AQbD TLC-densitometric method approach along with green fingerprint and whiteness assessment for quantifying two combined antihypertensive agents and their impurities.

Preserving the environment, reducing the amount of waste resulting from chemical trials, and reducing the amount of energy consumed have currently become a pivotal global trend. An analytical quality by design (AQbD) based eco-friendly TLC-densitometric method was implemented for quantifying two antihypertensive agents, captopril (CPL) and hydrochlorothiazide (HCZ), along with their impurities; captopril disulphide (CDS), chlorothiazide (CTZ) and salamide (SMD). The analytical target profile (ATP) was first identified, followed by selecting the critical analytical attributes (CAAs), such as retardation factors and resolution between the separated peaks. Critical method parameters (CMPs) that may have a crucial influence on CAAs were identified and emanated through the quality risk assessment phase. A literature survey-based preliminary studies were performed, followed by optimization of the selected CMPs through a custom experimental design to attain the highest resolution with optimum retardation factors. Moreover, method robustness was also tested by testing the design space. Complete separation of the drugs and their impurities was achieved using ethyl acetate: glacial acetic acid (6: 0.6, v/v) as a developing system applied to a 12 cm length TLC plate at room temperature with UV scanning at 215 nm. Calibration graphs were found to be linear in the ranges of (0.70-6.00), (0.10-2.00), (0.20-1.00), (0.07-1.50) and (0.05-1.00) µg/band corresponding to CPL, HCZ, CDS, CTZ, and SMD, respectively. Four different green metric tools were used to evaluate the greenness profile of the proposed method, and results showed that it is greener than the reported HPLC method. Method whiteness assessment was also conducted. Moreover, the method performance was evaluated following the ICH guidelines, and the outcomes fell within the acceptable limits. The developed method could be approved for routine assay of the cited components in their pharmaceutical formulations and bulk powder without interference from the reported impurities. The issue of concern is saving money, especially in developing countries.

Ecofriendly single-step HPLC and TLC methods for concurrent analysis of ternary antifungal mixture in their pharmaceutical products.

Two accurate, sensitive, and selective methods for simultaneous determination of miconazole nitrate (MIC), nystatin (NYS), and metronidazole (MET) in pure state or drug product were established and verified. First, RP-HPLC-DAD was designed. Separation was accomplished using a ZOBRAX Eclipse Plus RP-C8 column that was running under an isocratic elution of methanol: 0.05% aqueous solution of sodium dodecyl sulphate (40: 60 v/v), with a flow rate that was regulated at 0.8 mL/min. The column temperature was adjusted at 25 °C and diode array detector was monitored at 220 nm. The linearity range of the proposed method was achieved at the concentration of 5-50, 4-50, and 4-40 µg/mL and the attained retention time for the studied drugs was 2.52, 3.52 and 4.99 min for MIC, NYS, and MET, correspondingly. Second, a TLC-densitometric approach was used to resolve the three compounds. Resolution of the three cited drugs was carried out using TLC aluminum plates pre-coated with 0.25 mm silica gel 60 F254. A developing solvent comprised ethyl acetate: toluene: methanol: triethyl amine: formic acid (3: 1: 7: 0.3: 0.1 by volume) (pH = 5.5) was utilized and scanning of the resolved bands at 215 nm. Linearity of the developed TLC method was evaluated and evident to be 0.4-2, 0.4-2.2, and 0.4-2 µg/band for MIC, NYS, and MET, in that order. The suggested chromatographic methods were verified according to ICH directives. The findings of the developed chromatographic procedures were statistically compared with the results of the reported ones using student's t-test and F-test. Furthermore, two green assessment tools evaluated the indicated methods' level of greenness (GAPI and AGREE).

Greenness assessment of UPLC/MS/MS method for determination of two antihypertensive agents and their harmful impurities with ADME/TOX profile study.

