A Simple Validated LC-UV Method for the Simultaneous Determination of Brimonidine and Brinzolamide in the Presence of Brinzolamide's Degradation Product (Major Metabolite) in Rabbit Aqueous Humor.

A sensitive, specific, reliable and low-cost LC-UV method has been developed and validated for simultaneous quantification of brimonidine tartrate (BM) and brinzolamide (BZ) in rabbit aqueous humor (AH) in the presence of N-desethyl-brinzolamide (NDBZ); BZ is a major degradation product, and it is also considered to be its major metabolite. Dorzolamide hydrochloride (DZ) was used as an internal standard (IS). The analytes were extracted from rabbit AH samples by a simple pre-treatment utilizing ZnSO4.7H2O as a deproteinizing agent. The analytes were separated on a cyanopropyl Waters column (4.6 × 200 mm, 5 µm) with an isocratic mobile phase consisting of 25 mM ammonium acetate pH 4.5 (adjusted with 85% phosphoric acid):methanol:acetonitrile (94:4.5:1.5, v/v) at a flow rate of 1.0 mL min-1. The detection was done at 254 nm. The lower limit of quantification in rabbit AH was 100.0 ng mL-1. The method was validated according to EMA guidelines. The method was confirmed to be accurate, precise and linear over a range of 100.0-1000.0 ng mL-1 for BM and BZ. The method developed herein is simple, safe, eco-friendly, rapid and accurately applied for the quantification of BM and BZ, along with the successful separation of NDBZ, which is considered as a potential irritant degradation product of BZ.

Utilizing experimental design and desirability function in optimizing RP-HPLC method for simultaneous determination of some skeletal muscle relaxants and analgesics.

An experimental design and response surface methodologies using Plackett-Burman and Box-Behnken designs were applied for selecting and optimizing the most appropriate parameters which significantly affect the separation and quantitative estimation of five skeletal muscle relaxants and four analgesic drugs (baclofen, methocarbamol, dantrolene sodium, orphenadrine citrate, cyclobenzaprine hydrochloride, ketoprofen, etoricoxib, ibuprofen, and mefenamic acid) with a relatively short duration of analysis in a single run. For the separation of the nine drugs, an INERTSIL ODS-V3-5 µm C18 column (250 × 4.6 mm I.D.) was used with the optimum mobile phase conditions (45.15 mM ammonium acetate buffer pH 5.56 adjusted with acetic acid, acetonitrile, and methanol in a ratio of 30.5:29.5:40, v/v/v with a flow rate of 1.5 mL/min) and UV-detection at 220 nm. The optimized method was successfully subjected to the validation steps as described in ICH guidelines for linearity, precision, accuracy, robustness, and sensitivity. The optimized and validated method was effectively applied to determine the content of the studied drugs in their pharmaceutical preparations and to expand its applicability to the counterfeit estimation of etoricoxib in different brands of tablet dosage forms.

Resolution of overlapping spectra using chemometric manipulations of UV-spectrophotometric data for the determination of Atenolol, Losartan, and Hydrochlorothiazide in pharmaceutical dosage form.

Simultaneous determination of atenolol (ATN), losartan potassium (LOS), and hydrochlorothiazide (HCZ) in presence of HCZ impurity B was conducted by chemometric approaches and radial basis function network (RBFN) using UV-spectrophotometry without preliminary separation. Three chemometric models namely, classical least-squares (CLS), principal component regression (PCR), and partial least-squares (PLS) along with RBFN were utilized using the ternary mixtures of the three drugs. The multivariate calibrations were obtained by measuring the zero-order absorbance of the mixtures from 250 to 270 nm at the interval of 0.2 nm. The models were built covering the concentration range of (4.0 to 20.0), (3.8 to 20.2), and (0.9 to 50.1) µg mL-1 for ATN, LOS, and HCZ, respectively. The regression coefficient was calculated between the actual and predicted concentrations of the 3 drugs using CLS, PCR, PLS and RBFN. The accuracy of the developed models was evaluated using the root mean square error of prediction (RMSEP) giving satisfactory results. The proposed methods were simple, accurate, precise and were applied efficiently for the quantitation of the three components in laboratory-prepared mixtures, and in dosage form showing good recovery values. In addition, the obtained results were compared statistically with each other using ANOVA test showing non-significant difference between them.

