APPLICATION OF LIQUID CHROMATOGRAPHY COUPLED WITH MASS SPECTROMETRY IN THE IMPURITY PROFILING OF DRUG SUBSTANCES AND PRODUCTS

As the drug safety and efficacy is hampered in the presence of an impurity, the international regulatory agencies laid down stringent limits for the control of impurities in the active pharmaceutical ingredient and pharmaceutical formulations. The conventional approaches lack the characterization of impurities in trace levels, due to sensitivity issues, hyphenated techniques are preferred. Among the modern hyphenated techniques, liquid chromatography-mass spectrometry (LC-MS) has high sensitivity and can analyze large number of organic compounds in a short period of time. In the present study, the impurity profiling of various drug substances and products using LC-MS about past 6 years were retrospect for its importance, instrumentations, and applications

VALIDATED CHIRAL RP-UFLC METHOD FOR THE QUANTIFICATION OF CHLORTHALIDONE IN BULK AND PHARMACEUTICAL DOSAGE FORM

Objective: Development and validation of new, simple and reliable enantioselective reverse phase ultra-fast liquid chromatography (RP-UFLC) method for quantification of chlorthalidone in bulk and pharmaceutical dosage form.Methods: In the present study, the isocratic RP-UFLC method was developed on Phenomenex® Lux cellulose 4 column (250×4.6 mm, 5µ) and di-sodium hydrogen phosphate buffer (pH 3.6): methanol (40:60 v/v) as mobile phase. Elute was monitored at 240 nm with a flow rate of 1 ml/min.Results: The described method provided linear correlation (R2=0.999) between the range of 2-10 µg/ml. Chlorthalidone enantiomers showed good resolution with a retention time (tR) of 5.75 min and 7.46 min respectively. The precision of the method revealed that relative standard deviation is within the acceptable limit. The percentage recovery of each chlorthalidone enantiomers was found to be 99.98% and 100.09% respectively. The method was validated in accordance with ICH harmonized tripartite guidelines, validation of analytical procedures: text and methodology Q2 (R1).Conclusion: An economical, accurate, sensitive and precise RP-UFLC method was developed and fully validated for quality control analysis of chlorthalidone in pharmaceutical dosage form

SIMULTANEOUS ESTIMATION OF CLOPIDOGREL AND ATORVASTATIN IN HUMAN PLASMA USING BIO-ANALYTICAL RP-ULTRA FAST LIQUID CHROMATOGRAPHIC METHOD

This manuscript depicts a simple, accurate, sensitive, precise and robust Ultra fast liquid chromatographic (UFLC) method developed and validated for the simultaneous determination Clopidogrel and Atorvastatin in human plasma as per USFDA draft guidelines. In the current study, the analysis was performed on phenomenex C8 (250 × 4.6 mm, 5μm) column using phosphate buffer (pH-2.5) and acetonitrile (50: 50 v/v) as mobile phase at flow rate of 1.3 mL/min. In the current developed method, Clopidogrel and Atorvastatin eluted at a retention time of 4.06 and 10.31 min respectively. The proposed method is having linearity in the concentration range from 10 to 50μg/mL of Clopidogrel and Atorvastatin. The current method was validated with respect to linearity; precision, lowest limit of detection (LLOD), accuracy and recovery according to the USFDA guidelines. The system consisted of a pump (Shimadzu, prominence, UFLC), with 20 µl sample injector, along with a PDA detector at a wavelength of 243 nm and 220 nm for Atorvastatin and Clopidogrel respectively. Data was compiled using Shimadzu LC Solution software. A good linear relationship over the concentration range of 10-50µg/ml was shown. Validation of the method was carried out as per the USFDA draft guidelines. The method developed was found to be precise, accurate, specific, linear and selective. Statistical analysis shows that the method is selective and reproducible for the estimation of Clopidogrel and Atorvastatin in combined dosage forms