Hypertension is described by the world health organization (WHO) as a serious medical problem that significantly affects the heart, brain and kidneys. It is a major cause of premature death worldwide. The present study aims to quantify the combination of captopril (CPL), hydrochlorothiazide (HCZ) and their harmful impurities; captopril disulphide (CDS), chlorothiaizde (CTZ) and salamide (SMD). In-silico study was conducted for estimation of pharmacokinetic parameters (ADMET) as well as toxicity profile of the proposed impurities. The results showed that the three impurities under investigation had poor permeability to CNS and cannot pass the blood-brain barrier (BBB), reducing the likelihood of causing side effects in the brain. On the other hand, all studied impurities were found to be hepatotoxic. In consequence, a highly sensitive and green ultra-performance liquid chromatography- tandem mass spectrometric (UPLC/MS/MS) method was developed and validated for separation of the cited drugs in the presence of their harmful impurities; methanol and 0.1% formic acid (90:10, v/v) mixture was used as a mobile phase, eluted at a constant flow rate of 0.7 mL/min at room temperature. Detection was adopted using a tandem mass spectrometer in a positive mode only for CPL and negative mode for HCZ, CDS, CTZ and SMD. Separation was performed within 1 min. Calibration graphs were found to be linear in the ranges of (50.0-500.0 ng mL-1), (20.0-500.0 ng mL-1), (10.0-250.0 ng mL-1), (5.0-250.0 ng mL-1) and (20.0-400.0 ng mL-1) corresponding to CPL, HCZ, CDS, CTZ and SMD, respectively. Additionally, comparative study of greenness profile was established for the proposed and reported methods using five green metric tools. The proposed method was found to be greener than the reported HPLC method. The developed (UPLC/MS/MS) method was validated according to (ICH) guidelines and it was found to has greater sensitivity, shorter analysis time and lower environmental impact compared to the reported methods.

Comparative Chemometric Manipulations of UV-Spectrophotometric Data for the Efficient Resolution and Determination of Overlapping Signals of Cyclizine and Its Impurities in Its Pharmaceutical Preparations.

BACKGROUND: Cyclizine (CYZ), a commonly used antiemetic drug, has two pharmacopeial toxic impurities, 1-methylpiperazine (MPZ) and diphenylmethanol (DPM). When CYZ parenteral formulations are administered intravenously, both impurities are poisonous, toxic, and harmful to the human body. OBJECTIVE: Cyclizine was determined along with its hazardous impurities MPZ and DPM by green multivariate calibration using UV-spectroscopic data. METHODS: Three multivariate algorithms were used to resolve and quantify overlapped spectral signals: principal component regression (PCR), partial least squares (PLS), and synergistic intervals partial least squares (siPLS). A concentration set containing 16 distinct combinations of CYZ, MPZ, and DPM was randomly prepared, and the absorbance values of the concentration set were determined using the 376 point-wavelength set with an interval of 0.2 nm between 200 and 275 nm. RESULTS: Good linear correlations were established for CYZ, MPZ, and DPM in the concentration ranges of 5.00-25.0, 0.50-2.50, and 0.50-2.50 µg/mL, respectively. The ideal spectral range and associated combinations were chosen based on the lowest root mean error of prediction (RMSEP) and correlation coefficient values (r). The siPLS approach performed better than the PCR and PLS models. The combination of four subintervals, 1, 3, 4, and 7, demonstrated the greatest effect, with RMSEP values of 0.0272, 0.0053, and 0.0315 for CYZ, MPZ, and DPM, respectively, and correlation coefficients of 0.9991, 0.9999, and 0.9997, in order. Various assessment tools were used to evaluate and measure the greenness profile of the established methods. The proposed methods were validated using internal and external validation sets. CONCLUSIONS: The three methods were effectively used to determine CYZ in its pure form and parenteral formulations, as well as its toxic impurities. The acquired results were compared statistically to those obtained using the reported HPLC method. HIGHLIGHTS: Cyclizine and its toxic impurities can be determined spectrophotometrically by using the three developed chemometric models.