LC-MS/MS determination of atropine toxicity: Pre-analytical effect of blood collection tube and analytical matrix.

Atropine (ATR) intoxication is a recurrent case in emergency departments. The diagnosis is dependent on clinical evaluation and is supported by analytical assessment. The assay is limited by the rapid degradation/metabolism of ATR into TRP as well as the preanalytical factors impairing correct detection and diagnosis. In this study, an HPLC-MS/MS method was optimized for the simultaneous determination of ATR and TRP. The effect of analytical matrix and the impact of blood-collection tube type on the ATR analytical signal were investigated. Separation was achieved using water: 0.01% formic acid acidified methanol (40: 60, v/v) as a mobile phase and Inertsil® C18 column (5 µm; 4.6*150 mm) as a stationary phase. The retention-times were 2.6 and 6.5 min for ATR and TRP, respectively. A chromatographic shift (0.4 min) in ATR peak, but not TRP, was observed in biological samples from neat ones. The best analytical signal was observed when heparinized blood collection tubes were employed. The method was linear, accurate and precise in the ATR toxicity range enabling the detection of ATR intoxication down to a concentration of 0.1 ng/mL by applying a simple sample clean-up procedure. In conclusion, an HPLC-MS/MS method for the simultaneous determination of ATR and TRP is presented. The method highlights the chromatographic shift of ATR peak in biological samples that may induce false-negative detection and poses TRP as an alternative toxicological marker for ATR toxicity. Meanwhile the study recommends heparin tubes for blood-sample collection.

In Vivo Evaluation of the Pharmacokinetic Interaction between Levothyroxine and Amiodarone in Rat Plasma: Evaluation of Importance of Therapeutic Drug Monitoring during Co-Therapy.

Amiodarone-induced thyrotoxicosis (AIT) is a common condition in patients who are receiving amiodarone for cardiac arrhythmia. This risk is elevated in iodine-deficient regions. Levothyroxine is the standard treatment for patients with hypothyroidism. This investigation is concerned with the evaluation of the possible pharmacokinetic interaction between amiodarone and levothyroxine upon co-therapy in rats and to investigate the cause of thyrotoxicosis. A selective, sensitive and precise RP-HPLC method was developed for the simultaneous determination of levothyroxine and amiodarone in rat plasma. A stationary phase of C18 Xterra RP column and a mobile phase consisting of acetonitrile: acidified water with 0.1% trifluoracetic acid (pH = 4.8) with gradient elution were used. The experiment was conducted at ambient temperature with flow rate of 1.5 mL/min for the chromatographic separation and quantitation of the investigated drugs. Protein precipitation with methanol was applied for the analysis of the two drugs in rat plasma. The method was linear over concentration range of 5-200 µg/mL for both levothyroxine and amiodarone. The European Medicines Agency guideline was applied for the validation of the developed bioanalytical method. The method was successfully applied to in vivo pharmacokinetic study in which levothyroxine and amiodarone were quantified in plasma of rats after receiving an oral dose of levothyroxine and amiodarone. After the calculation of the pharmacokinetic parameters, a statistical analysis was performed to elucidate the existence of significant difference between test and control groups in rats. The combination of levothyroxine and amiodarone significantly decreased levothyroxine bioavailability in rats, making the therapeutic drug monitoring mandatory in patients receiving levothyroxine and amiodarone. In addition, the increased clearance of levothyroxine upon the co-administration with amiodarone may explain the reported hypothyroidism.

AGREE and ESA for Greenness Assessment of a Novel Validated RP-HPLC Method for Simultaneous Determination of Aspirin, Warfarin and Clopidogrel in Rat Plasma: Application to Pharmacokinetic Study of the Possible Interaction between the Three Drugs.

