCI with some pharmaceutical excipients is studied under stress conditions. The linear range of buspirone HCI is between 5 and 200 ng/microL with a limit of quantitation of 2.5 ng/microL. The intraassay percentage deviation is not more than 0.38%, and the day-to-day variation was not more than 0.80%. The selectivity, repeatability, linearity, range, accuracy, sample solution stability, ruggedness, and robustness show acceptable values.     

Determination of doxycycline in pharmaceuticals based on its degradation by Cu(II)/H<Subscript>2</Subscript>O<Subscript>2</Subscript> reagent in aqueous solution

The reaction between doxycycline, hydrogen peroxide and Cu(II) was investigated spectrophotometrically in aqueous solutions. The absorption spectra show that the degradation of antibiotic drug occurs in weak alkaline aqueous solutions. The kinetics of the reaction was studied by applying the initial-rate method and the relative rate constants were calculated at different temperatures. The kinetic-spectrophotometric method for the determination of doxycycline based on the kinetics of pseudo-first order reaction at 20°C is proposed. The effects of several molecules and ions usually present in pharmaceutical formulations were studied in order to assess their interference. The calibration graph was applied to the determination of doxycycline in capsule dosage form. The relative standard deviation was no greater than 3.80%. The results were compared to those obtained by the official HPLC method.     

A quantum-chemical investigation of deoxypeganine and its salts II. The photochemical oxidation of deoxypeganine hydrochloride

The stability of deoxypeganine hydrochloride under the action of light has been established. The kinetics of the oxidation of deoxypeganine hydrochloride in aqueous solution in the presence of deoxyvasicinone hydrochloride have been studied. The structures and electronic conformations of hypothetical models of the salt DOP HCl in the ground and excited states have been considered by the MO LCAO method in the AM1 approximation. The difference in the resistance of the molecules of the base deoxypeganine and its hydrochloride to photochemical oxidation is discussed on the basis of the results of experiments and calculations.Abbreviations HPLC high-performance liquid chromatography - MO LCAO molecular orbitals as linear combinations of atomic orbitals - AM1 Austin Model 1 - DOP HCl deoxypeganine hydrochloride - DOV HCl deoxyvasicinone hydrochloride Institute of the Chemistry of Plant Substances, Academy of Sciences of the Republic of Uzbekistan, Tashkent. Translated from Khimiya Prirodnykh Soedinenii, No. 5, pp. 729–733, September–October, 1993.     

Forced degradation studies of norepinephrine and epinephrine from dental anesthetics: Development of stability-indicating HPLC method and in silico toxicity evaluation

Injectable solutions containing epinephrine (EPI) and norepinephrine (NE) are not stable, and their degradation is favored mainly by the oxidation of catechol moiety. As studies of these drugs under forced degradation conditions are scarce, herein, we report the identification of their degradation products (DP) in anesthetic formulations by the development of stability-indicating HPLC method. Finally, the risk assessment of the major degradation products was evaluated using in silico toxicity approach. HPLC method was developed to obtain a higher selectivity allowing adequate elution for both drugs and their DPs. The optimized conditions were developed using a C18 HPLC column, sodium 1-octanesulfonate, and methanol (80:20, v/v) as mobile phase, with a flow rate of 1.5 mL/min, UV detection at 199 nm. The analysis of standard solutions with these modifications resulted in greater retention time for EPI and NE, which allow the separation of these drugs from their respective DPs. Then, five DPs were identified and analyzed by in silico studies. Most of the DPs showed important alerts as hepatotoxicity and mutagenicity. To the best of our acknowledgment, this is the first report of a stability-indicating HPLC method that can be used with formulations containing catecholamines.     