Quality by design approach for green HPLC method development for simultaneous analysis of two thalassemia drugs in biological fluid with pharmacokinetic study.

This work implements a combined experimental approach of analytical quality-by-design (AQbD) and green analytical chemistry (GAC) to develop an HPLC method for simultaneous determination of the two thalassemia drugs, deferasirox (DFX) and deferiprone (DFP), in biological fluid for the first time. This integration was designed to maximize efficiency and minimize environmental impacts, as well as energy and solvent consumption. To accomplish this goal, an analytical quality-by-design approach was performed, beginning with quality risk assessment and scouting analysis, followed by Placket-Burman design screening for five chromatographic parameters. Critical method parameters were thoroughly recognized and then optimized by using a two levels-three factors custom experimental design to evaluate the optimum conditions that achieved the highest resolution with acceptable peak symmetry within the shortest run time. The desirability function was used to define the optimal chromatographic conditions, and the optimal separation was achieved using an XBridge® HPLC RP-C18 (4.6 × 250 mm, 5 µm) column with ethanol : acidic water at pH 3.0 adjusted by phosphoric acid in the ratio of (70 : 30, v/v) as the mobile phase at a flow rate of 1 mL min-1 with UV detection at 225 nm at a temperature of 25 °C. Linearity was obtained over the concentration range of 0.30-20.00 µg mL-1 and 0.20-20.00 µg mL-1 for DFX and DFP, respectively, using 20.00 µg mL-1 ibuprofen (IBF) as an internal standard. The established method's greenness profile was evaluated and measured using various assessment tools, and the developed method was green. For the validation of the developed method, FDA recommendations were followed, and all the results obtained met the acceptance criteria. The suggested method was successfully used to study the pharmacokinetic parameters of DFX and DFP in rat plasma. Due to the substantial increase in bioavailability of the two iron chelating drugs, the results from this study strongly recommend their co-administration.

Spectrofluorimetric Approach for Quantification of Cyclizine in the Presence of its Toxic Impurities in Human Plasma; in silico Study and ADMET Calculations.

Cyclizine (CYZ); an antiemetic compound; is widely misused for its euphoric or hallucinatory effects, either by oral or intravenous routes. The concomitant abuse of CYZ among addicted adolescents contributes to neuromuscular disorders that are life-threatening. Consequently, with the company of 1-Methylpiperazine (MPZ) and diphenylmethanol (DPM, Benzhydrol) as pharmacopoeia-reported CYZ impurities, a novel spectrofluorimetric assay for the detection of CYZ, has been established either in human plasma samples or in its parenteral formulation. The native fluorescence of CYZ has been investigated under various conditions. Different parameters affecting relative fluorescence intensity of CYZ including diluting solvent, surfactant, plasma protein solvent, and pH were studied and optimized. The linearity obtained between the fluorescence intensity at emission wavelength 350 nm after excitation at 244 nm and the corresponding CYZ concentrations was in the range 10-1000 ng/mL for measurement of CYZ either in pure form or in human plasma samples, with a appropriate correlation coefficient (r = 0.9999) and 3.10 ng/mL as the limit of detection and 9.41 ng/mL as the limit of quantitation. The suggested procedure was created and validated in accordance with ICH guidelines for quantification of CYZ either in its pure form or its dosage form, and FDA guidelines for the assay of CYZ in human plasma. Finally, in silico study and ADMET predictions were conducted for the studied drug impurities to estimate their pharmacokinetic behaviors. The results showed that both CYZ impurities have higher cellular permeability and maximum tolerated doses, DPM has higher BBB and CNS permeability than MPZ, while MPZ exceeds DPM in total clearance and volume of distribution.

Nanoparticle-enhanced in-line potentiometric ion sensor for point-of-care diagnostics for tropicamide abuse in biological fluid.

