Validated HPLC-DAD and Spectrophotometric Methods for Simultaneous Determination of Ternary Mixture of Analgin, Caffeine, and Ergotamine.

BACKGROUND: Amigrain®, the market formulation of the ternary mixture of analgin, caffeine, and ergotamine, is used for the symptomatic treatment of migraine. OBJECTIVE: The aim of the work is to develop and validate an high-performance liquid chromatography-diode array-fluorescence (HPLC-DAD) method and a novel spectrophotometric method for simultaneous determination of analgin, caffeine, and ergotamine in their pharmaceutical formulation. METHOD: The HPLC separation of the ternary mixture was carried out using an Inertsil-C8 column and a gradient elution of mobile phase composed of acetonitrile and ammonium format buffer (pH 4.2), the ultraviolet detection for analgin and caffeine was carried out at λ = 280, 254 nm, and fluorometric detection for ergotamine was carried out at λ exc =310 nm, λ emm =360 nm. The two spectrophotometric methods were double divisor ratio spectra derivative (DDRD) and ratio dual wavelength (RDW) methods. The first method was used for determination of ergotamine at 355 nm and caffeine at 268 nm by the third and first derivative. The second one was based on using amplitude difference for the determination of caffeine and analgin. RESULTS: HPLC and spectrophotometric methods were applied over the concentration ranges of 50-400, 25-200, and 0.5-10 µg/mL and 10-35, 2-30, and 10-70 µg/mL for analgin, caffeine, and ergotamine for the two methods. CONCLUSIONS: The proposed methods were successfully applied for the determination of the cited drugs in their pharmaceutical formulation, and the obtained results were statistically compared with those of the reported methods without any significant difference. HIGHLIGHTS: The developed HPLC-DAD method has a high sensitivity for ergotamine. The spectrophotometric methods offer novelty, green solvent usage, and economic cost.

Simple protein precipitation extraction technique followed by validated chromatographic method for linezolid analysis in real human plasma samples to study its pharmacokinetics.

Fast and sensitive HPLC method was developed, optimized and validated for quantification of linezolid (LNZ) in human plasma using guaifenesin as an internal standard (IS). Analyte and IS were extracted from plasma by simple protein precipitation extraction technique using methanol as the precipitating solvent. The pretreated samples were injected in a mobile phase formed of acetonitrile:water:methanol (20:70:10v/v/v) in an isocratic mode at a flow rate of 1.5mL/min with UV detection at 251nm. Separation was done using Aglient ODS C18. The method showed linearity in the range of 0.75-50µg/mL with correlation coefficients equals to 0.9991. Precision and accuracy were in conformity with the criteria normally accepted in bio-analytical method validation. The RSDs for intra- and inter-day assays were <3.56 and 4.63%, respectively. The intra- and inter-day accuracies were 94.67-98.28% and 91.25-96.18%, respectively. The mean absolute recoveries ranged from 92.56±1.78 to 95.24±2.84. According to stability results, LNZ was stable in human plasma during the storage and analysis. LNZ a pharmacokinetic behavior was studied by applying the proposed analytical method.

Therapeutic drug monitoring and clinical outcomes in epileptic Egyptian patients: a gene polymorphism perspective study.

This work was performed to explore the effect of polymorphism in multidrug resistant genes on plasma phenytoin levels and patient outcome to evaluate its involvement in drug resistance and toxicity, which is usually associated with antiepileptic drugs. Therefore, we genotyped the adenosine triphosphate-binding cassette subfamily B member 1 (ABCB1) in 100 patients suffering from partial or generalized tonic-clonic seizures and receiving phenytoin and 50 healthy control subjects. Steady state plasma phenytoin levels were also determined in the epileptic patients. Patients were evaluated after 3 and 6 months and were classified either as drug resistant patients or responsive patients. Results revealed 37 patients with drug responsive epilepsy and 63 patients with drug resistant epilepsy. Genotyping of our patients and control subjects revealed a genotype distribution of CC, CT, TT: 55.50%, 38.00%, 6.50% for drug resistant patients, CC, CT, TT: 13.50%, 46.00%, 40.50% for drug responsive patients, and CC, CT, TT: 24.00%, 48.00%, 28.00% for the control subjects. Patients with drug-resistant epilepsy were more likely to have the CC than the TT genotype compared with either responsive patients (P < 0.0001) or control subjects (P < 0.0001). The C polymorphism was over-represented among patients with drug-resistant epilepsy as compared with either those with drug-responsive epilepsy (P < 0.001) or control subjects (P < 0.001). Of the total 100 epileptic patients, 13 patients had their plasma phenytoin levels exceeding the maximum safe concentration. These 13 patients were more likely to have TT genotype than the CC genotype compared with the remainder of patients who had their plasma phenytoin levels at 20 microg/mL or less. Responsive patients showed no deviation from the control group regarding the genotype (P > 0.05) or allele frequency (P > 0.05). In conclusion, because most of the antiepileptic drugs are multidrug resistant gene substrates, the ABCB1 is thus an important candidate gene for potentially influencing the response to antiepileptic drugs. Our findings suggest that using genotype data may make it possible to safely reduce the time required to reach an effective dose. Therefore, it is a priority to assess the utility of dose adjustment on the basis of genotype for these medicines that are substrates for this gene.