The green profile of the developed method is assessed and compared with previously reported methods. Percutaneous coronary intervention is a procedure where a catheter is utilized to place a stent in order to facilitate opening of the blood vessels in the heart. Triple antithrombotic therapy includes oral anticoagulation as warfarin and dual antiplatelet therapy (composed of aspirin and clopidogrel bisulfate). The aim of the current study was to evaluate the pharmacokinetic parameters of ASP, WAR and CLP and to investigate the possible interaction between the three drugs upon co-administration in rats. A selective and precise RP-HPLC method was developed for the simultaneous determination of ASP, WAR and CLP in rat plasma. Pharmacokinetic study was conducted in rats that received ASP, WAR and CLP as an application of the developed method. From the statistical evaluation of the pharmacokinetic parameters, it was observed that the co-administration of ASP, WAR and CLP significantly increased the ASP and CLP bioavailability in rats. A significant drug-drug interaction was confirmed in the current study. The elevated Cmax of ASP and CLP upon the co-administration of ASP, WAR and CLP may explain the reported bleeding.

Application of Box-Behnken Design and Response Surface Methodology for Selecting the Optimum RP-HPLC Conditions for the Simultaneous Determination of Paracetamol and Diclofenac Sodium Along With Three Skeletal Muscle Relaxants in Three Different Pharmaceutical Dosage Forms.

An isocratic HPLC method has been developed and validated for estimating paracetamol and diclofenac sodium simultaneously with three skeletal muscle relaxants, namely, methocarbamol, tizanidine hydrochloride and chlorzoxazone in their pure standard mixtures and in different multi-component dosage forms in a single chromatographic run. HPLC separation was achieved on a C18 Inertsil ODS-3V 5 µm column (250 × 4.6 mm) using a mobile phase mixture containing acetonitrile and 25 mM phosphate buffer (pH 7.4 adjusted with NaOH) in the proportion of (39.7:60.3, v/v) pumped at 1.2 mL/min flow rate with UV detection at 220 nm. An experimental design was used by applying Plackett-Burman design for screening the most critical predictors affecting the chromatographic separation and Box-Behnken design for optimizing the selected predictors and creating the response surface between the selected predictors and the interested responses. ICH recommendations were applied for validating the proposed method with regard to linearity, precision, accuracy, selectivity, limits of detection and quantitation, and robustness. There are many applications for the optimized method that can be applied for routine estimation of the cited drugs in laboratories of quality control and pharmaceutical industries to save money and time and to reduce material waste and effort.

The potential off-target neuroprotective effect of sister gliflozins suggests their repurposing despite not crossing the blood-brain barrier: From bioanalytical assay in rats into theory genesis.

Gliflozins are successfully marketed antidiabetic agents with a reported neuroprotective effect, and this study tests their blood-brain barrier crossing ability. Henceforward, a computational hypothesis interpreting their effects was reasonable after failure to cross into the brain. A chromatographic bioassay for canagliflozin, dapagliflozin, and empagliflozin was developed, validated, and applied to the rat's and rat's plasma and brain. HPLC method robustness was tested over two levels using Design of Experiment on MINITAB. It is the first method for gliflozins' detection in rats' brain tissue. The method was applied on 18 rats and six for each drug. Concentrations in plasma were determined but neither of them was detected in brain at the described chromatographic conditions. A computational study for the three drugs was endorsing two techniques. First, ligand-based target fishing reveals possible targets for gliflozins. They showed an ability to bind with human equilibrative nucleoside transporter 1, a regulator of adenosine extracellularly. Second, a docking study was carried out on this protein receptor. Results showed perfect alignment with a minimum of one hydrogen bond. Dapagliflozin achieved the lowest energy score with two hocking hydrogen bonds. This is proposing gliflozins ability to regulate equilibrative nucleoside transporter 1 receptors in peripheries, elevating the centrally acting neuroprotective adenosine.

Validated LC/MS/MS Method for the Determination of Rivastigmine in Human Plasma: Application to a Pharmacokinetic Study in Egyptian Volunteers to Determine the Effect of Gender and Body Mass Index.

The effect of gender and body mass index (BMI) on the pharmacokinetics of rivastigmine was studied in Egyptian human subjects using new bio-analytical validated LC/MS/MS method. In this study, Rivastigmine was estimated in human plasma using Escitalopram as an internal standard (IS). Rivastigmine and Escitalopram were extracted from human plasma samples by liquid-liquid extraction using diethyl ether (DEE)-dichloromethane (DCM) (70:30, v/v). Chromatographic separation was performed on a reversed phase C18 INERTSIL ODS column using 0.05% aqueous formic acid, acetonitrile in the ratio (50:50, v/v) as a mobile phase. Multiple reaction monitoring (MRM) was applied and operated by positive mode electrospray ionization. A significant difference between male and female Cmax (maximum plasma concentration) (P = 0.0205; CL = 95.4) was found using Mann-Whitney U test. Also, a moderate negative correlation was found between BMI and Tmax (time to peak plasma concentration) using spearman rho test. The calculated results confirm the difference of Rivastigmine pharmacokinetics between male and female subjects. Furthermore, it indicates that Rivastigmine dose adjustment may be necessary. The method was applied for the estimation of pharmacokinetic parameters in volunteers (n = 26, 17 male and 9 female) and the effects of gender and BMI were investigated.