Point of care (POC), also identified as on-site testing, has evolved as a rapid and accurate technique for drug of abuse screening and analysis. The aim of this work is to detect tropicamide (TPC) abuse in biological fluids; we selected rat plasma as example. We developed a disposal miniaturized, portable, green, and budget-friendly POC solid-state electrochemical sensor based on potentiometric transduction. To attain that, an innovative microfabricated electrode modified with conducting polymer poly(3-octylthiophene) (POT) has been placed on sensitized printed circuit board (PCB). A two-stage optimization process was implemented to develop the fabricated electrode. The first stage of the optimization process depends on screening various ionophores in order to enhance the sensor selectivity towards tropicamide. Copper nanoparticles exhibited the highest selectivity towards TPC. The second stage was utilizing a polymer as an ion-to-electron transducer layer between the copper nanoparticles impregnated ion sensing membrane and the microfabricated solid-contact ion-selective electrode. This polymer was added to boost the stability of the potential drift (1.2 mV/h) due to the hydrophobic behavior of the POT, which precludes the formation of an aqueous layer at the Cu electrode/polymeric membrane interface and improve the limit of detection (1.1 × 10-8 M). Nernstian potentiometric response was accomplished for TPC with a slope of 58.46 ± 0.43 mV/decade and E0 ∼ 189.39 ± 2.12 over the concentration range 1.0 × 10-7 to 1.0 × 10-2 M. The suggested sensor's intrinsic figure of merits include a quick response time (13 ± 2 s) and long life time (45 days). The proposed sensor has been successfully employed in the selective determination of TPC in pharmaceutical formulations, and biological fluids. When the results were compared to those of the official approach, there was no statistically significant difference. The Eco-Scale tool assessed and measured the greenness profile of the established method.

Determination of Sulphasalazine and Its Related Compounds by Simple, Smart, Validated, Green Spectrophotometric Methods.

BACKGROUND: Sulphasalazine (SZ) is a prodrug. Its active metabolite (mesalazine, MZ), which is also available in pharmaceutical formulations, and the major active metabolite of MZ (N-acetyl-5-aminosalicylic acid, AS) are commonly used for the treatment of inflammatory bowel diseases. OBJECTIVE: Two accurate, precise, sensitive, and specific spectrophotometric methods were developed and validated for determination of the studied components. METHODS: The first method is a modified ratio difference spectrophotometric method. In this method, SZ was determined by measuring the peak area from 410-500 nm, while MZ and AS were determined by measuring the difference of the selected amplitude values. The second method is a mean centering of ratio spectra spectrophotometric method. RESULTS: The developed methods were linear over the concentration ranges of 2-35, 2-30 and 1-25 µg/mL for SZ, MZ and AS, respectively. CONCLUSION: The developed methods were validated according to International Conference on Harmonization guidelines. They were successfully applied for determination of studied analytes. A greenness assessment was undertaken using three different tools. HIGHLIGHTS: Spectrophotometric methods were developed for determination of SZ and its related compounds for the first time. They were designated to be green and eco-friendly and their greenness profiles were evaluated using green solvents to keep the environment clean.

Lipophilicity study of different cephalosporins: Computational prediction of minimum inhibitory concentration using salting-out chromatography.

