Sidika Ertürk Toker

Determination of Rosuvastatin at Picogram Level in Serum by Fluorimetric Derivatization with 9-Anthryldiazomethane using HPLC

For the first time, a carboxyl group derivatization assay has been developed and validated for the determination of the cholesterol-lowering drug rosuvastatin in human serum at picogram level by high-performance liquid chromatography with fluorescence detection. The assay procedure involved a simple one-step liquid-liquid extraction of rosuvastatin with lovastatin as internal standard from serum with an ethyl acetate-methyl tertiary buthyl ether (1:1) mixture. After pre-column derivatization with 9-anthryldiazomethane at room temperature for one hour, the reaction mixture was injected onto a Phenomenex, Synergi C18 column (250 × 4.6 mm, 4 µ i.d.). The analytes were separated with a mobile phase composed of acetonitrile-water in gradient elution mode and detected at λ(em) = 410 nm, exciting at 366 nm. Calibration curves were constructed in concentration range of 0.01-20.0 ng/mL and limit of detection and limit of quantification values were found to be 0.68 and 2.30 pg/mL, respectively. To test suitability of the developed methods for clinic use, the pharmacokinetics of rosuvastatin were investigated after oral administration of a 20 mg rosuvastatin film tablet to a healthy volunteer and maximum plasma concentration, time to reach that concentration and elimination half life were found to be 17.5 ng/mL, 3.5 h and 18.09 h, respectively.

RP-HPLC Method for Determination of Oseltamivir Phosphate in Capsules and Spiked Plasma

A new, sensitive RP-HPLC method was developed for the determination of oseltamivir phosphate in capsules and plasma. The method was based on the reaction of the drug with 4-chloro-7-nitrobenzofurazan in borate buffer solution of pH 8.50. Isocratic chromatography was performed on a C18 column with acetonitrile–10 mM nitric acid (pH 3, 60 + 40, v/v) as the mobile phase with fluorescence detection (λex: 470 nm, λem: 541 nm). Mexiletine hydrochloride was used as an internal standard. Analytical parameters were evaluated. The calibration range was linear from 50.0–750.0 ng/ml. The mean percentage recovery in capsules and plasma were 99.95% and 95.42%, respectively.

Simultaneous determination of desloratadine and pseudoephedrine sulfate in tablets by high performance liquid chromatography and derivative spectrophotometry

Abstract Simple and accurate reversed phase liquid chromatographic and first derivative spectrophotometric methods have been developed for simultaneous determination of desloratadine and pseudoephedrine in combined dosage forms. The first method is based on the separation of both drugs by high performance liquid chromatography using a C18 column and 10 mm ortho-phosphoric acid: acetonitrile (77:23). The drugs are eluted at 1.0 ml/min flow rate and detected at 262 nm. Ondansetron was used as internal standard. The calibration graphs of desloratadine and pseudoephedrine were linear in the range of 1.0–10.0 μg/ml and 48–480 μg/ml, respectively. The detection limits of the method were found to be 0.14 μg/ml and 3.80 μg/ml for desloratadine and pseudoephedrine, respectively. The derivative spectrophotometric method depends on measuring the first derivative amplitudes at 280 and 244 nm for desloratadine and pseudoephedrine, respectively. The calibration graphs were linear in the range of 2.5–15.0 μg/ml for desloratadine and 120–720 μg/ml for pseudoephedrine. The detection limits were found at 0.38 μg/ml and 21.45 μg/ml for desloratadine and pseudoephedrine, respectively. The proposed methods were successfully applied to the determination of these drugs in commercially available tablets.

Development of sensitive spectrofluorimetric and spectrophotometric methods for the determination of duloxetine in capsule and spiked human plasma.

A new, sensitive and selective spectrofluorimetric method has been developed for the determination of duloxetine (DLX) in capsule and spiked human plasma. DLX, as a secondary amine compound, reacts with 7-chloro-4-nitrobenzofurazon (NBD-Cl), a highly sensitive fluorogenic and chromogenic reagent used in many investigations. The method is based on the reaction between the drug and NBD-Cl in borate buffer at pH 8.5 to yield a highly fluorescent derivative that is measured at 523 nm after excitation at 478 nm. The fluorescence intensity was directly proportional to the concentration over the range 50-250 ng/mL. The reaction product was also measured spectrophotometrically. The relation between the absorbance at 478 nm and the concentration is rectilinear over the range 1.0-12.0 µg/mL. The methods were successfully applied for the determination of this drug in pharmaceutical dosage form. The spectrofluorimetric method was also successfully applied to the determination of duloxetine in spiked human plasma. The suggested procedures could be used for the determination of DLX in pure form, capsules and human plasma being sensitive, simple and selective.

Expression of human cytochrome p450 3A4 gene in Schizosaccharomyces pombe

Heterologous expression systems can be utilized to great advantage in the study of cytochrome P450 enzymes. P450 3A4 is one of the major forms of cytochrome P450 found in liver. It is also involved in the metabolism of numerous widely used drugs and xenobiotics. In the present study human liver cytochrome P450 3A4 gene was transferred into the fission yeast Schizosaccharomyces pombe via two different S. pombe expression vectors carrying thiamine repressible promoter — nmt1 (pREP42) and constitutive promoter — adh1 (pART1). Heterologously expressed cytochrome P450 3A4 was detected in the cells grown in minimal (EMM) or rich medium (YEL) containing 0.5% (w/v) glucose. A typical cytochrome P450 peak for 3A4 was observed at 448 nm in microsomal fraction. The presence of heterologous expression of 3A4 form was also determined by SDS-PAGE and it molecular mass was identified as 52 kDa. The enzyme activity was confirmed by HPLC analysis, using testosterone as substrate.

Spectrophotometric Determination of Antidepressant Drug Duloxetine in Pharmaceutical Preparations using π-Acceptors

In this study, two simple and accurate spectrophotometric methods were presented for the determination of duloxetine hydrochloride (DLX) in pharmaceutical preparations. The methods were based on the reaction of DLX as n-electron donor with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) and 7,7,8,8-tetracyanoquinodimethane (TCNQ) as π-acceptors to give highly colored complex species. The colored products were quantitated spectro-photometrically at 477 and 841 for DDQ, TCNQ, respectively. All variables were studied in order to optimize the reaction conditions. Beers law was obeyed in the concentration ranges 10.0-50.0 and 15-60 μg mL−1 for DDQ and TCNQ method, respectively. The proposed methods have been successfully applied to the pharmaceutical analysis without any interference from excipients. The suggested procedures could be used for the determination of DLX in pharmaceutical preparations being sensitive, simple and selective.