Evaluation of the pattern and kinetics of degradation of adalimumab using a stability‐indicating orthogonal testing protocol

Forced degradation studies are crucial for the evaluation of the stability and biosimilarity. Here, adalimumab was subjected to oxidation, pH, temperature, agitation and repeated freeze–thaw in order to generate all possible degradation products. An orthogonal stability‐indicating testing protocol comprising SE‐HPLC, RP‐HPLC, TapeStation gel electrophoresis, dynamic light scattering (DLS), and functional receptor binding assay was developed and validated. The assay protocol was used for the assessment of the pattern and kinetics of aggregation/degradation of adalimumab. SE‐HPLC and DLS were used to show the formation of aggregates/fragments of adalimumab under nondenaturing conditions. TapeStation electrophoresis was performed under denaturing conditions to reveal the nature of aggregates. Results of the receptor binding assay agreed to those of SE‐HPLC and DLS which indicated that it can be used as an activity‐indicating assay for adalimumab. RP‐HPLC demonstrated excellent selectivity for adalimumab in the presence of its oxidized forms. The kinetics of degradation was studied in each case and the results showed that it followed the first‐order reaction kinetics. Correlation between the results supported the quality assessment of the tested product in industrial and clinical settings. This orthogonal protocol is a useful tool in stability assessment of monoclonal antibodies and a key criterion for the biosimilarity assessment.     

Development and validation of an HPLC method for tetracycline‐related USP monographs

A novel reversed‐phase HPLC method was developed and validated for the assay of tetracycline hydrochloride and the limit of 4‐epianhydrotetracycline hydrochloride impurity in tetracycline hydrochloride commercial bulk and pharmaceutical products. The method employed L1 (3 µm, 150 × 4.6 mm) columns, a mobile phase of 0.1% phosphoric acid and acetonitrile at a flow rate of 1.0 mL/min, and detection at 280 nm. The separation was performed in HPLC gradient mode. Forced degradation studies showed that tetracycline eluted as a spectrally pure peak and was well resolved from its degradation products. The fast degradation of tetracycline hydrochloride and 4‐epianhydrotetracycline hydrochloride in solution was retarded by controlling the autosampler temperature at 4 °C and using 0.1% H₃PO₄ as diluent. The robustness of the method was tested starting with the maximum variations allowed in the US Pharmacopeia (USP) general chapter Chromatography <621>. The method was linear over the range 80–120% of the assay concentration (0.1 mg/mL) for tetracycline hydrochloride and 50–150% of the acceptance criteria specified in the individual USP monographs for 4‐epianhydrotetracycline hydrochloride. The limit of quantification for 4‐epianhydrotetracycline hydrochloride was 0.1 µg/mL, 20 times lower than the acceptance criteria. The method was specific, precise, accurate and robust. Copyright © 2014 John Wiley & Sons, Ltd.     

Development of validated stability-indicating chromatographic method for the determination of fexofenadine hydrochloride and its related impurities in pharmaceutical tablets

ABSTRACT: A simple reversed phase high performance liquid chromatographic method with diode array detector (HPLC-DAD) has been developed and subsequently validated for the determination of fexofenadine hydrochloride (FEX) and its related compounds; keto fexofenadine (Impurity A), meta isomer of fexofenadine (Impurity B), methyl ester of fexofenadine (Impurity C) in addition to the methyl ester of ketofexofenadine (Impurity D). The separation was based on the use of a Hypersil BDS C-18 analytical column (250 × 4.6 mm, i.d., 5 μm). The mobile phase consisted of a mixture of phosphate buffer containing 0.1 gm% of 1-octane sulphonic acid sodium salt monohydrate and 1% (v/v) of triethylamine, pH 2.7 and methanol (60:40, v/v). The separation was carried out at ambient temperature with a flow rate of 1.5 ml/min. Quantitation was achieved with UV detection at 215 nm using lisinopril as internal standard, with linear calibration curves at concentration ranges 0.1-50 μg/ml for FEX and its related compounds. The optimized conditions were used to develop a stability-indicating HPLC-DAD method for the quantitative determination of FEX and its related compounds in tablet dosage forms. The drugs were subjected to oxidation, hydrolysis, photolysis and heat to apply stress conditions. Complete separation was achieved for the parent compounds and all degradation products. The method was validated according to ICH guidelines in terms of accuracy, precision, robustness, limits of detection and quantitation and other aspects of analytical validation.     

