Validation

ABSTRACTAn efficient and simple RP-HPLC method has been developed and validated for simultaneous determination of Repaglinide (REPA) and Metformin Hydrochloride (MET). The separation was carried out using mobile phase consisting of Phosphate buffer: Methanol (30:70). The column was used Luna 5u C18 (2) 100A of size 0.25m *4.6mm with flow rate of 1.0 mL/min. Drug peaks were well separated and were detected by a UV detector at 260 nm.. The described method was linear over concentration range of 10-50 ppm for assay of REPA and MET. The retention time of REPA and MET was found to be 4.31. The method has been validated according to ICH guidelines with respect to system suitability, specificity, precision, accuracy, ruggedness and robustness. Metformin limit of detection (LOD) and limit of quantification (LOQ) were 0.0386 mg/ml and 0.1169mg/ml respectively while LOD and LOQ for Repaglinide were 0.0339?g/ml and 0.1025?g/ml respectively and thus can be successfully applied for the routine analysis of REPA and MET in bulk and marketed dosage forms. Keywords: Repaglinide, Metformin Hydrochloride, RP-HPLC, Simultaneous estimation, Validation

ABSTRACTA new precise, accurate, robust and stability indicating high-performance liquid chromatographic method has been developed and validated for the simultaneous estimation of domperidone (DOM) and esomeprazole (ESO) in their pharmaceutical formulation. The proposed method carried out on Waters Symmetry C18 column (250 mm x 4.6 mm, 5.0 µm particle size) using an isocratic elution technique at a column temperature of 30 ?C. The mobile phase was a mixture of 0.01M sodium acetate buffer: methanol (45:55 v/v) and it was adjusted to pH 4.5 using glacial acetic acid with a flow rate of 1 mL/min and an injection volume of 20 µL. The retention times were 3.7 and 5.0 min with UV detection at 290 nm for DOM and ESO, respectively. The proposed method was linear over the concentration ranges of 0.04-60.0 and 0.08-120.0 µg/mL for DOM and ESO, respectively. Limit of detection and limit of quantitation values were 0.48 and 1.44 µg/mL for DOM and 0.47 and 1.43 µg/mL for ESO, respectively. The method also exhibited good levels of recovery from 100.23% to 101.59% for DOM and from 99.74% to 101.57% for ESO. From the validation study, it was found that the method was specific, rapid, accurate and reproducible. The high percentage of recovery and low relative standard deviation confirm the suitability of the method for routine pharmaceutical analysis of both drugs separately or in their combined dosage form. Keywords: HPLC; Esomeprazole; Domperidone; Validation; Formulation.

To assess the efficacy of therapeutic on Rotavirus diarrheal hospitalizations among children < 5 years of age   based on world health organization (WHO) guideline. Based on a longitudinal observational study, 1027 fecal samples were collected from children (less than 5 years), suffering from diarrhea attended at the Yemeni Swedish hospital (YSH) in Taiz , Yemen from January 2009  to December 2012 .  Rotavirus infection was detected by re – validated enzyme linkage immunosorbent assay (ELISA) on stool specimens of children. The treatment course consists of  two methods, namely, intravenous rehydration fluid therapy  (IV) for inpatient and oral rehydration fluid  therapy (OR) for outpatient with treatment of the major  symptoms, namely,  fever and vomiting based on anti – pyretic and anti – emetic if necessary . The efficacy of therapy quality outcomes was assessed clinically and reported. Firstly, the results of re - validated  ELISA method were precise to each analyte with percent relative standard deviation (RSD %) of intra-assay and inter – assay (< 5.0 %). Furthermore, the interval of accuracy for the method exhibited well recovery value of 93 - 100 % and the coefficient correlation (R2) value was  0.99 as a good linear method for Rotavirus infection. Secondly ,  A total of  581  out of  1027  (56.57 %) patients were admitted as inpatients for IV  fluid  therapy  ,  while  446  (43.43 %) were seen in the outpatient ward  receiving  OR  fluid  therapy . The recovery of patients was 98.10 % for IV and 98.43 % for OR, statistically, that was not significantly different (p> 0.05) . In conclusion , A successful   Rotavirus  treatment guidelines in Yemen will rely upon best sustained diagnosis by application Good Laboratory Practice (GLP)  which is clear in specific  – precise , reliable and  accurate method to detect the virus  and the best efficacy of therapy by Good Clinical Practice (GCP) which is clear in treatment  of  Rotavirus diarrhea which protects against dehydration by fluid therapy .