The chromatographic and lipophilicity characters of seven cephalosporins of different four classes (cephradine, cefaclor, cefprozil, cefixime, cefotaxime, ceftazidime and cefepime) were examined by salting out thin-layer chromatography (SOTLC). SOTLC using ammonium sulfate salt was employed to predict the lipophilicity of the proposed drugs via their retention behavior. The calculated RM0 values showed liner relationship with the molar concentration of ammonium sulfate in mobile phase in the range of 0.5-2.5 mol/L. Additionally, quantitative structure retention relationship (QSRR) was generated to figure out the relationship between the calculated chromatographic parameters (RM0 and C0) and log P of the studied cephalosporins. Good correlations were found between the chromatographically obtained retention parameters (RM0 and C0) and some molecular descriptors of the examined drugs. Furthermore, an efficient QSAR model was carried out using the calculated chromatographic parameters (RM0 and C0) and log P of the studied cephalosporins to predict minimum inhibitory concentration (MIC) and blood brain barrier (BBB) penetration of the examined drugs. The study was extended to separate and quantify the selected antibiotics in their pure forms and pharmaceutical formulations. Normal phase thin layer chromatographic (NP-TLC) method using a usable developing system of acetone: methanol: water: ammonium hydroxide: glacial acetic acid (90: 10: 18: 3: 2, by volume) was successfully applied to resolve the studied cephalosporins. Linearity was achieved in the range of 0.2-3 µg/mL for most of the studied antibiotics. The developed SOTLC method can be considered as a good start alternative to reversed phase thin layer chromatography (RP-TLC) for prediction of the lipophilic properties of examined cephalosporins. Moreover, the proposed NP-TLC densitometric method can be easily applied for quality control analysis of the chosen drugs and other structurally related components.

Development and validation of a stability indicating RP-HPLC-DAD method for the determination of bromazepam.

A reliable, selective and sensitive stability-indicating RP-HPLC assay was established for the quantitation of bromazepam (BMZ) and one of the degradant and stated potential impurities; 2-(2-amino-5-bromobenzoyl) pyridine (ABP). The assay was accomplished on a C18 column (250 mm × 4.6 mm i.d., 5 µm particle size), and utilizing methanol-water (70: 30, v/v) as the mobile phase, at a flow rate of 1.0 ml min-1. HPLC detection of elute was obtained by a photodiode array detector (DAD) which was set at 230 nm. ICH guidelines were adhered for validation of proposed method regarding specificity, sensitivity, precision, linearity, accuracy, system suitability and robustness. Calibration curves of BMZ and ABP were created in the range of 1-16 µg mL-1 with mean recovery percentage of 100.02 ± 1.245 and 99.74 ± 1.124, and detection limit of 0.20 µg mL-1 and 0.24 µg mL-1 respectively. BMZ stability was inspected under various ICH forced degradation conditions and it was found to be easily degraded in acidic and alkaline conditions. The results revealed the suitability of the described methodology for the quantitation of the impurity (ABP) in a BMZ pure sample. The determination of BMZ in pharmaceutical dosage forms was conducted with the described method and showed mean percentage recovery of 99.39 ± 1.401 and 98.72 ± 1.795 (n = 6), respectively. When comparing the described procedure to a reference HPLC method statistically, no significant differences between the two methods in regard to both accuracy and precision were found.

Development and Validation of Two Novel Chromatographic Methods: HPTLC and HPLC for Determination of Bromhexine Hydrochloride in Presence of Its Two Impurities.

Two simple, sensitive and validated chromatographic methods were developed for quantitative determination of bromhexine hydrochloride (BHX) in presence of its major impurities, impurity B (IMB) and impurity C (IMC), as specified by British Pharmacopoeia. First method (I) was high-performance thin layer chromatography-densitometry at which the chromatographic separation was performed using silica gel plates and developing system consisted of hexane:acetone:ammonia solution (9:0.5:0.08, by volume) with ultraviolet scanning at 240 nm and linearity was achieved in the ranges of 0.40-10.00, 0.20-2.00 and 0.20-2.00 µg/band of BHX, IMB and IMC, respectively. Also, second chromatographic method (II) was high-performance liquid chromatography (HPLC) where the separation was carried out on C18 column at isocratic mode at flow rate 1.5 mL/min. The mobile phase consisted of methanol:water (90:10, v/v) adjusted to pH 2.5 with O-phosphoric acid and temperature was adjusted to 40°C. The scanning wavelength was 240 nm. The chromatographic run time was 6 min. Linearity of this method was achieved in the ranges of 4.00-40.00, 0.20-10.00 and 0.50-10.00 µg/mL for BHX, IMB and IMC, respectively. The validation of these chromatographic methods was made according to International Conference on Harmonization guidelines. These methods were successfully applied for determination of BHX in its pharmaceutical formulation. Also, statistical comparison was attained between the developed methods and the reported HPLC method using Student's t-test and F-test, and the obtained results showed that there was not any significant difference between them concerning with accuracy and precision.