Influence of medium and temperature on the hydrolysis kinetics of propacetamol hydrochloride: determination using derivative spectrophotometry

Propacetamol hydrochloride (PRO) is a water-soluble prodrug of paracetamol (PA) which can be parenterally administered as analgesic for the treatment of postoperative pain, acute trauma, and gastric and/or intestinal disorders where oral administration is not possible. In these circumstances, PRO can be administered in physiologic or glucose solutions since it is rapidly and quantitatively hydrolyzed into PA by plasma estearases. We have studied the degradation kinetics of PRO in 5% glucose and 0.9% saline solutions at 4 degrees C and 25 degrees C (storage and room temperatures, respectively). The analytic technique used to determine PRO and PA quantitatively was first-derivative spectrophotometry. The degradation process of PRO can be best fitted to a second-order kinetics with independence of the medium used (saline or glucose solution). The hydrolysis kinetics of PRO conversion into PA depends on the temperature but not on the assay medium (saline or glucose solution). The degradation rate constants obtained for PRO were approximately 4.5 times higher at 25 degrees C than at 4 degrees C. The values of t(90%) for PRO were 3.17 h and 3.61 h at 25 degrees C, and 13.42 h and 12.36 h at 4 degrees C when the tests were performed in 5% glucose and 0.9% saline solutions, respectively.     

A quantitative HPLC method for determining lactide content using hydrolytic kinetics

A new method for quantitative analysis of lactide has been developed by applying chemical kinetics to a HPLC system. The most important advance is its practical approach to the quantification of analytes that are unstable in the HPLC mobile phase. In HPLC analysis, anhydrous mobile phases cannot separate lactide from impurities, and only mixtures of water and organic solvent can achieve effective separation. By selecting conditions for testing and studying the kinetics of lactide hydrolysis, extensive experiments revealed that lactide degradation can be treated as a pseudo-first-order reaction under the given HPLC conditions, and lactide content or purity can be quantitatively determined. This method is practical for measuring the purity of the intermediate lactide in polylactic acid (PLA) production and the lactide content in PLA. When lactide content is high, the relative standard deviation (RSD) of the measurements is <2.0%, while RSD is <5.0% at low levels, which indicates that the method is suitably precise.     

Validated HPLC and HPTLC stability-indicating methods for determination of alfuzosin hydrochloride in bulk powder and pharmaceutical formulations

Two sensitive, selective, and precise stability-indicating, high-performance liquid chromatography and high-performance thin-layer chromatography methods have been developed for the determination of alfuzosin hydrochloride in the presence of its degradation products. Alfuzosin.HCl was subjected to stress alkaline, acidic, oxidative, thermal, and photo-degradation. The drug could be well separated from the degradation products upon applying the two methods. Separation by HPLC was achieved using an Xterra RP18 column and acetonitrile/0.02 M KH2PO4 (pH=3) in a ratio of 20:80 as mobile phase. The flow rate was 1 mL/min. The linearity range was 0.25 to 11 microg/mL with mean percentage recovery of 100.26 +/- 1.54. The HPTLC method used ALUGRAM Nano-SIL silica gel 60 F254 plates; the optimized mobile phase was methanol/ammonia (100:1.2). Quantitatively the spots were scanned densitometrically at 245 nm. A second order polynomial equation was used for the regression. The range was 0.5-7 microg/spot. The mean percentage recovery was 100.13 +/- 1.67. Two main degradation products were obtained in most stress conditions, separated, and identified by FT-IR and NMR spectral analysis, from which the degradation pathway was proposed. The two methods were validated according to the International Conference on Harmonization. In addition, the HPLC method was used to study the kinetics of alkaline and acid degradation of the drug.     

