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Showing papers in "Journal of analytical and bioanalytical techniques in 2011"


Journal ArticleDOI
TL;DR: This paper attempts to shed light on some of the critical points to consider when performing bioanalytical method development and GLP compliant validation activities from a scientist’s perspective.
Abstract: Recently, there has been a heightened public awareness of drug safety across the globe. Nonclinical and clinical pharmacokinetic and toxicokinetic toxicology safety studies all require that the study samples be analyzed under the auspices of good laboratory practice (GLP) standards. GLPs are followed in order to ensure that safety studies are reliable and accurate. Several countries have issued or are in the process of issuing their own versions of bioanalytical guidance documents for performing method validation activities; however, each one is slightly different. These differences often make complying with regulatory requirements difficult and cumbersome. Several networking organizations have been working diligently to harmonize the various global bioanalytical guidance documents. This paper attempts to shed light on some of the critical points to consider when performing bioanalytical method development and GLP compliant validation activities from a scientist’s perspective.

59 citations


Journal ArticleDOI
TL;DR: In this paper, simple, accurate and reproducible UV spectrophotometric and HPLC method for simultaneous estimation of salbutamol and prednisolone (PRE) was developed.
Abstract: Simple, accurate, and reproducible UV spectrophotometric and HPLC method for simultaneous estimation of salbutamol (SAL) and prednisolone (PRE) was developed in the present work. The first developed method was Simultaneous equation method, wavelength selected are 227 nm for salbutamol and 244 nm for prednisolone respectively. Linearity was observed in concentration range of 6-20µg/ml for salbutamol as well as for prednisolone. Second developed method was RP-HPLC method using Thermo C18 column (4.6 mm i.d × 250 mm) and acetonitrile: 0.025M potassium dihydrogen orthophosphate buffer (pH adjusted to 3.5 with orthophosphoric acid) in the ratio of 30:70% v/v as mobile phase. For HPLC method, linearity was observed in the concentration range of 20-100µg/ml for salbutamol as well as for prednisolone and drugs was subjected to oxidation, hydrolysis, and heat to apply stress condition for degradation studies. Results of analysis were validated stastically and by recovery studies.

37 citations


Journal ArticleDOI
TL;DR: In this paper, a stability indicating ultra perfomance liquid chromatography (UPLC) method was developed for the simultaneous determination of sumatriptan succinate and Naproxen sodium in pharmaceutical dosage forms.
Abstract: A stability- indicating ultra perfomance liquid chromatography (UPLC) method was developed for the simultaneous determination of sumatriptan succinate and Naproxen sodium in pharmaceutical dosage forms. The chromatographic separation was achieved on C18, 50 mm × 4.8 mm, 1.8-μm particle size column. The mobile phase contains a mixture of 0.2% ortho phosphoric acid and acetonitrile as the mobile phase in gradient elution technique. The retention time of Sumatriptan and Naproxen was found to be 1.7 and 2.7 min respectively. The total runtime was 5 min within which two active compounds and degradation products were separated. This method allows the determination of 850-2565 μg mL -1 of sumatriptan succinate and 5000-15000 μg mL -1 of Naproxen sodium. The flow rate was 1.0 mL min -1 and the detection wavelength was 225 nm. The limit of detection (LOD) for sumatriptan succinate and Naproxen sodium was 1.9 and 1.5 μg mL -1 , respectively. The limit of quantification (LOQ) for sumatriptan succinate and Naproxen sodium was 6.3 and 4.8 μg mL -1 , respectively. This method was validated for accuracy, precision, linearity and robustness. sumatriptan succinate and Naproxen sodium were subjected to different ICH prescribed stress conditions of oxidative, acid, base, hydrolytic, thermal and photolytic degradation and the method was also found to be stability indicating.