The current paper was a shot to style a sustained unharness dosage kind victimisation varied grades of hydrophilic polymers, Hypromellose or Hydroxy- propyl alkyl radical polysaccharide (HPMC K15M, HPMC K100M) and Chitosan additionally incorporated as rate retarding material. Laboratory scale batches of Six tablet formulations were ready by wet granulation technique (Low shear). Micromeritic properties of the granules were evaluated before compression. Tablets were characterized as crushing strength, friability, weight variation, thickness, Drug content or assay and evaluated for in-vitro unharness pattern for fifteen unit of time victimisation Phosphate buffer of suitable pH at 37±0.5°C. Results and discussion: The results obtained discovered that HPMC K15M, Chitosan and Ethylcellulose at an acceptable concentration formulation (F6) was able to sustain the drug unharness for fifteen hours and followed Higuchi pattern similar Fickian diffusion. moreover production validation scale batches were designed supported laboratory scale best batch and charged for stability testing. it had been found that every one parameters were at intervals the limit of acceptance. There was no chemical interaction found between the drug and excipients throughout FT-IR and DSC study thought of in the present investigation. therefore it may be over that combinely polymers at an acceptable concentration will effectively be formulate to sustain the drug unharness.

The simple, sensitive, reliable and economically new method was developed for the estimation of Ramipril (RAM) and Telmisartan (TEL) by RP-HPLC in combined dosage form. After several trials with the different combinations and ratios of solvents, the present chromatographic parameters were optimized. It was found that potassium dihydrogenphosphate (pH 3.0): methanol: acetonitrile (30:20:50 v/v/v) was given satisfactory results. A C18 column (Agilent ODS UG 5 column) having dimensions of 4.5mmx250mm was used. The mobile phase was pumped at a flow rate of 1.0ml/min and the eluents were monitored at 210nm. System suitability was carried out by injecting six replicate injections of 100% standard concentration, number of theoretical plates, HETP(height equivalent Theoretical plate) and resolution were satisfactory. The optimized chromatograms confirm the presence of Ramipril and Telmisartan at Rt: 4.1 min and Rt: 5.11min respectively without any interference. The concentration range of 1-5µg/ml for RAM and 8-40µg/ml for TEL were linear with correlation coefficients 0.999 and 0.989 respectively. The percent recovery studies were found to be 99.5-99.88% and 99.93-99.99% w/w for RAM and TEL respectively which indicate method was accurate. The proposed method was precise and reproducible with %RSD of 0.93 for Ramipril and 0.41 for Telmisartan, respectively. The limits of detection and limits of quantification were 0.10µg/ml and 0.25µg/ml for Ramipril 0.32µg/ml and 0.78µg/ml for Telmisartan, respectively. The method was found to be robust and ruggedness and was well suitable for the estimation of commercial formulations of selected combinations.

A simple, sensitive, selective and stability indicating UPLC method has been developed for the quantitative determination of potential impurities of allopurinol active pharmaceutical ingredient. Allopurinol, its five impurities and degradation products were separated efficiently by using the mobile phase consisted of sodium perchlorate (10 mM, pH 3.0)and acetonitrile on a HSS T3P stationary phase in gradient elution profile. Forced degradation study confirmed that the newly developed method was specific and selective to the degradation products. The newly developed UPLC method was validated according to ICH guidelines considering five impurities to demonstrate specificity, precision, linearity, accuracy and stability indicating nature of the method. Regression analysis showed correlation coefficient value greater than 0.99 for allopurinol and its five impurities. Detection limit of impurities was in the range of 0.002–0.006% indicating the high sensitivity of the newly developed method. Accuracy of the method was established based on the recovery obtained between 91.7% and 106.6% for all impurities.

