Aytekin Temizer

Optimization of mobile phase in the separation of β-blockers by HPLC

Abstract β-blockers are generally determined using high-performance liquid chromatography (HPLC). Previous HPLC separations of β-blockers have often required a mobile phase containing three components; acetonitrile or methanol to control the retention; buffer to control the ionic strength and pH of the mobile phase; ion-pairing reagent to provide adequate retention of β-blockers or organic amines as masking agent to reduce peak tailing. Due to the complexity of the mobile phases employed, development of these assays can be a laborious process. Additionally, alkyl sulphonates and organic amines dramatically reduces the life-time reduction of silica based C18 columns. The results of this study demonstrated that the addition of tested alkyl sulphonates and organic amines is not essential for an adequate separation of β-blockers. In this study, we developed a simple HPLC method for the simultaneous separation of model β-blockers, atenolol, practolol, metoprolol, oxprenolol and propranolol. Atenolol, practolol, metoprolol, oxprenolol and propranolol adequately separated with high peak symmetries using a mobile phase consisted of methanol/acetonitrile/phosphate buffer (10 mM, pH 3.0) (15:15:70, v/v/v). By altering only the fraction of methanol with respect to acetonitrile, method development becomes a more efficient separation. Furthermore, atenolol, practolol, metoprolol, oxprenolol and propranolol can be detected up to 0.25, 5, 10, 50 and 10 ng ml−1. In this publication, we present the simultaneous separation of β-blockers having a wide range of polarity. It is proposed that this new mobile phase, consisting only acetonitrile, methanol and phosphate buffer can be used for the analysis of the several β-blockers presently in doping control analysis as well as others.

Gas chromatography-mass spectrometric analysis of clenbuterol from urine

Clenbuterol which is mostly used as an anabolic agent. It is also used for treatment of asthma. Clenbuterol was analysed from urine by using gas chromatography-mass spectrometry. GC-MS parameters were determined. Timolol was used as an internal standard. Extraction and derivatisation procedure of clenbuterol from urine were developed. Clenbuterol was extracted by using diethylether/ter-butanol (4:1; v:v) and pH 12 K2CO3/KHCO3 (3:2; w:w) buffer. MSTFA/NH4I (1 ml/10 mg) mixture was used for derivatization of clenbuterol. Selected ions of clenbuterol-bis-TMS were m/z: 405, 337, 336, 335, 300, and 227. Extraction yield and minimum detection limit of clenbuterol from urine were identified. Extraction yield was 94.30% and minimum detection was found 0.02 ng ml(-1) urine. It has been concluded that the GC-MS method is sensitive, accurate, precise, and reproducible for analysing of clenbuterol from urine.

Determination of ceftriaxone in aqueous humour and serum samples by differential-pulse adsorptive stripping voltammetry

Differential-pulse adsorptive stripping voltammetry was used to determine ceftriaxone in serum and aqueous humour samples. The method involved extraction of the ceftriaxone from serum samples with an Amberlite XAD-2 column followed by elution with methanol. The recovery was 97.6% with a relative standard deviation of 3.3% at a ceftriaxone concentration of 90.9 microg 1(-1). Peak currents of ceftriaxone were measured with a hanging mercury drop electrode at -0.78 V versus an Ag-AgCl reference electrode in pH 3.0 Britton-Robinson buffer. The calibration graph was linear from 0.02 to 1300 microg 1(-1). The method was applied to cataract cases and ceftriaxone levels were measured in aqueous humour and serum samples from patients who had received 1 or 2 g of ceftriaxone intravenously. Aqueous humour was added to the polarographic cell directly. The amounts of ceftriaxone in the aqueous humour and serum samples with respect to time were measured. The pharmacokinetic profiles for 1 and 2 g were compared.

Determination of lead and cadmium in urine by differential-pulse anodic stripping voltammetry

Differential-pulse anodic stripping voltammetry was used to determine Cd and Pb in urine. A hanging mercury drop electrode was chosen as the working electrode. Cd and Pb were determined separately after a digestion procedure and an additional pre-electrolysis step was used prior to the determination of Cd. Two deposition times and two standard additions were used to determine the concentrations of the metal ions.