Appraisal of the greenness profile of a chromatographic method for the simultaneous estimation of carbamazepine and oxcarbazepine, along with two potential impurities and three formulation excipients.

Structurally related carbamazepine (CBZ) and oxcarbazepine (OX) are two of the most commonly used antipsychotic drugs. The main impurities of CBZ, as described in both the USP and the BP, are iminodibenzyl (IMD) and iminostilbene (IST). Meanwhile, for non-pharmacopeial OX, the declared impurities include CBZ and IST. Prescribed oral suspensions of CBZ and OX contain additives including methyl paraben (MP), propyl paraben (PP) and sorbic acid (SA) as preservatives. An HPTLC method was introduced and developed for resolving the interference between CBZ, OX, their impurities, and the suspension additives in a single run, in addition to their quantitation with a high sensitivity that satisfies the USP requirements for the detection and quantitation of drug impurities. In the developed HPTLC method, CBZ and OX were measured in the range of 40-4000 ng per band, while IMD, IST, MP, PP and SA were in the range of 20-2000 ng per band, using a mixture of hexane : ethylacetate : formic acid : acetic acid (8 : 2 : 0.5 : 0.3, by volume) and UV scanning at 254 nm. The greenness profile of the method was evaluated by two different tools, the analytical Eco-Scale and the Green Analytical Procedure Index (GAPI), then a comparison between their results was conducted. This is the first time that the studied drugs, along with their impurities and suspension additives, were analyzed by a HPTLC method in a single run and within the limits required by the USP guidelines.

Validated spectrofluorometric determination of hypoglycemic combination, in pure form and pharmaceutical formulation using 9,10-phenanthraquinone reagent.

The aim of this work is to develop and validate a simple, rapid, highly sensitive and selective spectrofluorometric method for determination of Metformin HCl (MFH) and Glibenclamide (GLB) in their binary mixture without prior separation. The proposed method based on measuring the native fluorescence intensity of GLB at λemission = 348 nm, after excitation at λexcitation = 226 nm and measuring the fluorescence intensity of the fluorescent product produced from the derivatization of MFH using 9,10-phenanthraquinone in alkaline media at λemission = 416 nm after excitation at λexcitation = 240 nm. The proposed spectrofluorometric method allowed sensitive determination of the studied drugs with a limit of quantitation of 0.04 and 0.01 µg mL-1 for MFH and GLB, respectively, providing greater sensitivity than the reported one. Validation of the proposed method was carried out according to ICH guidelines with respect to linearity, accuracy, precision, and selectivity. The developed method was successfully applied for the determination of MFH and GLB in laboratory prepared mixtures and pharmaceutical formulation and the results obtained were statistically compared to those of the reported HPLC method with no significant difference between them.

Development and Validation of Stability Indicating High-Performance Liquid Chromatographic Method for Determination of Cyproheptadine Hydrochloride, its Impurity And Degradation Product.

This work presents a sensitive, accurate and selective RP-HPLC method for simultaneous determination of cyproheptadine HCl (CPH), its impurity B (dibenzosuberone) and CPH oxidative degradation product (10,11-dihydroxy-dibenzosuberone) in bulk powder and in pharmaceutical formulation. The RP-HPLC method depends on isocratic elution using C8 column and mobile phase consisting of 0.05 M KH2PO4 buffer:methanol (35:65, v/v, pH = 4.5) at a flow rate of 2 mL/min, and the eluant was monitored at 245 nm. Good resolution was obtained with tR values of 3.05, 7.54 and 6.17 min for CPH, impurity and oxidative degradate, respectively. The proposed method has been validated as per ICH guidelines using pure forms of CPH, its impurity and degradation product in pharmaceutical formulation with an accuracy of 100.48, 100.16 and 100.11, respectively. Additional spiking experiments yielded an accuracy of 100 ± 1.6%. Repeatability and intermediate precision results indicated acceptable low <2% RSD values. Moreover, the developed method's statistical results were favorably compared to the previously reported method results regarding both accuracy and precision. The developed method can be applied for analysis of the three components in quality control laboratories.