Two simple, sensitive selective accurate and economical spectrophotometric methods Method A&B for the determination of feropenem in bulk drug and pharmaceutical formulations (tablets) have been described in the present work. Method - A is based on the formation of  red colored ion-association complex between feropenem and methiline blue (MB) exhibiting absorption maximum at 650nm and obeying Beer’s law in the concentration range of 4-20µg/ml. The Method - B is based on the formation of Ion-association complex between feropenem and safranine-o(SFO) to yield an yellow colored chromogen exhibiting absorption maximum at 520nm(Method B) and obeying Beer’s law in the concentration range of 4-20µg/ml. Statistical analysis of the results has been carried out for the proposed methods revealing high accuracy and good precision. The proposed methods developed by the author could be successfully extended to the commercial pharmaceutical formulations (tablets) containing feropenem.

A simple RP-HPLC compatible method for the injectable suspension dosage of combined medroxyprogesterone acetate and estradiol cypionate was developed and validated according to ICH and USP guidelines. The chromatographic separation was achieved by using the Zorbax Eclipse C18 column (50 mm×4.6 mm, 2.7 µm) with gradient elution technique at a flow rate of 1.0 ml/min. The UV detection was performed at 225 nm. The linearity of medroxyprogesterone acetate over the concentration range was 49.65 to 744.69 μg/ml and 10.15 to 152.28 μg/ml for estradiol cypionate respectively. The accuracy was evaluated by means of spike recovery method and the result showed in the range of 99.7% to 101.4% for medroxyprogesterone acetate and 98.6% to 101.8% for estradiol cypionate. The specificity of the method showed that the analyte was not interfered by the presence of co-formulated substances. The robustness of the study was found agreeable; hence it proves that the method was robust. The stability of the analyte was found stable for 24 hours. The developed method was successfully employed for the determination of combined medroxyprogesterone acetate and estradiol cypionate in injectable suspension dosage forms.

A simple, accurate, precise and rapid UV spectrophotometric simultaneous equation method has been developed for the simultaneous estimation of Melatonin and Pyridoxine in tablet dosage form. The stock solution was prepared in methanol. 243 and 271nm, an absorbance maxima were selected for Melatonin and Pyridoxine respectively. Beer’s law obeyed the concentration range of 3-9 μg/ ml and 10-30 μg/ml, for Melatonin and Pyridoxine. The results of analysis were validated statistically and by recovery studies. Accuracy and precision are within the limit. The % RSD for the recovery study was less than 2. The developed method was validated as per International conference Harmonization (ICH) guideline for its accuracy, precision, Limit of detection and Limit of quantitation. The proposed method can be effectively applied for the simultaneous estimation of both drugs in tablet dosage form.

A simple, accurate, and precise HPTLC method has been developed and validated for the simultaneous estimation of Atorvastatin Calcium (ATO) and Aspirin (ASP) from bulk drug and Dosage form. The method employed TLC aluminum plates precoated with silica gel 60 GF 254 as the stationary phase. The solvent system comprised Toluene: Ethyl Acetate: Methanol: Acetic acid [7:2:1.5:0.1v/v/v/v]. This system was found to give good result for both the drugs (Rf value: of ASP 0.43cm and ATO 0.53cm). Spectrodensitometric scanning-integration was performed at a wavelength of 235nm.The calibration curve was found to be linear within the concentration range of 20ng/spot to 100ng/spot for ATO and 30 to 150ng/spot for ASP. The regression data for calibration curve shows good linear relationship with r2 = 0.9989 and 0.9990 for ATO and ASP respectively. The method was validated in accordance with the requirements of ICH guidelines. The method was successfully applied for determination of drug in bulk and Dosage form. Thus, the proposed method can be used successfully for routine analysis of ATO and ASP from bulk and dosage form.