31 citations


Journal ArticleDOI
TL;DR: In this paper, the authors describe the development and subsequent validation of a stability indicating reverse-phase high-performance liquid chromatography method for the simultaneous estimation of salbutamol sulphate and theophylline in tablet dosage forms.
Abstract: The study describes development and subsequent validation of a stability indicating reverse-phase high- performance liquid chromatography method for the simultaneous estimation of salbutamol sulphate and theophylline in tablet dosage forms. A reversed-phase phenomenax C-18 column (250 mm × 8 mm i.d., particle size 10 μm) column with mobile phase consisting of acetonitrile and phosphate buffer 65:35 (v/v) (pH 4.2 ± 0.02, adjusted with triethylamine) was used. The flow rate was 1.2 mL min -1 and effluents were monitored at 235 nm. The retention times (t R ) of salbutamol sulphate and theophylline were found to be 5.33 min and 13.36 min, respectively. The method was validated in terms of linearity, range, specificity, accuracy, precision, limit of detection (LOD) and limit of quantitation (LOQ). The linearity for both the drugs was found in the range of 2-64 μg mL -1 . The % recoveries of salbutamol sulphate and theophylline were found to be 99.41 and 101.11, respectively. The utility of the procedure is verified by its application to marketed formulations that were subjected to accelerated degradation studies. The method distinctly separated the drug and degradation products even in actual samples. The products formed in marketed tablet dosage forms are similar to those formed during stress studies.

31 citations


Journal ArticleDOI
TL;DR: In this article, a TLC-densitometry method was developed for simultaneous determination of tamsulosin hydrochloride and finasteride in tablet dosage form.
Abstract: Summary A new simple, precise, accurate and selective TLC-densitometry method has been developed for simultaneous determination of tamsulosin hydrochloride and finasteride in tablet dosage form. Chromatographic separation was performed on aluminum plate precoated with silica gel 60 F 254 using toluene: n-propanol: triethylamine (3.0:1.5:0.2 v/v) as mobile phase. Detection was carried out densitometrically at 260 nm. The RF value of tamsulosin hydrochloride and finasteride were 0.32 and 0.54, respectively. The reliability of the method was assessed by evaluation of linearity which was found to be 200 – 1200 ng/spot for tamsulosin hydrochloride and 1000 - 6000 ng/spot for finasteride. Accuracy of the method was accessed by percentage recovery and found to be 99.77 ± 0.71 % for tamsulosin hydrochloride and 99.75 ± 0.86 % for finasteride. The method can be used for routine analysis of tamsulosin hydrochloride and finasteride in tablet dosage form.

21 citations


Journal ArticleDOI
TL;DR: The results show that remarkable global changes in protein abundance occurred in the B. subtilis proteome as a result of CaCl2 or chelator 58 treatments compared to control cells, and suggest an important role of Ca2+ ions in B. subtitle cells.
Abstract: While the role of calcium binding proteins (CaBPs) in cell signaling pathways and homeostasis is well established in eukaryotic cells, the physiological function of CaBPs in prokaryotes is unknown. Although several CaBPs have been identified and sequences predicted in a variety of prokaryotic genomes, biochemical and functional characterization is lacking. We hypothesize that CaBPs play an important role in Ca2+ homeostasis and that Ca2+ ions regulate several processes in bacterial cells. The purpose of this work was to study the effects of Ca2+ in the B. subtilis proteome, to identify CaBPs altered (increased or decreased) by the addition of Ca2+ -chelators (EGTA, BAPTA) or CaCl2, and to examine Ca2+ homeostasis in B. subtilis cells utilizing various analytical techniques. 45Ca-autoradiography and antibody-crossreactivity were used to detect CaBPs. These proteins were identified by LC-MS/MS. Intracellular calcium levels [Ca2+]i were measured using the photoprotein aequorin. Our results show that remarkable global changes in protein abundance occurred in the B. subtilis proteome as a result of CaCl2 or chelator 58 treatments compared to control cells. Six proteins appeared to be modulated by high levels of extracellular Ca2+. These proteins were increased after Ca2+ -chelator treatments and reduced upon Ca2+ addition. Moreover, these proteins bound radioactive 45Ca2+, and showed a shift in molecular weight in the presence of Ca2+ /EGTA. B. subtilis cells thightly regulate cytosolic Ca2+ levels. Taken together, these results suggest an important role of Ca2+ ions in B. subtilis.

