Niveen A. Mohamed

Voltammetric determination of azithromycin at the carbon paste electrode

Azithromycin (AZ) is the first member of a class of macrolide azalides antibiotics called azolides. A simple and selective square-wave voltammetric (SWV) method has been developed for the determination of azithromycin in pure form, in pharmaceutical preparation and in biological samples. Determination of azithromycin was accomplished with hand-make carbon paste electrode (CPE) in oxidative screen mode. The counter and reference electrodes were a Pt wire and a Ag/AgCl, respectively. Various parameters that can influence the peak signal (effect of buffer, ionic strength, accumulation time, pH and the composition of the paste) have been scrutinized. The best results were obtained in acetonitrile-aqueous 1M sodium acetate-acetic acid buffer (pH 4.6) containing 0.1M KCl (1:9; v/v) using a 15% paraffin oil CPE. The limits of detection and quantification of the pure drug are 0.463 and 1.544ppb (with the correlation coefficient, r=0.9785and the standard deviation, S.D.=0.1 (n=5), for the accumulation time of 60s), respectively. The method was successfully applied to the determination of the drug in urine and two forms of pharmaceutical formulations. Recoveries were 99.2-100.5% with S.D.=0.1-and 0.8% (n=5).

Binding analysis of nilvadipine to plasma lipoproteins by capillary electrophoresis-frontal analysis

Capillary electrophoresis coupled with frontal analysis (HPCE/FA) was applied to the ultramicro analysis of enantioselective binding of nilvadipine (NV), a calcium channel blocker, to plasma lipoproteins. The drug lipoprotein mixed solution was hydrodynamically introduced into a non-coated fused silica capillary for capillary electrophoresis. Since NV has no electric charge in the run buffer (pH 7.4), the unbound NV moved towards the cathodic end by electroosmotic flow, which was faster than the electrophoretic migrations of negatively charged lipoproteins and the bound NV. Once unbound NV migrated apart from lipoprotein, and bound NV was quickly released from the protein to maintain the binding equilibrium. Thus, NV migrated as a zone with a plateau region. The concentration of NV in this plateau region appearing on the electrophorogram was the same as the unbound NV concentration in the initial sample solution. It was found that the binding of NV to high-density lipoprotein (HDL), low-density lipoprotein (LDL) and oxidized LDL was non-specific and not enantioselective. Partition-like binding to the lipid part of these lipoproteins seemed to occur dominantly. The total binding affinities of NV to LDL were about seven times stronger than those to HDL, and the oxidation of LDL enhanced the binding affinity significantly.

Spectrofluorimetric method for determination of some angiotensin II receptor antagonists

A simple, rapid, accurate and highly sensitive spectrofluorimetric method has been developed for determination of some angiotensin II receptor antagonists (AIIRAs), namely Losartan potassium (Los-K), Irbesartan (Irb), Valsartan (Val) and Candesartan cilexetil (Cand) in pure forms as well as in their pharmaceutical dosage forms. All the variables affecting the relative fluorescence intensity (RFI) were studied and optimized. Under the optimum conditions, linear relationships with good correlation coefficients (0.9982–0.9991) were obtained over the concentration range from 0.006 μg/mL to 1.7 μg/mL. Good accuracy and precision were successfully obtained for the analysis of tablets containing each drug alone or combined with hydrochlorothiazide (HCTZ) without interferences from the co-formulated HCTZ or the additives commonly present in tablets.

Selective micellar electrokinetic chromatographic method for simultaneous determination of some pharmaceutical binary mixtures containing non-steroidal anti-inflammatory drugs

A simple and selective micellar electrokinetic chromatographic (MEKC) method has been developed for the analysis of five pharmaceutical binary mixtures containing three non-steroidal anti-inflammatory drugs (NSAIDs). The investigated mixtures were Ibuprofen (IP)–Paracetamol (PC), Ibuprofen (IP)–Chlorzoxazone (CZ), Ibuprofen (IP)–Methocarbamol (MC), Ketoprofen (KP)–Chlorzoxazone (CZ) and Diclofenac sodium (DS)–Lidocaine hydrochloride (LC). The separation was run for all mixtures using borate buffer (20 mM, pH 9) containing 15% (v/v) methanol and 100 mM sodium dodecyl sulphate (SDS) at 15 kV and the components were detected at 214 nm. Different factors affecting the electrophoretic mobility of the seven investigated drugs were studied and optimized. The method was validated according to international conference of harmonization (ICH) guidelines and United States pharmacopoeia (USP). The method was applied to the analysis of five pharmaceutical binary mixtures in their dosage forms. The results were compared with other reported high performance liquid chromatographic methods and no significant differences were observed.

