Rapid Ultra-performance Liquid Chromatographic Method Research Articles

STABILITY INDICATING ULTRA PERFORMANCE LIQUID CHROMATOGRAPHIC METHOD FOR SIMULTANEOUS DETERMINATION OF PHENYLEPHRINE HYDROCHLORIDE, CHLORPHENIRAMINE MALEATE, PARACETAMOL, GUAIPHENESIN AND BROMHEXINE HYDROCHLORIDE IN BULK AND PHARMACEUTICAL FORMULATION

Objective: The objective of the present study is to develop simple, rapid, sensitive, accurate and economic stability-indicating ultra-performance liquid chromatographic (UPLC) method for the simultaneous quantification of phenylephrine hydrochloride, chlorpheniramine maleate, paracetamol, guaiphenesin and bromhexine hydrochloride in bulk and tablet dosage form.
 Methods: The separation of drugs in the chromatographic column was accomplished on Hibar C18 (100 mm x 2.1 mm, 1.6 µm) column at a detection wavelength of 220 nm. The mobile phase was a combination of sodium phosphate monobasic monohydrate buffer (pH was adjusted to 3.5 with orthophosphoric acid) and acetonitrile in the ratio of 70:30 % v/v which was pumped at a flow rate of 0.3 ml/min. The column temperature was maintained at 30 °C and the injection volume was 0.3 µl. Forced degradation studies of drugs were carried out using acid, base, peroxide, light and heat.
 Results: All the five drugs have been eluted within 3 min. The retention times were found to be 0.834 min, 1.199 min, 1.600 min, 1.979 min and 2.525 min for phenylephrine, chlorpheniramine maleate, paracetamol, guaiphenesin and bromhexine respectively. The correlation coefficient (r2) was found to be 0.999 for all the drugs. The recovery levels were found to be in the range of 99.17 % to 100.69 %. RSD values of drugs were found to be below 2 %. The results of limit of detection and quantitation specified the sensitivity of the developed method. Significant degradation of drugs as a result of stress studies was found in acid, base and peroxide, but they were slightly degraded in photolytic and thermal conditions. The method has effectively resolved the degraded products. All the validation parameters were found to be within the limits according to International Conference on Harmonization (ICH) guidelines.
 Conclusion: A simple and rapid UPLC method was established for the determination of five drugs. Hence, the proposed method can be employed for the quality control of specified drugs in bulk and pharmaceutical formulation even in the presence of degradation products.

Development and validation of a simple and rapid UPLC method for the in-vitro estimation of (-)-epigallocatechin-3-gallate in lipid-based formulations

(-)-Epigallocatechin gallate (EGCG) is a catechin found in green tea that has potential health benefits, such as anti-oxidant, anti-carcinogenic and anti-inflammatory effects. A rapid and sensitive Ultra-Performance Liquid Chromatographic (UPLC) method was developed and validated for the estimation of (-)-epigallocatechin-3-gallate in lipid-based formulation. The UPLC method was conducted on C18 analytical column (50 mm × 2.1 mm, 1.8 μm particle size). The mobile phase consisted of a mixture of acetic acid (1%, v:v; pH = 3), acetonitrile and water at volume ratio of 13:15:72 delivered at a flow rate of 0.5 mL/min. The diode array detector (DAD) acquisition wavelength was set at wavelengths 210 and 280 nm. Caffeine was used as internal standard. The tested validation parameters, i.e., selectivity, linearity, accuracy, precision, and sensitivity (Limit of detection and limit of quantification) were determined at both wavelengths. Results revealed that caffeine and EGCG peaks were eluted at retention times of 0.55 and 0.85 minutes, respectively. The calibration curve was linear over the concentration range of 10-60 μg/mL, with coefficients of determination (r2) of 0.9993 and 0.9998 nm at 210 and 280 nm, respectively. All the validation parameters were found within the acceptable range. The proposed method was successfully applied for the quantitation of EGCG in lipid-based formulation and statistical analysis with a reported method showed no significant difference at p < 0.05. Therefore, the proposed analytical method for EGCG can be considered as a rapid, selective and accurate analytical method that can be used for the quantitative analysis of EGCG.