19 citations


Journal ArticleDOI
TL;DR: This review gives information regarding various stages involved in development and validation of analytical methods like LC, HPLC, MS.
Abstract: Analytical methods development and validation play important roles in the discovery, development, and manufacture of pharmaceuticals. The official test methods that result from these processes are used by quality control laboratories to ensure the identity, purity, potency, and performance of drug products. This review gives information regarding various stages involved in development and validation of analytical methods like LC, HPLC, MS.

18 citations


Journal ArticleDOI
TL;DR: A rapid and sensitive RP-HPLC method with UV detection and UV spectrophotometric method for routine pharmaceutical quality control of aceclofenac, paracetamol and tizanidine in tablets formulation was developed as mentioned in this paper.
Abstract: A rapid and sensitive RP-HPLC method with UV detection and UV spectrophotometric method for routine Pharmaceutical quality control of aceclofenac, paracetamol and tizanidine in tablets formulation was developed. Chromatography was performed by using Phenomenex-Luna C 18 (250 x 4.6 mm, 5μ) with a mobile phase containing Methanol: Water (90:10v/v), flow rate 1mL/min, wavelength at 256 nm. Linearity observed over the concentration range between 5-30μg/mL, 10-60μg/mL and 2-12μg/mL for aceclofenac, paracetamol and tizanidine respectively. The UV spectrophotometric method was performed at 277, 248 and 323 nm by derivative spectrophotometry with simultaneous equation method. The entire three drugs obey Beer’s law in the concentration range between 5-30 μg/mL, 2-16 μg/mL and 2-30 μg/mL respectively. The proposed methods were simple, rapid, precise, accurate and sensitive and can be used for routine quality control in pharmaceuticals.

15 citations


Journal ArticleDOI
TL;DR: This review examines the most commonly used techniques for typing and/or characterizing bacteria for epidemiological purposes and includes a historical perspective to help explain why certain techniques may be preferred over others, as well as a prediction of the future directions that epidemiologists may go in applying molecular technologies for their work.
Abstract: The evolution of molecular technologies has had a major impact on many fields of research, including epidemiology. At the core of this branch of epidemiology is the need for high specificity typing of disease agents: to confirm trace back of disease to origin, to monitor the spread of disease causing strains, to study population dynamics of the disease strain, to discern endemic/enzootic from epidemic/epizootic infections, to detect the presence of multiple strain (s) in the population and/or individual, to identify modes of transmission of the disease agent from host to host, and to address other epidemiological questions or issues. Molecular subtyping has been generally found to be better than most traditional phenotypic subtyping methods because it is usually more discriminating and less influenced by the organisms’ responses to environmental cues. A large number of molecular techniques have been adapted for application to epidemiological issues, and different techniques are needed for different aspects of investigation. This review examines the most commonly used techniques for typing and/or characterizing bacteria for epidemiological purposes. It includes a historical perspective to help explain why certain techniques may be preferred over others, as well as a prediction of the future directions that epidemiologists may go in applying molecular technologies for their work.

11 citations


Journal ArticleDOI
TL;DR: In this article, a simple, selective, precise densitometric HPTLC method for analysis of 6-gingerol both in chloroform extract of Zingiber officinalis and in laxiwin formulation has been developed and validated.
Abstract: A simple, selective, precise densitometric HPTLC method for analysis of 6-Gingerol both in chloroform extract of Zingiber officinalis and in laxiwin formulation has been developed and validated. The method employed HPTLC aluminium plates precoated with silica gel 60F-254 as the stationary phase. The phytoconstituent, 6-Gingerol was separated with mobile phase toluene: ethyl acetate (7:3 %, v/v). A compact band was obtained for 6-gingerol at R F 0.33±0.02. Densitometric analysis of 6-gingerol was carried out in the absorbance mode at 254 nm. The linear regression analysis data for the calibration plots showed good linear relationship with r 2 = 0.9981 with respect to peak area in the concentration range 150–900 ng per spot. The mean value of slope and intercept were 4.103 and 398.6 with respect to peak area. The method was validated for precision, recovery and robustness. The limits of detection and quantitation were 24.13 and 63.83 ng per spot, respectively. This novel Statistical analysis proves that the method is repeatable and selective for the analysis of 6-Gingerol in extract and in pharmaceutical formulations.

