Synchronized spectrofluorimetric determination of ponatinib and curcumin as an effective therapeutic combination in laboratory prepared mixtures and human plasma samples.

Curcumin is a natural product that is frequently utilized in cancer prevention and treatment. The significant benefit of vegetable-derived nutraceuticals in combination with widespread cytostatic medication such as ponatinib is to reduce toxicity and side effects. In this paper, we focus the study on analytical quantification of ponatinib and curcumin through highly sensitive synchronous spectrofluorometric method. Applying this method at Δλ = 160 nm, each of ponatinib and curcumin could be measured at 303 and 412 nm without interference from each others. The diverse experimental factors impacting the performance of the method were studied and optimized. The method exhibited a reasonable linearity in the ranges of 5.0-60.0 and 10.0-200.0 ng/mL for ponatinib and curcumin, respectively with detection limits of 1.48 and 1.22 ng/mL and quantitation limits of 4.49 and 3.68 ng/mL, respectively. The anticipated method was employed for the assessment and evaluation of the studied drugs in the spiked human plasma samples. The mean % recoveries in plasma samples (n = 6) for each of ponatinib and curcumin were 99.84 ± 1.86 and 100.06 ± 2.72, accordingly. The developed method was validated in conformity with the requirements of International Council of Harmonization (ICH).

Multi-spectroscopic and molecular docking studies for binding interaction between fluvoxamine and human serum albumin.

In the present study, different spectroscopic techniques have been used to study the binding interaction between the antidepressant drug fluvoxamine and human serum albumin under simulated physiological conditions (pH 7.4). The utilized spectroscopic techniques include fluorescence emission spectroscopy, synchronous fluorescence spectroscopy, UV-Vis absorption spectroscopy, Fourier Transform Infrared spectroscopy (FT-IR), and molecular modeling methods. The obtained results revealed that the formation of a complex between human serum albumin and fluvoxamine was responsible for quenching the native fluorescence of human serum albumin. The results indicated that the quenching mechanism between human serum albumin and fluvoxamine was static. Besides, the binding constant (K), number of binding sites (n), thermodynamic parameters (ΔH, ΔS, and ΔG), and binding forces were calculated at three different temperatures (298, 310, and 318 K). These data proposed that hydrophobic forces were the principal intermolecular forces stabilizing the complex. From the molecular docking results, it could be deduced that fluvoxamine was inserted into sub-domain II A (site I) of human serum albumin and led to a slight change in human serum albumin conformation.

Conventional and first derivative synchronous spectrofluorimetric methods for the simultaneous determination of cisatracurium and nalbuphine in biological fluids.

Cisatracurium besylate has been determined by fast and highly sensitive spectrofluorimetric method based on measuring the fluorescence intensity of its methanolic solution at 312 nm after excitation at 230 nm (Method I). The linearity occurred over the concentration range of 10.0-130.0 ng/mL with detection limit of 1.07 ng/mL. The method was further extended for the determination of the studied drug in spiked human plasma with good percentage recoveries (97.43-103.50%). Cisatracurium is co-administered with nalbuphine during surgery. The simultaneous determination of both drugs was based on synchronous spectrofluorimetric technique. First derivative synchronous spectrofluorimetric amplitude was measured in methanol at Δ λ = 60 nm and each drug could be estimated at the zero crossing point of the other. Hence, cisatracurium could be measured at 284.6 nm while nalbuphine at 276.3 nm (Method II). The method was linear over the ranges of 50.0-750.0 ng/mL and 0.5-7.0 µg/mL with the detection limits of 2.16 ng/mL and 0.04 µg/mL for cisatracurium and nalbuphine, respectively. The method was further extended for the simultaneous determination of both drugs in spiked human urine with mean percentage recoveries of 99.99 ± 2.06 and 99.53 ± 6.17 for cisatracurium and nalbuphine, respectively. Both methods were validated in agreement with Guidelines adopted by International Council of Harmonization (ICH).

First derivative synchronous spectrofluorimetric method for the simultaneous determination of propofol and cisatracurium besylate in biological fluids.