SELECTIVE REVERSED PHASE HIGH PERFORMANCE LIQUID CHROMATOGRAPHY FOR THE SIMULTANEOUS DETERMINATION OF SOME PHARMACEUTICAL BINARY MIXTURES CONTAINING NSAIDS

A simple, rapid, sensitive, and economic reversed-phase High Pressure Liquid Chromatography method was developed for the simultaneous evaluation of five binary mixtures in their pharmaceutical preparations. The investigated mixtures were Ibuprofen(IP)-paracetamol(PC), Ibuprofen(IP)-Chlorzoxazone(CZ), Ibuprofen(IP)- Methocarbamol(MC), Ketoprofen(KP)-Chlorzoxazone(CZ), and Diclofenac sodium(DS)-Lidocaine hydrochloride(LC). The chromatographic separation was performed using isochratic mode at a flow rate 1 mL/ min, ODS C18 column, and UV detection at 254 nm. The mobile phase consisted of acetonitrile:phosphate buffer, pH 5 (60:40). The parameters affecting separation of the investigated drugs were studied. Under the optimum assay conditions, the method gave linear calibration curves with good correlation coefficients. The limits of detection (LOD) and limits of quantitation (LOQ) ranged from 0.34–5.33 µg/mL and 1.03–16 µg/mL, respectively, indicating high sensitivity of the method. The assay of the investigated binary mixtures is completed in less than 5 min. The proposed method was successfully applied to the simultaneous analysis of the drugs in their pharmaceutical binary mixtures in dosage forms with good recoveries and no significant differences between proposed and reported method. This method can be routinely applied to the quality control of the investigated binary mixtures in their pharmaceutical formulations.

Dual separation mode for simultaneous determination of antihypertensive drug combinations by high-performance liquid chromatography.

A simple, reproducible and efficient dual separation mode high performance liquid chromatographic (HPLC) method was developed for simultaneous determination of antihypertensive drug combinations including; hydrochlorothiazide (HCTZ), valsartan (VAL), amiloride (AML) and captopril (CAP). The newly developed Platinum™ column, which provides a dual-mode separation with its polar and non-polar sites, was used for rapid separation of these co-administered drugs. Good resolution was obtained when Platinum™ column was used compared with C(18) column. Additionally, simple isocratic mode with mobile phase containing methanol and 0.02 mole L(-1) phosphate buffer adjusted to pH 3.0 (45:55, v/v) was used for separation. The flow rate was 0.5 mL min(-1) and effluent was monitored at 270 nm. All the investigated drugs were completely separated within less than 6 min. The linearity range obtained for the developed HPLC method was 0.5-100 μg mL(-1) with detection limits of 0.13-1.2 μg mL(-1) for all the studied drugs. The method was validated in accordance with the requirements of ICH guidelines and shown to be suitable for intended applications. The method was successfully used for determination of the studied drugs in pure form and pharmaceutical dosage forms without prior need for separation. The method is valuable for quality control laboratories for simultaneous determination of these co-administered antihypertensive drugs in binary, ternary and quaternary mixtures.

A Specific High-Performance Thin-Layer Chromatography with Fluorescence Detection for the Determination of Some α1-Blockers

A specific high-performance thin-layer chromatography (HPTLC) method with fluorescence detection was developed for determination of selective α1-blockers, including terazosin (TER), doxazosin, prazosin, and alfuzosin. So far, this is the first HPTLC method using fluorescence detection mode for the determination of α1-blockers. It was designated to combine the specificity attributed to the drugs native fluorescence, the high sample throughputs of the HPTLC methods, and the use of environmentally safe reagents. Good resolution was obtained using simple mobile phase composed of acetonitrile and phosphate buffer (0.02 mole/L) adjusted to pH 3.0 (30:70, v/v). The method used fluorescence detection mode and was validated according to the International Conference on Harmonization guidelines. The emission intensity was measured using optical filter K400 after excitation at 332 nm. The correlation coefficient in the range 0.9984–0.9997 was obtained in the concentration range 5–100 ng/band. The limits of detection and quantification were 3.52–6.91 and 11.76–23.02 ng/band, respectively. Forced degradation stability for TER was studied using the developed HPTLC method. The proposed method was successfully applied for the determination of the studied α1-blockers in pharmaceutical dosage forms and in human plasma without the need for complicated extraction and cleanup steps.