A Turn-On-Type Fluorescence Resonance Energy Transfer Eco-friendly Method for Nitazoxanide Quantification in Pharmaceutical Dosage Form and Spiked Plasma: Evaluation of Greenness Profile Using Different Assessment Tools.

A brand-new class of anti-infective drugs that work against bacteria, viruses, and protozoan parasites is nitazoxanide and related thiazolides. Thiazolides have also been shown to cause cell cycle arrest and apoptotic cell death in cancer cells in recent years. In this study, an eco-friendly, spectrofluorimetric technique that is verified, easy, and sensitive has been proposed for quantifying nitazoxanide (NTZ), a broad-spectrum antiparasitic drug. When NTZ is reduced with zinc (Zn) powder in an acidic media, a highly fluorescent product is produced. To get the highest sensitivity, different experimental conditions impacting the response were examined and optimized. Following excitation at 299 nm, scanning of the fluorescent product was done at 440 nm. The intensity of the fluorescence was proportional to the drug concentration in the range of 0.1-0.6 µg/mL. The approach was validated according to International Conference on Harmonization (ICH) guidelines, and the outcome was satisfactory. The detection and quantitation limits were calculated to be 0.013 and 0.038 µg/mL, respectively. The suggested technique was successful in analyzing commercially available NTZ dosage forms. Furthermore, the proposed technique was used to assess NTZ levels in human plasma and it was bio-analytically validated according to European Medicines Agency (EMA) guidelines. The suggested method can be used in quality control laboratories as well as in pharmacokinetic studies. In order to picture the green profile of the developed method, four greenness assessment tools have been applied. National Environmental Methods Index (NEMI), analytical Eco-Scale Assessment (ESA), Green Analytical Procedure Index (GAPI) and Analytical Greenness metric (AGREE) are the relatively most widely used metrics. So, they were utilized to perform a detailed greenness comparison between the proposed method and some of the reported methods for the determination of NTZ. The developed method was found to be an excellent green method with the highest AGREE score.

Application of Box-Behnken experimental design and response surface methodology for selecting the optimum RP-HPLC conditions for the simultaneous determination of methocarbamol, indomethacin and betamethasone in their pharmaceutical dosage form.

An isocratic RP-HPLC method has been developed for the separation and determination of methocarbamol (MTL), indomethacin (IND), and betamethasone (BET) in combined dosage form using an Inertsil ODS-3v C18 (250 × 4.6 mm, 5 µm) column with UV- detection at 235 nm. Experimental design using Box-Behnken design (BBD) was applied to study the response surface during method optimization and to achieve a good separation with a minimum number of experimental runs. The three independent parameters were pH of buffer, % of acetonitrile and flow rate of the mobile phase while the peak resolution of IND from MTL and the peak resolution of BET from IND (R2) were taken as responses to obtain mathematical models. The composite desirability was employed to optimize a set of responses overall (peak resolutions). The predicted optimum assay conditions include a mobile phase composition of acetonitrile and phosphate buffer (pH 5.95) in a ratio of 79:21, v/v, pumped at a flow rate of 1.4 mL min-1. With this ideal condition, the optimized method was able to achieve baseline separation of the three drugs with good resolution and a total run time of less than 7 min. The linearity of MTL, IND, and BET was determined in the concentration ranges of 5-600 µg mL- 1, 5-300 µg mL- 1, and 5-300 µg mL- 1 and the regression coefficients were 0.9994, 0.9998, and 0.9998, respectively. The average percent recoveries for the accuracy were determined to be 100.41 ± 0.60%, 100.86 ± 0.86%, and 100.99 ± 0.65% for MTL, IND, and BET, respectively. The R.S.D.% of the intra-day precision was found to be less than 1%, while the R.S.D.% of the inter-day precision was found to be less than 2%. The RP-HPLC method was fully validated with regard to linearity, accuracy, precision, specificity, and robustness as per ICH recommendations. The proposed method has various applications in quality control and routine analysis of the investigated drugs in their pharmaceutical dosage forms and laboratory-prepared mixtures with the goal of reducing laboratory waste, analysis time, and effort.