Propofol and cisatracurium besylate have been simultaneously determined using a highly sensitive first derivative synchronous spectrofluorometric method. The method is based on measuring first derivative synchronous spectrofluorimetric amplitude at Δλ = 40 nm with a scanning rate of 600 nm/min. The different experimental parameters affecting the fluorescence intensity of the two drugs were carefully studied and optimized. The amplitude-concentration plots were rectilinear over the range 40.0-400.0 ng/mL and 20.0-280.0 ng/mL for propofol and cisatracurium, respectively with lower detection limits of 4.0 and 2.35 ng/mL and quantification limits of 12.1 and 7.1 ng/mL for propofol and cisatracurium, respectively. The proposed method was successfully applied for the determination of the two compounds in synthetic mixtures and in commercial ampoules. The high sensitivity attained using the proposed method allowed the simultaneous determination of both drugs in spiked plasma samples. The mean % recoveries in spiked human plasma (n = 3) were 96.53 ± 0.90 and 96.20 ± 1.64 for each of propofol and cisatracurium, respectively. The method was validated in compliance with International Council of Harmonization (ICH) Guidelines.

Green analytical chromatographic assay method for quantitation of cyclobenzaprine in tablets, spiked human urine and in-vitro dissolution test.

Cyclobenzaprine hydrochloride, a skeletal muscle relaxant has been determined using an ecofriendly micellar HPLC method in its pure form and tablets. The chromatographic determination was performed using C8 monolithic column (100mm×4.6mm i.d., 5µm particle size) and micellar eluent which was composed of sodium dodecyl sulfate (0.15M), n-propanol (15%), 0.02M orthophosphoric acid (pH 4.5) and 0.3% triethylamine using UV detection of effluent was set at 225nm. The calibration plot showed good linearity over concentration range from 2-40µg/mL. The assay results were statistically validated for linearity, accuracy, precision and specificity according to ICH guidelines. Additionally, regarding USP guidelines, the uniformity of tablets content and in-vitro dissolution test of the tablets was tested using the proposed method. Simple and rapid applicability of the developed method allowed determination of the drug in its pure and tablet dosage forms. Moreover, the major advantage of micellar HPLC technique is to determine the drug in biological fluids without prior extraction steps. Depending on this, the estimation of cyclobenzaprine in spiked human urine was so simple without traditional tedious procedures. The proposed method offers the advantages of sensitivity and simplicity in addition to short analysis time which didn't exceed 6 minutes.

Simultaneous HPLC determination of alfuzosin, tamsulosin and vardenafil in human plasma and pharmaceutical formulations using time programmed fluorescence detection.

Alfuzosin and tamsulosin are recently co-administrated with vardenafil to treat symptoms of benign prostatic hyperplasia and erectile dysfunction. A highly sensitive and simple liquid chromatographic method was developed and validated for the simultaneous determination of the three drugs using moxifloxacin as an internal standard. Isocratic separation was achieved within 7.0 min using phenyl-hexyl column (250 × 4.6 mm i.d.) and a mobile phase composed of acetonitrile/0.25% phosphoric acid (30:70, v/v) at pH 3.0. The analysis was performed at a flow rate of 1.2 mL/min with fluorescence detection at 246/450 nm for Alfuzosin and vardenafil, and 226/322nm for tamsulosin using time programming technique. The proposed method was linear over the concentration ranges of 5.0-50.0ng/mL, 10.0-200.0ng/mL and 20.0-400.0ng/mL for alfuzosin, vardenafil and tamsulosin, with limits of detection of 0.56ng/mL, 0.98ng/mL and 2.81 ng/mL in a respective order. The developed method was successfully applied to determine the studied drugs in dosage forms and human plasma samples and the results were satisfactory as revealed by statistical analysis of the data.

Micelle-enhanced spectrofluorimetric determination of amlexanox in bioadhesive buccal tablets: application to content uniformity testing.