10 citations


Journal ArticleDOI
TL;DR: In this article, a simple sensitive and rapid method was developed for quantification of tacrolimus in rat whole blood utilizing triple Quadrupole LC/MS, which had a total chromatographic run time of 2.5 min; and linear calibration curves over the concentration range of 20.931 -1000.
Abstract: A simple, sensitive and rapid method was developed for quantitation of tacrolimus in rat whole blood utilizing triple Quadrupole LC/MS. An aliquot of 0.1 mL of whole blood sample was extracted with t-butyl methyl ether using a Heidolph vortexer. The chromatographic separation was performed by using chromolith fast gradient HPLC RP 18e (2mmX50mm.i.d) column with a mobile phase of 90% methanol and 10% 2mM ammonium acetate buffer followed by MS/MS detection. The analyte was quantitated in negative ionization mode. Multiple reaction monitoring (MRM) using the transition m/z 802.4→560.2 and m/z 808.4→548.6 was performed to quantify tacrolimus with IS (pimecrolimus), respectively. The method had a total chromatographic run time of 2.5 min; and linear calibration curves over the concentration range of 20.931 -1000.703 ng/mL.The lower limit of quantification (LLOQ) was 20.931 ng/mL. Use of sodium citrate (3.85%) as an anticoagulant in rat whole blood and samples were stable for at least the time required to assay the number of samples that could be placed in the auto sampler which is maintaining temperature of 10°C. The between and within batch precision and accuracy of the method were determined by using 6 quality control samples. The highest %CV 478.908 ng/mL (8.01% within run & 3.07 between run), with other %CV<5%. The recovery ranged 23.92% for tacrolimus over range of 50.285 to 798.179 ng/mL and was 18.52% for pimecrolimus respectively. The validated method was successfully applied to the quantification of tacrolimus concentration in rat whole blood.

Journal ArticleDOI
TL;DR: Stability-indicating Liquid Chromatographic and bioassay methods were validated and employed in the fluconazole stability studies and showed to be an unstable drug, indicating that special care must be taken during the handling, storage and quality control of this therapeutic agent.
Abstract: Chemical degradation of drugs may result in altered therapeutic efficacy and even toxic effects. Therefore, understanding the factors that change the stability of pharmaceuticals and identifying ways to guarantee their stability are important. In this work stability-indicating Liquid Chromatographic (LC) and bioassay methods were validated and employed in the fluconazole stability studies. The correlation of sample results from both methods was evaluated. Fluconazole raw material stability was investigated in aqueous, acid (0.1 M HCl), alkaline (0.1 M NaOH) and oxidative (3% v/v H2O2) reflux for 6 hours, by LC method. Fluconazole capsules were exposed to UVC (254 nm, 66 and 180 days), climatic chamber (40°C, 75% RH, 90 days) and oven (60°C, 60 days), these samples were analyzed by LC and bioassay methods It was found that the drug is degraded (10% decrease) with arising of a possible degradation product in an oxidative medium and UVC exposure, in all the others conditions fluconazole remained chemically stable (higher than 98%) when analyzed by LC. However when the capsules stressed samples were evaluated through bioassay very low antifungal activity was found (about 30%). Fluconazole showed to be an unstable drug and it indicates that special care must be taken during the handling, storage and quality control using appropriated methods to analyze this therapeutic agent. This work suggests monitoring the fluconazole stability by bioassay and the stability-indicating LC methods.

Journal ArticleDOI
TL;DR: In this article, the retention time (tR) of N-butyl-N-(3-carboxypropyl) nitrosamine (BCPN) was found to be randomly damping.
Abstract: N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) has been widely used in rodents as an invaluable experimental tool for investigation of bladder cancer (BCA). The urinary level of its metabolite, N-butyl-N-(3-carboxypropyl) nitrosamine (BCPN) was reported to be a very reliable predicative parameter of BCA. However, in determination of the urinary BCPN we found the retention time (tR) of BCPN was randomly damping. The tR values of the authentic BCPN at 5, 10, 20, 50, and 100 ppm were 28.48, 27.59, 27.43, 28.00, and 28.32 min comparing with 28.23 min of the urinary BCPN in HPLC analysis, similarly, 17.30 min for the urinary and the 18.00 min for the authentic BCPN in GC/MS analysis. To interpret such a damping, we theoretically proposed that a certain transient skewed molecular interaction could occur during the chromatographic separation, which would cause a certain degree of fluctuation on the tR of target molecules. Conclusively, the retention time of a chemical is not a definite value as often considered in HPLC and GC/MS analyses. In reality it fluctuates depending mainly upon the interaction among a cluster of coexisting molecules, in particular, when operated at higher concentrations.