Assessment of alteration in antiviral plasma concentration across dialysis days: computational and analytical study.

Aim: Protein-bound uremic toxins (PBUTs) may displace drugs from the plasma proteins and render them more liable to clearance. This study aims to investigate the possible interplay between PBUTs and directly acting antivirals (DAAs). Methods: PBUT plasma protein binding was compared to those of paritaprevir (PRT), ombitasivir (OMB) and ritonavir (RTV) in silico to assess the possible competitive displacement. The three drugs were LC-MS/MS determined in seven patients across dialysis and non-dialysis days and results were compared. Results & conclusion: Results showed that the PBUT exhibited a lower binding than DAA reducing the liability of their competitive displacement. This was echoed by an unaltered plasma concentration across dialysis days. Results may indicate that PBUT accumulation may have limited effect on disposition of DAA.

DoE Screening and Optimization of Liquid Chromatographic Determination of Nicotinic Acid and Six Statins: Application to Pharmaceutical Preparations and Counterfeit Detection.

An isocratic reversed-phase high performance liquid chromatographic method has been developed and validated to simultaneously determine nicotinic acid, pravastatin sodium, rosuvastatin calcium, atorvastatin calcium, pitavastatin calcium, lovastatin sodium and simvastatin sodium in focus on counterfeit drug detection. Thin-layer chromatography, nuclear magnetic resonance and mass spectrometry have been additionally performed to verify the identification of adulterants of counterfeit herbal medicines. Chromatographic separation was carried out on Inertsil® ODS-3 C18 (4.6 × 150 mm, 5 µm) with isocratic mobile phase elution containing a mixture of acetonitrile: methanol: 25 mM potassium dihydrogen phosphate buffer, pH 2.86 adjusted with 0.1 M o-phosphoric acid (48: 30: 22, v/v/v), at a flow rate of 1 mL/min and with UV detection at 238 nm. The design of experiment methodology, Plackett-Burman and Box-Behnken designs, was used to screen and optimize the mobile phase composition. The validation of the method was also carried out under the International Conference on Harmonization guidelines. The developed method was sensitive, accurate, simple, economical and highly robust, in addition to the comprehensiveness and novelty of this method for separating the seven drugs. The results were statistically compared with the reference methods used Student's t-test and variance ratio F-test at P < 0.05.

Bioanalytical LC-MS/MS method for simultaneous estimation of atorvastatin, its major active metabolites and ezetimibe.

Background: Hyperlipidemia is one of the most common chronic diseases encountered globally, and atorvastatin (ATV) is mainly metabolized into two major active metabolites. Methodology: Hence, we aimed to estimate ATV and ezetimibe (EZE) simultaneously in the presence of ATV major and active metabolites using a validated LC-MS/MS method. Conclusion: The proposed method was linear (r2 >0.99), accurate (92.02-109.94%) and precise (CV% ≤14) over the concentration range of 0.50-120 ng/ml, 0.20-48 ng/ml, 0.50-120 ng/ml and 0.20-48 ng/ml for ATV, EZE, 2-hydroxy ATV and 4-hydroxy ATV, respectively. The applied liquid-liquid extraction gave rise to reliable extraction recoveries of 84.91 ± 1.14%, 85.20 ± 1.62%, 85.46 ± 0.41% and 105.46 ± 2.35% for ATV, EZE, 2-hydroxy ATV and 4-hydroxy ATV, respectively.

Analysis of Metformin and Five Gliptins in Counterfeit Herbal Products: Designs of Experiment Screening and Optimization.