A highly sensitive, simple and rapid spectrofluorimetric method was developed for the determination of Amlexanox (AMX) in its bioadhesive buccal tablets. The proposed method is based on measuring the native fluorescence of the methanolic solution of AMX at 400 nm after excitation at 242 nm in 0.2 M borate buffer (pH 10) and 0.5% w/v sodium dodecyl sulfate (SDS) solution. The interaction of AMX with SDS was studied, and the enhanced fluorescence intensity was exploited to develop an assay method for the determination of AMX. The relative fluorescence intensity-concentration plot was rectilinear over the range 5.0-80.0 ng/mL, with a lower detection limit of 0.57 ng/mL and a lower quantification limit of 1.74 ng/mL. The proposed method was successfully applied to the analysis of AMX in its commercial tablets. Moreover, content uniformity testing was conducted by applying official USP guidelines. Statistical evaluation and comparison of the data obtained using the proposed and comparison methods revealed good accuracy and precision for the proposed method.

Spectrofluorimetric determination of amisulpride and bumidazone in raw materials and tablets.

A highly sensitive, simple and rapid spectrofluorimetric method was developed for the determination of amisulpride (AMS) and bumidazone (BUM) in tablet form. The proposed method is based on measuring the native fluorescence of the studied drugs in methanol at 360 and 344 nm after excitation at 276 and 232 nm for AMS and BUM, respectively. The fluorescence-concentration plots were rectilinear over the ranges of 5.0-60.0 ng/mL for AMS and 0.5-5.0 µg/mL for BUM. The lower detection limits were 0.70 ng/mL and 0.06 µg/mL, and the lower quantification limits were 2.0 ng/mL and 0.18 µg/mL for AMS and BUM, respectively. The method was successfully applied for the analysis of AMS and BUM in commercial tablets. Statistical evaluation and comparison of the data obtained using the proposed and comparison methods revealed good accuracy and precision for the proposed method.

Stability-Indicating Spectrofluorimetric Methods for the Determination of Metolazone and Xipamide in Their Tablets. Application to Content Uniformity Testing.

A highly sensitive, simple and rapid stability-indicating spectrofluorimetric method was developed for the determination of metolazone (MET) and xipamide (XPM) in their tablets. The proposed method is based on the measurement of the native fluorescence of MET in methanol at 437 nm after excitation at 238 nm and XPM in alkaline methanolic solution at 400 nm after excitation at 255 nm. The fluorescence-concentration plots were rectilinear over the range of 2.0- 20.0 ng/mL for MET and 0.2- 2.0 µg/mL for XPM, with lower detection limits (LOD) of 0.35 ng/mL and 0.02 µg/mL and a lower quantification limit (LOQ) of 1.05 ng/mL and 0.07 µg/mL for MET and XPM, respectively. The method was successfully applied to the analysis of MET and XPM in their commercial tablets and the results were in good agreement with those obtained using the official and comparison methods, respectively. Furthermore, content uniformity testing of the studied pharmaceutical tablets was also conducted. The application of the proposed method was extended to stability studies of MET and XPM after exposure to different forced degradation conditions, such as acidic, alkaline, oxidative and photolytic degradation conditions, according to ICH Guidelines. Moreover, the method was utilized to investigate the kinetics of the alkaline, acidic and photolytic degradation of MET. The apparent first-order rate constants and half-life times were calculated. Proposals for the degradation pathways for both MET and XPM were postulated.

Highly sensitive HPLC method for assay of aliskiren in human plasma through derivatization with 1-naphthyl isocyanate using UV detection.

A simple and sensitive HPLC method has been developed for the determination of aliskiren in human plasma through derivatization with 1-naphthyl isocyanate. The separation was achieved on a C18 column using a mobile phase consisting of acetonitrile/water/phosphoric acid (45:55:0.01, v/v/v, pH 3.2) in a flow rate of 1mL/min with UV detection at 230nm. Caffeine was used as an internal standard. The factors influencing the derivatization reaction yields were carefully studied and optimized. The method was linear over the concentration range of 5-400ng/mL with a detection limit of 0.5ng/mL and a limit of quantification of 1.0ng/mL. The relative standard deviation was less than 4.2% for both intra-day assay and inter-day assay results. No interferences from endogenous compounds were encountered. The percentage recovery was in the range 97.1-98.6%. The method is suitable for routine therapeutic drug monitoring and for pharmacokinetic studies.