Kinetic spectrophotometric method for determination of amlodipine besylate in its pharmaceutical tablets

A simple and sensitive kinetic spectrophotometric method has been developed and validated for determination of amlodipine besylate (AML). The method was based on the condensation reaction of AML with 7-chloro-4-nitro-2,1,3-benzoxadiazole in an alkaline buffer (pH 8.6) producing a highly colored product. The color development was monitored spectrophometrically at the maximum absorption λmax 470 nm. The factors affecting the reaction were studied and the conditions were optimized. The stoichiometry of the reaction was determined, and the reaction pathway was postulated. Moreover, both the activation energy and the specific rate constant (at 70 °C) of the reaction were found to be 6.74 kcal mole−1 and 3.58 s−1, respectively. The initial rate and fixed time methods were utilized for constructing the calibration graphs for the determination of AML concentration. Under the optimum reaction conditions, the limits of detection and quantification were 0.35 and 1.05 μg/mL, respectively. The precision of the method was satisfactory; the relative standard deviations were 0.85–1.76%. The proposed method was successfully applied to the analysis of AML in its pure form and tablets with good accuracy; the recovery percentages ranged from 99.55±1.69% to 100.65±1.48%. The results were compared with that of the reported method.

An efficient one-pot reaction for selective fluorimetric determination of cefpodoxime and its prodrug

Cefpodoxime proxetil (CFP), an oral third-generation cephalosporin, is a prodrug that is de-esterified in vivo to its active metabolite, cefpodoxime acid (CFA). Therefore, this study aimed to develop a facile and efficient one-pot reaction for selective and sensitive determination of CFA and its prodrug (CFP). The method was based on single-step reaction between CFP or CFA and 1,2-naphthoquinone-4-sulfonate (NQS) as a selective derivatizing reagent in alkaline medium without heating, extraction or reduction steps as usual for NQS derivatization reactions. The fluorescence of the formed NQS-derivative was monitored directly at emission wavelength of 440 nm after excitation at 330 nm. The method can easily be implemented in plating facilities by operators and/or incorporated in on-line derivatization reaction. The correlation coefficients of 0.9991 and 0.9984 were obtained in the concentration ranges of 50-2000 ng mL(-1) for CFA and CFP, respectively. The detection limits were 9.17 and 9.48 ng mL(-1) for CFA and CFP, respectively. The method was validated in accordance with the requirements of ICH guidelines and shown to be suitable for their efficient and sensitive determinations. The developed method was successfully applied for selective determination of CFP in pure form and in pharmaceutical dosage forms as well as CFA in human urine after single dose of CFP without prior need for separation. The method is valuable for quality control laboratories for monitoring of CFP and its active metabolite CFA.

Highly sensitive and selective high-performance liquid chromatography method for bioequivalence study of cefpodoxime proxetil in rabbit plasma via fluorescence labeling of its active metabolite.

Cefpodoxime proxetil (CFP), a broad-spectrum third-generation cephalosporin, has been used most widely in the treatment of respiratory and urinary tract infections. For bioequivalence study of CFP in rabbit plasma, it was necessary to develop a highly sensitive and selective high-performance liquid chromatographic (HPLC) method with fluorescence (FL) detection. The pre-column labeling of cefpodoxime acid (CFA) (active metabolite) with an efficient benzofurazan type fluorogenic reagent, 4-N,N-dimethyl aminosulfonyl-7-fluoro-2,1,3-benzoxadiazole (DBD-F) was carried out in the present study in 100mM borate buffer (pH=8.5) at 50°C for 15min. The obtained fluorescent products were separated on C18 column with an isocratic elution of the mobile phase, which consists of 10mM phosphate buffer (pH=3.5)/CH3CN (70:30, v/v). The fluorescent product (DBD-CFA) was detected fluorimetrically at 556nm with an excitation wavelength of 430nm. Cefotaxime sodium was used as internal standard. The method was validated according to the requirements of US-FDA guidelines. The correlation coefficient of 0.999 was obtained in the concentration ranges of 10-1000ngmL(-1). The limits of detection and quantification (S/N=3) were 3 and 10ngmL(-1), respectively. Plasma CFA levels were successfully determined in rabbit with satisfactory precision and accuracy. The proposed HPLC-FL method was successfully applied to study bioequivalence in rabbits for two formulations of different brands contained CFP (prodrug) in a randomized, two-way, single-dose, crossover study and all pharmacokinetic parameters for the two formulations were assessed.