EX VIVO ANTICOAGULANT ACTIVITY OF 1, 3, 4-OXADIAZOLE DERIVATIVES

Objective: The present medication for the management of arterial thromboembolism (ATE) disorders by anticoagulant therapy highlights its lacunae due to recurrent ATE episodes and indicates the need for better anticoagulant agents with clinical advantage.Methods: The anticoagulant study was performed for increase in prothrombin time (PT) and activated partial thromboplastin time (aPTT) at a test dose of 25 mg kg-1.Results: The results of ex vivo anticoagulant evaluation revealed that the tested compounds 3a-3q did not exhibit a significant increase in PT with respect to acenocoumarol (1 mg kg-1) employed as the reference drug for increase in PT. While the compounds, 3a-3q exhibited minimal increase in aPTT in comparison to unfractionated heparin (500 IU kg-1) employed as the reference drug for increase in aPTT. Among all the tested compounds, only compound 3q exhibited moderate anticoagulant activity with an increase in PT (33 ± 0.4 s) to that of the reference drug acenocoumarol (48 ± 0.5 s).Conclusion: The anticoagulant efficacy investigation highlights that the synthesized compound 3q could be considered for further clinical studies to ascertain its possible hit as anticoagulant agents.Â

NOVEL UV SPECTROPHOTOMETRIC METHOD FOR THE DETERMINATION OF TERIFLUNOMIDE IN TABLET DOSAGE FORM

Objective: The current work is intended towards the development of a novel, simple, and precise UV spectrophotometric method for the estimation of teriflunomide (TEF) present in the marketed formulation. Methods: Acetonitrile was used as asolvent and the absorbance of the drug was measured at the absorbance maxima of TEF, UV 284 nm. Results: Calibration curve plotted in concentration range 5-10 µg /ml exhibited excellent linear relationship with line equation y = 0.0858x-0.0223 and r2 value of 0.9996. The method was found to comply all the validation parameters as per the ICH guideline indicating the sensitivity of the method towards analyte. Conclusion: The method can be used satisfactorily for the routine analysis of TEF present in marketed formulation

RP-UFLC METHOD DEVELOPMENT AND VALIDATION FOR SIMULTANEOUS ESTIMATION OF CLOPIDOGREL, PANTOPRAZOLE AND ROSUVASTATIN IN HUMAN PLASMA: DRUG INTERACTION STUDIES

Objectives: Objective of our present study was to develop a novel ultra fast liquid chromatographic method for quantitative simultaneous estimation of Clopidogrel, Pantoprazole & Rosuvastatin in human plasma and to validate the developed method following USFDA guidelines.Methods: In the current study, the analysis was performed on phenomenex C8 (250 × 4.6 mm, 5μm) column using phosphate buffer (pH-2.5) and acetonitrile (45: 55 v/v) as mobile phase at flow rate of 1.2 mL/min. The system consisted of a pump (Shimadzu, prominence, UFLC), with 20 µl sample injector, along with a PDA detector at a wavelength of 254, 243 nm and 220 nm for Clopidogrel, Pantoprazole and Rosuvastatin respectively. Data was compiled using Shimadzu LC Solution software.Results: In this developed method Clopidogrel, Pantoprazole & Rosuvastatin, eluted at a retention time of 2.566, 5.002 and 9.301 min respectively. The proposed method is having linearity in the concentration range from 5 to 50μg/mL of Clopidogrel, Pantoprazole & Rosuvastatin. The current method was validated with respect to accuracy, linearity; precision, lowest limit of quantification (LLOQ) and recovery according to the USFDA guidelines.  A good linear relationship over the concentration range of 5-50µg/ml was shown. Validation of the method was carried out as per the USFDA draft guidelines.Conclusion: A novel specific, accurate, precise UFLC method was developed for quantitative simultaneous estimation of Clopidogrel, Pantoprazole & Rosuvastatin in human plasma and validated. The developed method is suitable and economic for routine analysis and pharmacokinetic studies of Clopidogrel, Pantoprazole & Rosuvastatin simultaneously. The method developed was found to be precise, accurate, specific, linear and sensitive. Statistical analysis shows that the method is reproducible and selective for the estimation of Clopidogrel, Pantoprazole & Rosuvastatin in dosage form of patient plasma.Â