Journal ArticleDOI
TL;DR: A quick and sensitive reversed phase high performance liquid chromatography (HPLC) method has been developed in Indian surgical patients to determine the concentration of Propofol In human plasma as mentioned in this paper.
Abstract: A quick and sensitive reversed phase high performance liquid chromatography (HPLC) method has been developed in Indian surgical patients to determine the concentration of Propofol In human plasma. Propofol can be isolated from human plasma by adding 1ml precipitating solution which consists of acetonitrile and perchloric acid (67:33) mixture, which also contains dibutylpthalate (1mg/ml) as an internal standard. The sample is mixed for two minute on a vortexer. The plasma substance precipitated by acetonitrile and perchloric acid are further separated by centrifugation. The supernatant is directly injected into the HPLC system with the help of autosampler. The analysis was carried out using column 250 × 4.6 mm column packed with10-μm Spherisorb reversed phase octadecyl silane particles (C 18 ). The 500ml of mobile phase (67:33:0.04) consisted of 335ml of acetonitrile and 165ml of distilled water and 200μl of acetic acid maintaining the pH 4.0.The flow rate of the mobile phase was 1.5ml/ min. propofol was monitored by a UV detector at a 270nm wavelength. The limit of detection of propofol (in human plasma) was found to be 0.0001μg/ml while limit of quantification was found to be 0.001μg/ml for a 20ul injection volume.

Journal ArticleDOI
TL;DR: In this paper, a simple, sensitive, rapid and economic high performance thin layer chromatographic method was developed for determination of amlodipine besylate and olmesaratan medoxomil in human plasma by liquid liquid extraction using paracetamol as an internal standard.
Abstract: A simple, sensitive, rapid and economic high performance thin layer chromatographic method has been developed for determination of amlodipine besylate and olmesaratan medoxomil in human plasma by liquid liquid extraction using paracetamol as an internal standard. The plasma sample was extracted using mixture of methanol: acetonitrile (3.0: 0.1). A concentration range from 500-3000 ng/spot of amlodipine besylate and olmesaratan medoxomil was used for calibration curve respectively. The percent recovery of amlodipine besylate and olmesaratan medoxomil was found to be 90.41 and 90.64 % respectively. The mobile phase consists of chloroform: methanol (9:1, v/v). Densitometric analysis was carried out at wavelength 254 nm. The Rf values for amlodipine besylate, olmesaratan medoxomil and paracetamol was 0.50 ± 0.05, 0.26 ± 0.05 and 0.66 ± 0.05 respectively. The stability of amlodipine besylate and olmesaratan medoxomil in plasma was confirmed during three freeze-thaw cycles at ‘-20°C’, on bench during 24 h and post preparative during 48 h. The proposed method was validated statistically and by performing recovery study for determination of amlodipine besylate and olmesaratan medoxomil in human plasma by liquid liquid extraction.

Journal ArticleDOI
TL;DR: A highly senstive and rapid LC-MS/MS method has been developed and validated for the estimation of desvenlafaxine in rabbit plasma and the results met the acceptance crieteria.
Abstract: A highly senstive and rapid LC-MS/MS method has been developed and validated for the estimation of desvenlafaxine in rabbit plasma. The chromatographic separation was performed with 0.2% formic acid: methanol at flow rate of 0.4 mL/min on Symmetry Shield RP18 column with a total run time of 3.0 min. TheMS/MS ion transitions monitored were 263.90? 58.10 for desvenlafaxine and 281.30--> 86.10 for IS (metaprolol). Method validation and pre-clinical sample analysis were performed as per FDA guide lines and the results met the acceptance crieteria. The lower limit of quantification achieved was 0.5 ng/mL and the linearity was observed from 0.5 to 1500 ng/mL. This novel method has been applied to pharmacokinetic study of desvenlafaxine in rabbits.