Determination of Ranitidine in a Biological Material by Using Differential Pulse Adsorptive Stripping Voltammetry

Abstract A method has been developed to determine Ranitidine in stomach tissue of patients treated with this drug by using differential pulse adsorptive stripping voltammetry. A hanging mercury drop electrode was chosen as the working electrode. Extraction of ranitidine from the stomach tissue has been accomplished by using liquid-liquid extraction with chloroform. A peak at -0.56 V vs Ag/AgCl at Britten-Robinson buffer pH 2.3 has been taken for calibration curve which has been found linear in between 5.26×10−10 M to 1.27×10−5 M. Its precision was estimated by 10 successive measurernents of 2×10−7 M ranitidine The relative standard deviation was 2.8 %. The percent recovery has been found as 81 %.

Determination of ceftriaxone in biological material by differential-pulse adsorptive stripping voltammetry

Differential-pulse adsorptive stripping voltammetry was used to determine sub-micromolar concentrations of ceftriaxone in plasma. A hanging mercury drop electrode was chosen as the working electrode. A simple clean-up procedure was developed in which ceftriaxone was extracted from blood plasma with the non-ionic resin Amberlite XAD-2 and eluted with methanol. The recovery from plasma was 97.6% using a 1.52 x 10(-4) M stock ceftriaxone solution. The method was applied to caesarean cases, and total ceftriaxone levels were measured in the maternal and umbilical cord blood. The amount of ceftriaxone transmitted to the baby on administration of the drug to the mother before the caesarean operation was found to be in the range 0.067-0.17%.

Determination of methotrexate in human blood plasma by adsorptive stripping voltammetry

The use of adsorptive stripping voltammetry to measure sub-micromolar concentrations of methotrexate in plasma has been investigated. A simple clean-up procedure has been developed in which methotrexate is extracted from blood plasma with Amberlite XAD-2, which is a non-ionic resin, and eluted with methanol. Recovery for plasma analysis was 80%.

In vitro effects of thyroxine on the mechanical properties of erythrocytes

In vitro effects of thyroxine on erythrocyte deformability and mechanical fragility were observed. Deformability of erythrocytes was improved in a dose dependent manner by thyroxine. Mechanical hemolysis was found to be lower if thyroxine was included in erythrocyte suspensions at concentrations close to the physiological levels (10(-9)M). These changes might be related to the alterations of intracellular calcium concentration, as in the erythrocyte suspensions containing 10(-9)M thyroxine, intracellular calcium concentration was found to be 30 times lower than the control suspensions which did not contain thyroxine. Thyroxine also reduced the mechanical hemolysis ratio in calcium loaded cells. These observations suggest that thyroxine might play some role in the regulation of the mechanical properties of erythrocytes which might be mediated via the effects on calcium metabolism.

Electroanalytical determination of vinca alkaloids used in cancer chemotherapy

A differential pulse polarographic method has been developed for determination of the antineoplastic agents vincristine and vinblastine at ng ml level, in biological fluids such as plasma and urine. The vincristine and vinblastine are extracted from urine with Amberlite XAD-2. Linear calibration plots are obtained for both over the concentration range 0.005-5 mug ml . The relative standard deviations found were 1.7% for analysis of the pure drugs, 7.3% for urine and 8.6% for plasma.

Determination of betaxolol in human aqueous humour by high-performance liquid chromatography with fluorescence detection.

A reversed-phase high-performance liquid chromatographic method is described for the determination of betaxolol in human aqueous humour. Betaxolol and the internal standard metoprolol were extracted with cyclohexane and separated on a reversed-phase column (Luna C(18), 250 x 4.6 mm, 5 microm) with a mobile phase containing acetonitrile-phosphate buffer (40:60, v/v) at a flow-rate of 0.8 ml/min. The column effluent was monitored with a fluorescence detector at 227 nm (excitation) and 301 nm (emission). The retention times for metoprolol and betaxolol were 3.55 and 5.63 min, respectively. The recovery from aqueous humour was found to be 71.6% for betaxolol at 1.25 microg/ml. The within-day and day-to-day accuracy values were in the range of 96.17-105.2% for betaxolol at 0.1, 4 and 12 microg/ml (n=6), within-day and day-to-day precision values were less than 10% for betaxolol at the concentrations given above. The detection limit corresponding to the signal-to-noise ratio of 3:1 was 15 ng/ml. The presented method was suitable for measuring betaxolol levels in human aqueous humour samples obtained from patients after topical administration.