NOVEL HPTLC-DENSITOMETRIC METHOD FOR THE ESTIMATION OF TERIFLUNOMIDE IN TABLET DOSAGE FORM

Objective: The current work is intended towards the development of a novel, simple and precise high-performance thin layer chromatographic (HPTLC) method coupled with a densitometer for the estimation of teriflunomide (TEF) present in the marketed formulation. Methods: The chromatographic development was performed on aluminum plates coated with silica gel 60 F254 using toluene: ethyl acetate: glacial acetic acid (7.5:2: 0.5 v/v/v) as the mobile phase. Densitometric scanning was achieved at the absorbance maxima, UV 284 nm. Results: Well separated band was observed with Rf value 0.46. The calibration curve plotted in the concentration range 100-700ng/band exhibited an excellent linear relationship with the r2 value of 0.9928. The method was found to comply with all the validation parameters as per the ICH guidelines. Conclusion: The method ensures minimal use of mobile phase with minimal run time compared to other reported analytical methods. This validated method can be used by quality control laboratories for the routine quantitative analysis of tablets consisting of Teriflunomide

DETERMINATION OF OCTANOL-WATER PARTITION COEFFICIENT OF NOVEL COUMARIN BASED ANTICANCER COMPOUNDS BY REVERSED-PHASE ULTRA-FAST LIQUID CHROMATOGRAPHY

Objective: The present study aims at the development of a reversed phase ultra-fast liquid chromatography (RP-UFLC) method for measurement of the lipophilicity (log P) between n-octanol and water for the newly synthesized coumarin derivatives in our laboratory.Methods: The synthesized compounds were dissolved in methanol and analyzed using XTerra RP18 column as the stationary phase and a mixture of methanol (0.25% v/v octanol) and buffer as the mobile phase with isocratic elution.Results: In this study we concentrated on the relationship between a reversed-phase ultra-fast liquid chromatography (RP-UFLC) retention parameters and log P of our synthesized compounds. Furthermore, a good correlation and very close values were obtained between the experimentally determined log P values and values obtained from Chemdraw.Conclusion: The developed method was found to be insensitive to any of the impurities present and moreover it requires very little sample for analysis

SPECTROPHOTOMETRIC METHOD FOR THE DETERMINATION OF AMIKACIN IN PURE AND PHARMACEUTICAL DOSAGE FORM

Objective: The aim of the study was to develop an easy, sensible and rapid method for the estimation of amikacin in both pure and marketed formulation using the spectrophotometric method.Methods: Due to lack of chromophoric group in the amikacin, it was derivatized with 0.1 mmol chloranillic acid reagent. For the estimation of amikacin, Shimadzu UV-1700 model spectrophotometer with UV probe software was used. The method was based on simple charge transfer complexation of the drug with a p-chloranillic acid reagent to give a purple coloured product which was measured at 524nm against blank solution.Results: The derivatised product of amikacin was detected at a wavelength of 524 nm. Linearity was observed with the concentration range of 20-100 µg/ml with a regression coefficient of 0.9803. Results of all the parameters were within the acceptance criteria with % RSD less than 2.Conclusion: The spectroscopic method was validated as per ICH guidelines and was found to be applicable for routine quantitative analysis of amikacin in marketed formulations also. The results of linearity, precision, accuracy LOD and LOQ were within the specified limits. The method is highly sensitive, robust, reproducible and specific.Â

Colorimetric Determination of Cefadroxil and Ceftriazone in Pharmaceutical Dosage Forms

Purpose: To develop a simple, rapid and selective method for the spectrophotometric determination of cefadroxil and ceftriaxone using 1, 2- napthaquinone-4- sulfonic acid sodium.Methods: The method was based on the derivatization of cefadroxil and ceftriaxone with 1, 2-naphthaquinone-4- sulfonic acid sodium in alkaline medium to yield orange-colored products.Results: The reaction products of cefadroxil and ceftriaxone at their respective max of 475 and 480 nm showed linearity in the concentration range of 10 - 100 and 25 - 175 ìg/ml, respectively. Relative standard deviations of 0.82 % for cefadroxil and 0.95 % for ceftriaxone were obtained. Recoveries of cefadroxil tablets and ceftriaxone injection were in the range of 100.66 ± 0.98 and 99.38 ± 0.84 %, respectively.Conclusion: Recovery studies gave satisfactory results indicating that none of the major additives/excipients interfered with the assay method. Therefore, the proposed method is simple, rapid, precise and convenient for the assay of cefadroxil and ceftriaxone in commercial preparations