Simultaneous determination of aliskiren and hydrochlorothiazide in tablets and spiked human urine by ion-pair liquid chromatography.

An alternative method for analysis of aliskiren (ALI) and hydrochlorothiazde (HCT) in combined dosage forms by ion-pair reversed phase high performance liquid chromatography was developed and validated. The pharmaceutical preparations were analyzed using a C18 column (250 mm x 4.6 mm, 3 microm) with a mobile phase consisting of 25% methanol, 50% sodium monobasic phosphate aqueous solution containing 6 mM tetrabutylammonium bromide and 25% water at pH 7.2. Isocratic analysis was performed at a flow rate of 1 mL/min and a column temperature of 30 degrees C under direct UV detection at 210 nm. Paracetamol was used as internal standard. The validation was performed according to the ICH guidelines. The proposed method was linear over the concentration range of 0.250 to 60 and 0.1 to 10 microg/mL for ALI and HCT, respectively. The limits of detection and quantitation (LOD and LOQ) were 0.075 and 0.198 microg/mL, respectively, for ALI and 0.04 and 0.062 microg/mL, respectively, for HCT. The method proved to be specific, sensitive, precise and accurate with mean recovery values of 101.1 +/- 0.32% and 100.9 +/- 0.41% for ALI and HCT, respectively. The method robustness was evaluated by means of an experimental design. The proposed method was applied successfully to spiked human urine samples with mean recoveries of 98.8 +/- 0.36% and 98.1 +/- 0.21% for ALI and HCT, respectively.

Spectrofluorimetric determination of oseltamivir phosphate through derivatization with o-phthalaldehyde. Application to pharmaceutical preparations with a preliminary study on spiked plasma samples.

A simple and sensitive spectrofluorimetric method has been developed and validated for the determination of oseltamivir phosphate (OST) in pharmaceutical preparations. The method is based on the reaction between oseltamivir phosphate and o-phthalaldehyde in presence of 2-mercapto-ethanol in borate buffer, pH 10.8, to give a highly fluorescent product measured at 450 nm after excitation at 336 nm. The different experimental parameters affecting the development and stability of the reaction product were studied and optimized. The fluorescence intensity-concentration plot is rectilinear over the range 0.05-1.0 µg/mL, with a lower detection limit of 5 ng/mL and limit of quantitation of 16 ng/mL. The developed method was successfully applied to the analysis of the drug in its commercial capsules and suspension, mean recoveries of OST were 99.97 ± 1.67% and 100.17 ± 1.18%, respectively (n = 3). Statistical comparison of the results obtained by the proposed and comparison method revealed no significant difference in the performance of the two methods regarding accuracy and precision. The proposed method was further extended to in vitro determination of the studied drug in spiked human plasma as a preliminary investigation; the mean recovery (n = 3) was 98.68 ± 5.8%. A reaction pathway was postulated.

Stability-indicating HPLC method for determination of naftazone in tablets. Application to degradation kinetics and content uniformity testing.

A simple, sensitive, stability-indicating HPLC method was developed and validated for the quantitative determination of the vasoprotective drug, naftazone in presence of its degradation products. The analysis was carried out on a Nucleosil 100-5 phenyl column (250 mm × 4.6 mm, 5 µm) using a mobile phase consisting of methanol-0.02 M sodium dihydrogen phosphate mixture (60:40, v/v) of pH 6.0. The analyses were performed at ambient temperature with a flow rate of 1.0 mL/min and UV detection at 270 nm. The method showed good linearity over the concentration range of 0.1-10.0 µg/mL with a lower detection limit of 0.032 and quantification limit of 0.096 µg/mL. The suggested method was successfully applied for the analysis of naftazone in its commercial tablets. Moreover, it was utilized to investigate the kinetics of alkaline, acidic and oxidative degradation of the drug. The apparent first-order rate constants, half-life times, and activation energies of the degradation process were calculated. The pH-rate profile curve was derived. Furthermore, the proposed method was successfully applied to the content uniformity testing of naftazone tablets.

Stability-indicating micelle-enhanced spectrofluorimetric method for determination of loratadine and desloratadine in dosage forms.