Evaluation of vinburnine in pharmaceuticals by smart spectrophotometric methods; full stability study.

Vinburnine (VNB) is a vinca alkaloid used as a vasodilator to enhance cerebral circulatory insufficiency. It is a cyclic amide containing drug which is expected to be sensitive to hydrolytic degradation. The degradation profile of VNB was studied in this work following ICH recommendations for stability study. The drug was sensitive only to degradation with NaOH with the formation of the carboxylic acid derivative, identified by IR and 1H NMR analyses as 2-((1S,12bS)-1-ethyl-1,2,3,4,6,7,12,12b-octahydroindolo[2,3-a] quinolizin-1-yl) acetic acid, (DEG). In this study five simple, smart and univariate stability indicating spectrophotometric methods were developed and validated for simultaneous determination of VNB and DEG for the first time. The developed methods include; Dual Wavelength Method (DWM), Dual Wavelength Resolution Method (DWRM), Factorized Absorbance Difference Method (FADM), Advanced Absorbance Subtraction Method (AASM), and Derivative Amplitude Factor Method (DAFM). These methods were capable of determination of VNB and DEG over the ranges of 1-30 and 3-50 µg/mL, respectively. The proposed methods were simple, smart, specific, and could be applied for analyzing synthetic mixtures of VNB and DEG and were successfully applied for determination of the drug in commercially available capsules. The obtained results of these methods were statistically compared with the reported HPLC one using student's-t and F- tests, where no significant difference was observed. Validation of the developed methods was applied according to ICH recommendations and all the results were within the acceptable limits.

The convenient use of fluorescamine for spectrofluorimetric quantitation of pramipexole in pure form and pharmaceutical formulation; application to content uniformity testing.

Pramipexole is a selective dopamine receptor agonist which is used in the treatment of Parkinson's disease and restless legs syndrome. The present work illustrates the development and validation of a sensitive and selective spectrofluorometric method for quantitation of pramipexole (PMP) through its interaction with fluorescamine at pH 7.5 using aqueous borate buffer to produce a highly fluorescent product. The fluorescent intensity of the formed product was measured at 480 nm after excitation at 391 nm. Experimental factors that could influence the formation, stability and the fluorescence intensity of the formed product were investigated and optimized. The linearity of the proposed method was achieved in the concentration range of 0.05-2.0 µg/mL. The quantitation and detection limits were 47 and 15 ng/mL, respectively. The proposed method has been validated in respect to guidelines of ICH and pharmaceutical tablets of PMP were successfully analyzed. Moreover, the method was applied for studying the content uniformity test according to the guidelines of United States Pharmacopeia.

Stability indicating spectrophotometric methods for quantitative determination of bromazepam and its degradation product.

Four simple, sensitive and selective stability indicating spectrophotometric methods are presented for quantitative determination of the benzodiazepine drug; bromazepam (BMZ) and one of its reported potential impurities and degradation product; 2-(2-amino-5-bromobenzoyl) pyridine (ABP) in methanol. Method A, is isoabsorptive point coupled with D0 method, where good linearity was obtained by measuring the absorbance of BMZ at 264 nm (Aiso) in the concentration range of 2-25 µg mL-1, and the absorbance of ABP at its λmax 396 nm in concentration range of 0.5-24 µg mL-1. Method B, is ratio subtraction; the absorbance was measured at 233 nm for BMZ using 20 µg mL-1 of ABP, while ABP was determined directly at its λmax 396 nm using methanol as a solvent. Method C, was based on measuring the total peak amplitude of the first derivative of the ratio spectra (DD1) of BMZ from 301 to 326 nm using 10 µg mL-1 of ABP as a divisor and determination of ABP at peak amplitude of 293 nm using 5 µg mL-1 of BMZ as a divisor. In method D, ratio difference method, good linearity was achieved for determination of BMZ and ABP by measuring the differences between the amplitudes of ratio spectra at 312 nm and 274 nm and differences between the amplitudes of ratio spectra at 274 nm and 312 nm, respectively. The stability of BMZ was investigated under different ICH recommended forced degradation conditions. The suggested methods were then successfully applied for determination of BMZ in its pharmaceutical formulations.

