Quantitative structure-retention relationships of azole antifungal agents in reversed-phase high performance liquid chromatography.

Artificial neural network (ANN) is a learning system based on a computational technique which can simulate the neurological processing ability of the human brain. It was employed for building of the quantitative structure-retention relationships (QSRRs) model of antifungal agents-imidazoles or triazoles by structure. Computed molecular descriptors together with the percentage of acetonitrile in mobile phase (v/v) and buffer pH, being the most influential HPLC factors, were used as network inputs, giving the retention factor as model output. The multilayer perceptron network with a 9-5-1 topology was trained by using the back propagation algorithm. Good correlation between experimentally obtained data and ones predicted by using QSRR-ANN on previously unseen data sets indicates good predictive ability of the model.

Chemometrically assisted development and validation of LC-UV and LC-MS methods for simultaneous determination of torasemide and its impurities.

Complete evaluation of chromatographic behavior and establishment of optimal experimental conditions for determination of torasemide and its four impurities are determined by experimental design. Fractional factorial and 3(n) full factorial design were employed for efficient and rapid optimization of liquid chromatography-ultraviolet and liquid chromatography-mass spectrometry (LC-MS) methods. Separation is achieved on a Zorbax SB C(18) analytical column (250 x 4.6 mm, 5 µm) with mobile phase consisting of acetonitrile and 10 mM ammonium formate (pH 2.5 with formic acid) in gradient mode. The flow rate is 1 mL min(-1), the temperature of the column is 25 °C and UV detection is performed at 290 nm. The efficiency of ionization in electrospray ionization is higher than in atmospheric pressure chemical ionization mode; therefore, it is further used for analysis of torasemide and its impurities. Both methods meet all validation criteria. The calibration curves show high linearity with the coefficients of correlation (r) greater than 0.9982. The obtained recovery values (95.78-104.92%) and relative standard deviation values (0.12-5.56%) indicate good accuracy and precision. Lower limit of detection (LOD) and limit of quantitation (LOQ) values are obtained with the LC-MS method, indicating higher sensitivity of the proposed method.

Development and validation of reversed phase high performance liquid chromatographic method for determination of moxonidine in the presence of its impurities.

A simple, rapid, isocratic reversed-phase high-performance liquid chromatographic method was developed and validated for the analysis of moxonidine and its impurities in tablet formulations. The chromatographic separation was achieved on a Symmetry shield C18 column (250 mm × 4.6 mm, 5 µm) by employing a mobile phase consisting of methanol-potassium phosphate buffer (0.05 M) mixture (15:85, v/v) (pH 3.5) at a flow rate of 1 ml min⁻¹; detection at 255 nm. Central composite design technique and response surface method were used to evaluate the effects of variations of selected factors (buffer pH value, column temperature, methanol content) in order to achieve the best isocratic separation within short analysis time (less than 10 min), as well as for robustness test considerations. The method fulfilled the validation criteria: specificity, linearity, accuracy, precision, limit of detection and limit of quantitation. The method was successfully applied for the analysis of commercial moxonidine tablets.

Determination of carbamazepine and its impurities iminostilbene and iminodibenzyl in solid dosage form by column high-performance liquid chromatography.

An accurate and precise RP-HPLC method was developed and validated for the determination of carbamazepine and its impurities iminostilbene and iminodibenzyl in a tablet formulation with fluphenazine as an internal standard. Buffer-methanol (50 + 50, v/v) was used as the mobile phase. During validation, specificity, linearity, precision, accuracy, LOD, LOQ, and robustness of the method were tested. The method was proven to be specific against placebo interference. Linearity was evaluated over the concentration range of 100-500, 0.05-0.25, and 0.1-0.5 microg/mL, and the r values were 0.9994, 0.9997, and 0.9979 for carbamazepine, iminostilbene, and iminodibenzyl, respectively. Intraday precision of the method was good, and RSD was below 2% for all analytes. The accuracy of the method ranged from 100.69 to 102.10, 99.76 to 102.66, and 99.26 to 100.08% for carbamazepine, iminostilbene, and iminodibenzyl, respectively. LOD was 0.0125, 0.025, and 0.05 microg/mL and LOQ was 0.05, 0.05, and 0.1 microg/mL for carbamazepine, iminostilbene, and iminodibenzyl, respectiviely. Robustness of the method was proven by using a chemometric approach. The method was successfully applied to the analysis of commercially available carbamazepine tablets and showed good repeatability, with RSD below 2%.

Study of forced degradation behavior of eletriptan hydrobromide by LC and LC-MS and development of stability-indicating method.