Quantification of seven β-lactam antibiotics and two β-lactamase inhibitors in human plasma using a validated UPLC-MS/MS method

There is an increasing interest in monitoring plasma concentrations of β-lactam antibiotics. The objective of this work was to develop and validate a rapid ultra-performance liquid chromatographic method with tandem mass spectrometric detection (UPLC-MS/MS) for simultaneous quantification of amoxicillin, ampicillin, cefuroxime, cefazolin, ceftazidime, meropenem, piperacillin, clavulanic acid and tazobactam. Sample clean-up included protein precipitation with acetonitrile and back-extraction of acetonitrile with dichloromethane. Six deuterated β-lactam antibiotics were used as internal standards. Chromatographic separation was performed on a Waters ACQUITY UPLC system using a BEH C18 column (1.7μm, 100mm×2.1mm) applying a binary gradient elution of water and acetonitrile both containing 0.1% formic acid. The total run time was 5.5min. The developed method was validated in terms of precision, accuracy, linearity, matrix effect and recovery. The assay has now been successfully used to determine concentrations of amoxicillin/clavulanic acid, cefuroxime and meropenem in plasma samples from intensive care patients.

Determinação rápida de oseltamivir em cápsulas por cromatografia líquida de ultraeficiência com detector por arranjo de diodos

A simple and rapid ultra-performance liquid chromatographic method for determination of oseltamivir in capsules was developed and validated. The mobile phase consisted of 5 mmol/L triethylammonium buffer (pH 3.0) and acetonitrile (70:30, v/v). Separation was performed in a Hypersil Gold® column, with octylsilil as stationary phase (100 x 2.1 mm, p.d. 1.9 µm). Chromatography run time was 1.2 min. The method presented adequate specificity, linearity, precision, ruggedness and accuracy and was adequate for determination of oseltamivir in capsules.

Validated UPLC method for the fast and sensitive determination of steroid residues in support of cleaning validation in formulation area

An ultra performance liquid chromatographic (UPLC) method was developed for simultaneous determination of seven steroid (dienogest, finasteride, gestodene, levonorgestrel, estradiol, ethinylestradiol, and norethisterone acetate) active pharmaceutical ingredient (API) residues. A new, generic method is presented, with which it is possible to verify the cleaning process of a steroid producing equipment line used for the production of various pharmaceuticals. The UPLC method was validated using an UPLC™ BEH C18 column with a particle size of 1.7μm (50mm×2.1mm) and acetonitrile–water (48:52, v/v) as mobile phase at a flow rate of 0.55ml/min. Method development and method validation for cleaning control analysis are described. The rapid UPLC method is suitable for cleaning control assays within good manufacturing practices (GMP) of the pharmaceutical industry.

Microanalysis of β-Lactam Antibiotics and Vancomycin in Plasma for Pharmacokinetic Studies in Neonates

Rational dosing of antibiotics in neonates should be based on pharmacokinetic (PK) parameters assessed in specific populations. PK studies of neonates are hampered by the limited total plasma volume, which restricts the sample volume and sampling frequency. Available drug assay methods require large sample volumes and are labor-intensive or time-consuming. The objective of this study was to develop a rapid ultra-performance liquid chromatographic method with tandem mass spectrometry detection for simultaneous quantification of amoxicillin, meropenem, cefazolin, cefotaxime, deacetylcefotaxime, ceftriaxone, and vancomycin in 50 microl of plasma. Cleanup consisted of protein precipitation with cold acetonitrile (1:4) and solvent evaporation before reversed-phase chromatographic separation and detection using electrospray ionization tandem mass spectrometry. Standard curves were prepared over a large dynamic range with adequate limits of quantitation. Intra- and interrun accuracy and precision were within 100% +/- 15% and 15%, respectively, with acceptable matrix effects. Coefficients of variation for matrix effects and recovery were <10% over six batches of plasma. Stability in plasma and aqueous stocks was generally sufficient, but stability of meropenem and ceftriaxone in extracts could limit autosampler capacity. The instrument run time was approximately 3.50 min per sample. Method applicability was demonstrated with plasma samples from an extracorporeal membrane oxygenation-treated neonate. Different beta-lactam antibiotics can be added to this method with additional ion transitions. Using ultra-performance liquid chromatography mass spectrometry, this method allows simple and reliable quantification of multiple antibiotics in 50 microl of plasma for PK studies of neonates.