BACKGROUND: Drug counterfeiting is a rising problem due to difficulties with identifying counterfeit drugs and the lack of regulations and legislation in developing countries. OBJECTIVE: This study aims to develop a robust and economic reversed phase high performance liquid chromatography (LC) method for simultaneously determining metformin HCl, vildagliptin, saxagliptin, alogliptin benzoate, sitagliptin phosphate monohydrate, and linagliptin to target counterfeiting. METHODS: Plackett-Burman (PB) and Box-Behnken (BB) designs were used to screen and optimize the mobile phase composition. Chromatographic separation was carried out on an Inertsil® ODS-3 C18 column with isocratic elution mode and the mobile phase was a mixture of acetonitrile-methanol-ammonium formate buffer, pH 3.5 (25:10:65, v/v/v). This method was applied to analyze synthetic drugs in three traditional Chinese and Indian herbal medicines. To identify the adulterants, thin-layer chromatography (TLC), nuclear magnetic resonance (NMR), and mass spectrometry (MS) were used on counterfeit herbal medicines. RESULTS: The developed method is sensitive, simple, rapid, economical, accurate, and highly robust. Student's t-test and variance ratio (F-test at P < 0.05) were used to compare the results statistically with the reference methods. CONCLUSION: The study found that the analyzed herbal medicines were adulterated with metformin and the quantification of anti-diabetic counterfeits was therefore applied. HIGHLIGHTS: This study determined counterfeited anti-diabetic drugs in Indian and Chinese traditional herbal medicines(THMs). Design-of-experiment, PB, and BB designs were used. Method validation was also performed in accordance with the International Conference on Harmonization guidelines.

Analysis of Aspirin, Prasugrel and Clopidogrel in Counterfeit Pharmaceutical and Herbal Products: Plackett-Burman Screening and Box-Behnken Optimization.

An isocratic reversed-phase high-performance liquid chromatographic method has been developed and validated for the simultaneous determination of aspirin, prasugrel HCl and clopidogrel bisulfate in the presence of clopidogrel-related compound (impurity-A) in focus on counterfeit. This method was used to determine counterfeited antiplatelet drugs in two substandard Indian pharmaceutical products sold on the market in Yemen and two traditional herbal medicines sold on the market in China. Thin layer chromatography and mass spectrometry of counterfeit herbal medicines have additionally been carried out to verify the identification of adulterants. Chromatographic separation was performed on Inertsil ® ODS-3 C18 (4.6 × 250 mm, 5 µm) with isocratic mobile phase elution containing a mixture of acetonitrile: (25 mM) potassium dihydrogen phosphate buffer, pH 2.7 adjusted with 0.1 M o-phosphoric acid (79: 21, v/v), at a flow rate of 1 mL/min and UV detection at 220 nm. Designs of experiment methodology, Plackett-Burman and Box-Behnken designs were used for the screening and optimization of the mobile phase composition. The method validation was also performed in accordance with the International Council on Harmonization (ICH) guidelines. The method developed for routine analysis was found to be sensitive, simple, accurate and highly robust. The results were statistically compared to reference methods using Student's t-test and variance ratio F-test at P < 0.05.

Hydroxychloroquine Safety Outcome within Approved Therapeutic Protocol for COVID-19 Outpatients in Saudi Arabia.

BACKGROUND: Global healthcare is challenged following the COVID-19 pandemic, since late 2019. Multiple approaches have been performed to relieve the pressure and support existing healthcare. The Saudi Arabian Ministry of Health (MOH) launched an initiative to support the National Healthcare System. Since the 5th of June 2020, 238 outpatient fever clinics were established nationwide. This study aimed to assess the safety outcome and reported adverse events from hydroxychloroquine use among suspected COVID-19 patients. METHOD: A cross-sectional study included 2,733 patients subjected to MOH treatment protocol (hydroxychloroquine) and followed-up within 3-7 days after initiation. Data was collected through an electronic link and cross-checked with the national database (Health Electronic Surveillance Network, HESN) and reports from the MOH Morbidity and Mortality (M&M) Committee. RESULTS: 240 patients (8.8%) discontinued treatment because of side effects (4.1%) and for non-clinical reasons in the remaining (4.7%). Adverse effects were reported among (6.7%) of all studied participants, including mainly cardiovascular (2.5%, 0.15% with QTc prolongation), and gastrointestinal (2.4%). No Intensive Care Unit admission or death were reported among these patients. CONCLUSION: Our results show that hydroxychloroquine for COVID-19 patients in mild to moderate cases in an outpatient setting, within the protocol recommendation and inclusion/exclusion criteria, is safe, highly tolerable, and with minimum side effects.