The objective of the present study was to report the stability profile of novel antimigrain drug Eletriptan hydrobromide based on the information obtained from forced degradation studies. The drug was subjected to acid (0.1-1 mol L(-1) HCl), neutral and base (0.1-1 mol L(-1) NaOH) hydrolysis and to oxidative decomposition (3-15% (v/v) H(2)O(2)). Photolysis and thermo degradation at 75 degrees C were carried out in methanol solution and in solid state with both Eletriptan hydrobromide bulk drug and the tablet formulation. The products formed under different stress conditions were investigated by LC and LC-MS. The experimental conditions for LC were chosen by employing experimental design and multicriteria decision making methodology. These powerful tools enabled the accomplishment of satisfactory resolution with the shortest possible analysis time. Analytes were separated on a C(18) column (XTerra, 150 mm x 3.9 mm, 5 microm) with the mobile phase composed of methanol-water solution of TEA (pH 6.52, 1%, v/v) (30:70, v/v) pumped at 1 mL min(-1) flow rate. The column temperature was set at 50 degrees C and the detection at 225 nm using DAD detector. The LC method was suitably modified for LC-MS analysis which was further used to characterize the arisen degradation products. The possible degradation pathway was outlined based on the results. The drug appeared to be instable towards every stress condition but oxidation. The stability was not jeopardized even under more exaggerated conditions such as increased temperature of the solutions to 75 degrees C, increased strength of acid/alkali solutions and prolonged testing period. Validation of the LC-DAD method was carried out in accordance with ICH guideline. The method met all required criteria and was applied when testing the commercially available tablets.

Multicriteria optimization methodology in development of HPLC separation of mycophenolic acid and mycophenolic acid glucuronide in human urine and plasma.

Multicriteria optimization methodology was applied for development of isocratic reversed-phased HPLC method for simultaneous determination of mycophenolic acid (MPA) and mycophenolic acid glucuronide (MPAG) in human urine and plasma. In the first stage of method development, pH value of the water phase, percentage of acetonitrile, temperature of the column and flow rate of the mobile phase were investigated using fractional factorial design. Afterwards, the optimal conditions were found employing central composite design and Derringer's desirability function. Two goals were considered, the retention factor of the MPAG to be in the range, between 0.8 and 1.118 which allowed well separation of MPAG from the urine and plasma peaks, and the shortest possible total analysis run time. Then, the obtained sigmoid functions were used to transform the optimization criteria into the desirability values. The satisfactory chromatographic conditions were obtained with mobile phase consisted of acetonitrile-phosphate buffer (pH 2.4; 0.04 M KH(2)PO(4)) (28:72, v/v). The separation was performed on C(18) Chromolith column (100 mm x 4.6 mm) with flow rate of 5 mL/min, the temperature of the column was 25 degrees C and the chosen wavelength for simultaneous determination of MPA and MPAG was 215 nm. The MPAG eluted at 0.552 min and the duration of run was 3.092 min. Afterwards, the method was subjected to validation. Linearity was observed over the concentration range of 1-50 microg/mL for MPA and 1-500 microg/mL for MPAG in urine and 1-60 microg/mL for MPA and 1-70 microg/mL for MPAG in plasma matrix. The method showed intra-day and inter-day precision with relative standard deviation lower then 5% and accuracy as recovery (%) between 100+/-5%.

Application of experimental design in optimization of solid phase extraction of mycophenolic acid and mycophenolic acid glucuronide from human urine and plasma and SPE-RP-HPLC method validation.

The aim of this study was to develop and optimize a solid phase extraction (SPE) procedure for purification of mycophenolic acid (MPA) and its metabolite mycophenolic acid glucuronide (MPAG) in biological samples. During optimization process chemometric approach was applied. First, in screening experiments fractional factorial design (FFD) was used for selecting the variables which affected the extraction procedure. The ionic strength of the phosphate buffer in the washing step and the percentage of acetonitrile in the elution step were statistically significant for the recovery of MPAG while the percentage of acetonitrile and pH of the washing solution were statistically significant for that of MPA. Afterwards, the significant variables were optimized using central composite design (CCD). The developed SPE method included phosphate buffer (pH 2.4; 0.056 M) in the washing step, and the mixture of acetonitrile and phosphate buffer of which pH was adjusted to 2.4 (70:30, v/v) in the elution step. The investigation was applied to both urine and plasma and the nature of biological matrix appeared to be of no importance. The extraction from both matrixes showed good repeatability with relative standard deviations up to 6% for MPAG and 8% for MPA, and recovery around 100% for both substances. Furthermore, new SPE-RP-HPLC method for determination of MPA and MPAG in both humane urine and plasma has been validated. The great advantage of this method is the chromatographic run of only 3 min.

Investigation of chromatographic conditions for the separation of cefuroxime axetil and its geometric isomer.

The optimal chromatographic conditions for the separation of the syn- and anti-geometric isomers of cefuroxime axetil applying RP-HPLC and micellar liquid chromatography (MLC) methods were investigated. The possibility to separate diastereoisomers of syn- and anti-cefuroxime axetil was observed. Investigations were performed using three columns, two classical silicas and one with hybrid particle technology. Three aqueous-organic and one micellar mobile phases were used. The best results were achieved using micellar mobile phase. Optimization study was performed using different micellar mobile phases. MLC method is sensitive and applicable in purity and stability testing.