A highly sensitive and simple spectrofluorimetric method was developed for the determination of loratadine (LRT) and desloratadine (DSL) in their pharmaceutical formulations. The proposed method is based on investigation of the fluorescence spectral behaviour of LRT and DSL in a sodium dodecyl sulphate (SDS) micellar system. In aqueous solution of acetate buffer of pH 4.5, the fluorescence intensities of both LRT and DSL were greatly enhanced (240%) in the presence of SDS. The fluorescence intensity was measured at 438 nm after excitation at 290 nm for both drugs. The fluorescence-concentration plots were rectilinear over the range 0.05-2.0 µg/mL for both LRT and DSL, with lower detection limits of 5.13 × 10(-3) and 6.35 × 10(-3) µg/mL for LRT and DSL, respectively. The method was successfully applied to the analysis of the two drugs in their commercial tablets, capsules and syrups, and the results were in good agreement with those obtained with the official or comparison methods. The proposed method is specific for the determination of LRT in the presence of other co-formulated drugs, such as pseudoephedrine. The application of the proposed method was extended to stability studies of LRT and DSL after exposure to different forced degradation conditions, such as acidic, alkaline and oxidative conditions, according to ICH guidelines.

Determination of oxybutynin in pharmaceuticals via reaction with mixed acids anhydrides: application to content uniformity testing.

Sensitive and simple spectrophotometric (Method I) and spectrofluorimetric (Method II) methods were developed and validated for the determination of oxybutynin HCl (OXB) in its dosage forms. The method was based on the reaction of OXB with malonic acid anhydride in acetic acid anhydride to form a highly yellow colored product that was measured at 375 nm spectrophotometrically. The same reaction product exihibits strong fluorescence that was measured at 440 nm after excitation at 390 nm. The factors affecting formation and stability of the reaction product were carefully studied and optimized, and the reaction mechanism was postulated. The absorbance-concentration plot is rectilinear over the range 4-40 µg/mL with LOD of 1.12 µg/mL and LOQ of 3.39 µg/mL. The fluorescence-concentration plot is rectilinear over the range 0.5-6 µg/mL with LOD of 0.11 µg/mL and LOQ of 0.33 µg/mL. The method was applied to the analysis of commercial tablets Detronin® and Uripan®. Statistical comparison of the results with those of the reference method revealed good agreement and proved that there were no significant difference in the accuracy and precision between the two methods respectively. The study was extended to content uniformity testing.

Simple and sensitive spectrofluorimetric method for the determination of pregabalin in capsules through derivatization with fluorescamine.

A new, simple and sensitive spectrofluorimetric method has been developed for the determination of pregabalin (PG) in capsules. The method is based on the reaction between pregabalin and fluorescamine in borate buffer solution of pH 10 to give a highly fluorescent derivative that is measured at 487 nm after excitation at 390 nm. The different experimental parameters affecting the development and stability of the reaction product were carefully studied and optimized. The fluorescence intensity concentration plot was rectilinear over the range of 0.01-0.3 µg mL⁻¹ with a lower detection limit of 0.0017 µg mL⁻¹ and limit of quantitation of 0.005 µg mL⁻¹. The developed method was successfully applied to the analysis of the drug in its commercial capsules. The mean percentage recovery of PG in its capsule was 99.93±1.24 (n = 3). Statistical comparison of the results with those of the comparison method revealed good agreement and proved that there was no significant difference in the accuracy and precision of the two methods. A proposed reaction pathway was postulated.

First derivative synchronous fluorescence spectroscopy for the simultaneous determination of sulpiride and mebeverine hydrochloride in their combined tablets and application to real human plasma.