Development and validation of stability indicating chromatographic methods for simultaneous determination of citicoline and piracetam.

Citicoline and piracetam were subjected separately to different stress conditions as recommended by the international conference on harmonization. In addition, new stability indicating thin layer chromatographic and ultra high performance liquid chromatographic methods have been developed and validated for simultaneous determination of citicoline and piracetam in presence of their degradation products. Separation on the proposed thin layer chromatographic method was carried out using a developing system containing methanol:chloroform:ammonium chloride buffer (9:1:2, v/v/v) on silica gel plates at 230 nm. On the other hand, the mobile phase in the ultra high performance liquid chromatographic method was composed of water (containing 0.1% triethylamine):ethanol (92:8, v/v). The flow rate was 1 mL/min and ultraviolet detection was at 230 nm. Moreover, results of the developed methods were statistically compared to those obtained by the reported high-performance liquid chromatography method and no significant difference between them was found. The greenness profile of ultra high performance liquid chromatographic method was assessed and compared with those of the previously published high-performance liquid chromatography methods, it was noticed that the proposed ultra high performance liquid chromatographic method more environmentally friendly and greener than other methods.

Ecofriendly chromatographic methods for determination of co-prescribed drugs, olanzapine and metformin, in rat plasma.

Background: Olanzapine (OLZ) is one of most recommended drugs for the treatment of schizophrenia while metformin (MET) is the most commonly used hypoglycemic agent. Aim: Development and validation of two green, sensitive and accurate chromatographic methods for the simultaneous determination of OLZ along with the co-prescribed, MET. Materials & methods: TLC-densitometric method with a developing system consisting of methylene chloride:methanol:ethyl acetate:triethylamine (4:4:5:0.1, by volume) and a reversed-phase (RP)-HPLC method where the chromatographic separation was performed using ethanol:water mixture (50: 50, v/v) as a mobile phase. Results: TLC-densitometric method had linearity over concentration ranges of 160-4000 ng/band for OLZ and 150-4500 ng/band for MET, while RP-HPLC method was linear and validated over concentration range of 300-20000 ng/ml for OLZ and MET. Conclusion: Pharmacokinetic study was successfully performed and suggested the possibility of co-administration of MET with OLZ and their further formulation in one pharmaceutical preparation to enhance patient's compliance.

Spectrofluorimetric approach for determination of citicoline in the presence of co-formulated piracetam through fluorescence quenching of eosin Y.

A simple, sensitive, and precise spectrofluorimetric method has been developed and validated for quantitation of citicoline in its pharmaceutical formulations. The proposed method based on quantitative quenching effect of citicoline on the native fluorescence of Eosin Y via developing of a binary complex reaction between the cited drug and Eosin Y in acidic medium using acetate buffer pH = 3.6. The quenching of the fluorescence of eosin was measured at 540 nm after excitation at 518 nm. Calibration graph was achieved in the range of 300-3000 ng/mL with 0.9996 as correlation coefficient and 291.0 and 93.86 ng/mL as quantitation and detection limits, respectively. The developed method considered as the first developed spectrofluorimetric one for quantitation of citicoline with high sensitivity and validated according to ICH guidelines. The selectivity of the proposed method was investigated by studying the interference of piracetam as co-formulated drug with CIT in pharmaceutical formulation, therefore the developed method could be used for routine quality control of citicoline in its pharmaceutical formulations either alone or in combination with piracetam.