A Rapid Stability-Indicating LC Method for Ziprasidone Hydrochloride

A simple and rapid reversed-phase liquid chromatographic method was developed for the related substances determination and quantitative evaluation of ziprasidone hydrochloride, which is used as an antipsychotic agent. Forced degradation studies were performed on bulk sample of ziprasidone hydrochloride using acid, base, oxidative hydrolysis, thermal stress and photolytic degradation. Mild degradation of the drug substance was observed during thermal stress and considerable degradation observed during base hydrolysis. The chromatographic method was fine tuned using the samples generated from forced degradation studies. Good resolution between the peaks corresponds to synthetic impurities and degradation products from the analyte were achieved on YMC Pack Pro C18 column using the mobile phase consists of a mixture of 0.05% v/v of phosphoric acid in water and acetonitrile. The stressed test solutions were assayed against the qualified working standard of ziprasidone hydrochloride and the mass balance in each case was close to 99.7% indicating that the developed method was stability-indicating. Validation of the developed method was carried out as per ICH requirements.     

Validation of UV Spectrophotometric Method for Fexofenadine Hydrochloride in Pharmaceutical Formulations and Comparison with HPLC

Abstract

A simple, reproducible, accurate, and effective spectrophotometric method was developed and validated for the quantitation of the antihistamine fexofenadine in capsules and coated tablets. Ethanol was used as solvent and the absorbance at the wavelength of 220 nm was employed to the quantitation of the drug. The method validation was fulfilled through the evaluation of the analytical parameters of linearity, precision, accuracy, limits of detection, and quantitation and specificity. The method was linear (r=0.9999) at concentrations ranging from 8.0 to 20.0 µg ml^?1, precise (RSD intra‐day=0.29; 0.18; 0.39; RSD inter‐day=0.12 for capsules and RSD intra‐day=0.13; 0.16; 0.13; RSD inter‐day=0.13 for coated tablets), accurate (percentage recovery=99.97% for capsules and 100.51% for tablets), sensitive (limits of detection and quantitation of 0.10 and 0.29 µg ml^?1, respectively) and specific. The method was compared to a high performance liquid chromatography (HPLC) method, which was previously developed to the same drug. The results showed no significant difference between the methods in fexofenadine hydrochloride quantitation.     

Assay of four stereoisomers of desacetyl diltiazem hydrochloride. Application toin vitro chiral inversion studies

Summary Direct resolution of four stereoisomers of the related compound of diltiazem hydrochloride, namely desacetyl diltiazem hydrochloride, was studied by both normal and reversed-phase chiral HPLC. The four stereoisomers were completely resolved on a Chiralcel OF column. The technique developed was applied to a chiral inversion study of desacetyl diltiazem hydrochloride. This inversion was observed neither in the solid state, in aqueous solution at 100°C for 3 h nor under visible light for 10h, but was observed in aqueous solution under UV irradiation.The (+)-(2S, 3S)-cis-desacetyl diltiazem hydrochloride degraded with a half-life of 1.9 h in aqueous solution under UV and epimerized to (+)-(2R, 3S)-trans-desacetyl diltiazem hydrochloride. Similarly, (+)-(2S, 3S)-cis-desacetyl diltiazem hydrochloride degraded about three times faster than diltiazem hydrochloride. Reverse epimerization of (+)-(2R, 3S)-trans-desacetyl diltiazem hydrochloride to (+)-(2S, 3S)-cis-desacetyl diltiazem hydrochloride was not observed.The overall degradation was the result of two competitive processes, the epimerization and the decomposition of the benzothiazepin ring. The degradation and epimerization rate of (+)-(2S, 3S)-cis-desacetyl diltiazem hydrochloride in solution under UV depended upon the solvent, the aqueous pH, and concentration.     

Development and validation of an HPLC method for mefloquine hydrochloride determination in tablet dosage form

A simple HPLC method for determination of mefloquine hydrochloride in tablets was developed and validated. The separation was carried out on an Xterra RP18 (250 x 4.6 mm id, 5 pm particle size) analytical column. The mobile phase was 0.05 M monobasic potassium phosphate buffer (pH 3.5)-methanol (40 + 60, v/v). The flow rate and wavelength were set to 1 mL/min and 283 nm, respectively. The method was specific for mefloquine hydrochloride in the presence of hydrolytic, oxidative, and photolytic degradation products. It was also linear, precise, accurate, and robust, being suitable for routine QC analyses and stability studies. The developed HPLC method was compared to a previously described spectrophotometric method.