A rapid, simple and highly sensitive first derivative synchronous fluorometric method has been developed for the simultaneous analysis of binary mixture of sulpiride (SUL) and mebeverine hydrochloride (MEB). The method is based upon measurement of the synchronous fluorescence intensity of these drugs at ∆λ = 100 nm in water. The different experimental parameters affecting the fluorescence of the two drugs were carefully studied and optimized. The fluorescence-concentration plots were rectilinear over the range of 0.05-1 µg/mL and 0.2-3.2 µg/mL for SUL and MEB respectively with lower detection limits (LOD) of 0.006 and 0.01 µg/mL and quantification limits (LOQ) of 0.0.02 and 0.05 µg/mL for SUL and MEB, respectively. The proposed method was successfully applied for the determination of the two compounds in synthetic mixtures and in commercial tablets. The high sensitivity attained by the proposed method allowed the determination of both of SUL and MEB metabolite (veratic acid) in real human plasma samples applying second derivative synchronous fluorometric technique. The mean% recoveries (n = 3) for both MEB metabolite (veratic acid) and SUL were 99.82 ± 2.53 and 98.84 ± 6.20 for spiked human plasma respectively, while for real human plasma, the mean% recoveries (n = 3) were 91.49 ± 4.25 and 91.36 ± 8.46 respectively.

Validated Stability indicating Spectrophotometric Method for the Determination of Acetazolamide in Dosage Forms through Complex Formation with Palladium (II).

A simple and sensitive spectrophotometric method was developed for the determination of acetazolamide (ACM) in pure form and pharmaceutical preparations. The proposed method is based on the complex formation of acetazolamide with Palladium (II) chloride in acetate buffer pH5.4 and measuring the absorbance at 308 nm. The absorbance- concentration plot was rectilinear over the concentration range of 5-70 µg/ml with a minimum detection limit (LOD) of 0.98 µg/ml, limit of quantification (LOQ) of 2.96 µg/ml, and a molar absorptivity ζ=2.7 × 10(3) L/mol.cm. The factors affecting the absorbance of the formed complex were carefully studied and optimized. The composition of the complex as well as its stability constant was also investigated. The proposed method was applied for the determination of acetazolamide in its tablets and the results obtained were favorably compared with those obtained using the official method. A proposal of the reaction pathway was postulated.

A New Spectrophotometric Method for Determination of Phenylpropanolamine HCl in its Pharmaceutical Formulations via Reaction with 2,3,5,6-tetrachloro-1,4-benzoquinone.

A selective and simple spectrophotometric method has been developed for the determination of phenylpropanolamine HCl (PPA) in its dosage forms. The method was based on the formation of a colored N-vinyl chlorobenzoquinone derivative of PPA through its reaction with 2,3,5,6-tetrachloro-1,4-benzoquinone in presence of acetaldehyde. The colored product exhibits maximum absorbance at 650 nm. Different experimental parameters affecting formation and stability of the product were carefully studied and optimized. The stoichiometry of the reaction was determined, and the reaction pathway was postulated. The absorbance concentration plot was rectilinear over the range of 5-100 µg/mL with Limit of Detection (LOD) and Limit of Quantitation (LOQ) of 0.244 µg/mL and 0.74 µg/mL respectively. The analytical performance of the method was fully validated, and the results were satisfactory. The proposed method was successfully applied to the determination of PPA in its commercial dosage forms including tablets, capsules and syrups with good recoveries. Statistical comparison of the results with those of the comparison method showed good agreement and proved that there was no significant difference in the accuracy and precision between the reference and the proposed methods. The mechanism of the reaction pathway was postulated.

Spectrophotometric Determination of the Antidepressants Sertraline and Paroxetine HCl using 2,4-Dinitrofluorobenzene.

A simple and sensitive spectrophotometric method was developed for the determination of each of sertraline (SER) and paroxetine HCl (PXT) in dosage forms. The method is based upon reaction of PXT and SER with 2,4-dinitrofluorobenzene (DNFB) to form colored products. The absorbance of the products were measured at 375and 390 nm for SER and PXT respectively. The absorbance concentration plots were rectilinear over the concentration rang of 1-10 and 2-20 µg/mL with lower detection limits (LOD) of 0.11 and 0.28 µg/mL and quantification limits (LOQ) of 0.32 and 0.85 µg/mL for SER and PXT, respectively. The developed method was successfully applied for the determination of SER and PXT in dosage forms. The common excipients and additives did not interfere in their determinations. There was no significant difference between the results obtained by the proposed and the reference methods regarding Student t-test and the variance ratio F-test respectively. A proposal of the reaction pathway was postulated.