Study of gender-related pharmacokinetics of ezogabine in Egyptian volunteers by a validated LC-MS/MS bioanalytical method.

Gender-based pharmacokinetics and/or pharmacodynamics differences can result in differences in treatment which can accordingly affect the drug safety and/or efficacy. A new validated bio-analytical LC-MS/MS method was developed for the estimation of ezogabine, a third-generation antiepileptic drug, in human plasma using oxcarbazepine as an internal standard (IS) and to study the gender effect on the pharmacokinetic parameters in Egyptian human subjects. Liquid-liquid extraction of plasma samples was performed with diethyl ether: dichloromethane. The separation was accomplished in an isocratic mode with a mobile phase of a mixture of 5 mM ammonium acetate: methanol: acetonitrile pumped on a reversed phase C18 INERTSIL ODS-3 (5 µm, 150 × 4.6 mm). Multiple reaction monitoring was applied and operated by positive mode electrospray ionization. Male and female Cmax (p = 0.0308; CL = 95) and t1/2 (p = 0.0301; CL = 95) were found to be significantly different using Mann-Whitney U test. These findings highlight the difference of ezogabine pharmacokinetics among populations. Further, gender-based ezogabine dose adjustment may be considered.

Bio-analytical methods for investigating the effect of age, body mass index and gender on the PK/PD ratio of antibiotics.

The effectiveness of antibiotics (ABs) is governed by achieving the adequate pharmacokinetic (PK)/pharmacodynamics (PD) ratio. In this study, fast LC-MS/MS methods were developed and validated for the bioanalysis of cefaclor (CFC), ciprofloxacin (CFC), roxithomycin (RXM) and clindamycin (CLD). Chromatographic separation was performed on a C18 Zorbax-Eclipse Plus (3.5 µm, 100 × 4.6 mm) using isocratic elution. Detection was performed by positive electrospray ionization. The methods were applied for the assessment of PK parameters in volunteers (n = 101, 64 male and 37 female) and the effects of age, body mass index (BMI) and gender were investigated. Good linearity (r2 ≥ 0.99), accuracy (>86%), precision (CV% ≤ 11) and extraction recovery (>83%) were observed for CFC, CFX, RXM and CLD. Application to PK studies revealed that age and BMI affected the Thalf and the AUC of RXM and CLD (p < 0.023). Gender difference affected the critical PK parameters of the four ABs (Thalf (U = 18; P = 0.036) of CFC, the Cmax of CFX (U = 30; P = 0.017), the Thalf (U = 23; P = 0.009) and AUC (U = 26; P = 0.008) of RXM and CLD), respectively. These results highlight the significance of age and BMI variations for RXM and CLD dosing. Furthermore, it indicates that the gender difference may be considered when adjusting the AB dose.

Validated Liquid Chromatographic Method for the Determination of (Canagliflozin, Dapagliflozin or Empagliflozin) and Metformin in the Presence of (1-Cyanoguanidine).

A simple and accurate liquid chromatographic method has been developed and validated for the determination of either canagliflozin, dapagliflozin propandiol monohydrate or empagliflozin and metformin in presence of metformin major degradation product;1-cyanoguanidine. The Liquid Chromatographic (LC) method was based on isocratic elution on Prontosil (Lichrosorb 100-5-NH2) column using a mobile phase consisting of NaH2PO4 buffer (10 mM, pH 2.8):acetonitrile (18.5:81.5, v/v), at a flow rate of 2 mL/min-1. Quantitation was achieved with UV detection at 225 nm. The validation of the method was assessed according to International Conference on Harmonization (ICH) guidelines. Linearity, accuracy and precision were satisfactory over the concentration ranges of 12.5-100, 3.75-30, 0.3075-2.46, and 0.3125-2.5 µg/mL for metformin HCl, canagliflozin, dapagliflozin propandiol monohydrate and empagliflozin, respectively. Limits of detection and quantitation were found to be 0.068, 0.135, 0.077 and 0.069 µg/mL and 0.206, 0.410, 0.233 and 0.210 µg/mL for metformin HCl, canagliflozin, dapagliflozin propandiol monohydrate and empagliflozin, respectively. The developed method is suitable for the quality control and routine analysis of the cited drugs separately or in combinations.