Three techniques for the determination of perindopril through derivatization with 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole.

In addition to its pure form, three accurate, rapid, and simple methods have been established for determining perindopril (PRD) in its tablet form. At pH 9.0 using a borate buffer, developing the three designated methods was successful according to the reaction between PRD and 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) and the formation of a chromogen (with a yellow color) measurable at 460 nm using the spectrophotometric method (Method I). In addition, the produced chromogen was assessed using the spectrofluorimetric method (Method II) at 535 nm following excitation at 461 nm. Afterward, the same reaction product was separated and determined using the HPLC method with fluorescence detection (Method III). A Promosil C18 stainless steel column (Q7 5 mm particle size, 250-4.6 mm) has proven suitable for separation. The mobile phase adjustment was made at pH 3.0, with a 1.0 mL min -1 flow rate; its composition was methanol-sodium dihydrogen phosphate, 0.02 M (60: 40, v/v). Through concentration ranges of 5.0-60.0, 0.5-6.0, and 1.0-10.0 µg mL-1, the calibration curves were rectilinear for Methods I, II, and III, respectively, with limits of quantification (LOQ) of 1.08, 0.16 and 0.19 µg mL-1 as well as limits of detection (LOD) of 0.36, 0.05 and 0.06 µg mL-1. The developed methods were implemented to estimate PRD in tablets, and a comparison between the obtained outcomes utilizing the developed methods as well as obtained from the official method revealed that they were comparable. The official BP method was based on dissolving PRD in anhydrous acetic acid and titrating with 0.1 M perchloric acid, then the potentiometric determination of the end-point. The designated methods were also implemented in content uniformity testing with satisfying results. The reaction pathway proposal was speculated, and according to ICH Guidelines, the statistical evaluation of the data was performed. The three proposed methods were confirmed to be green, eco-friendly and safe to environment using Green Analytical procedure index (GAPI) method.

Concurrent estimation of some co-administered antimicrobial drugs applying conventional and first derivative synchronous fluorescence spectroscopy techniques.

For the estimation of some co-administered antimicrobials, two highly accurate and precise spectrofluorimetric methods were developed. Fluconazole (FLZ) is co-administered with either ciprofloxacin (CPR) or ofloxacin (OFX) for the treatment of certain microbial infections. On the other hand, another antimicrobial drug, vancomycin (VNC) is co-administered with ciprofloxacin (CPR) for peritonitis treatment. In method I, conventional spectrofluorimetry has been introduced for the concurrent quantitative estimation of FLZ in presence of OFX or CPR. While in method II, a first derivative synchronous spectrofluorimetric technique was adapted for quantitation of VNC and CPR co-administered combination. Both of them were utilized for estimation of the considered drugs in raw materials, laboratory prepared mixtures, dosage forms, and biological fluids. Method I was relied on simultaneous measuring of the native fluorescence of FLZ and OFX or CPR without any overlapping between the emission spectra of each binary mixture (FLZ / OFX) and (FLZ / CPR). Fluorescence intensities were measured at 283.0, 483.0 and 436.0 nm after excitation at 262.0, 292.0 and 275.0 nm for FLZ, OFX and CPR, respectively. Method II was utilized the synchronous fluorescence intensity of VNC and CPR in methanol at Δλ = 40 nm. The first derivative synchronous spectra were calibrated at 297.0 nm for VNC and at 379.5 nm for CPR. Different variables influencing conventional and synchronous fluorescence intensities of the four antimicrobials under investigation were precisely optimized. Both methods were successfully investigated for the determination of the studied drugs in plasma. The linear data analysis for the calibration curves reveals a good relationship in the ranges of 1.0-10.0, 0.25-2.5 and 0.06-0.6 µg/mL for FLZ, OFX and CPR for method I with limits of detection 0.144, 0.038 and 0.007 µg/mL and limits of quantitation of 0.437, 0.114 and 0.021 µg/mL for FLZ, OFX and CPR, respectively. Linearity range for method II was 0.5 -10.0 µg/mL for VNC and CPR with detection limits of 0.127 and 0.110 µg/mL and quantitation limits of 0.380 and 0.334 µg/mL for VNC and CPR, respectively. International Council on Harmonization ICH Q2 (R1) Guidelines were followed in the developed methods validation. The achieved outcomes were statistically compared with those found by the reported ones, and no significant difference was observed.

Eco-friendly fluorimetric approaches for the simultaneous estimation of the co-administered ternary mixture: etoposide, moxifloxacin and nalbuphine.

Antineoplastic drugs, etoposide (ETO), are widely used in leukaemia. A patient with leukaemia has a relative infection with pneumonia treated by fluoroquinolones as moxifloxacin HCL (MOX). Because opioid analgesic as nalbuphine HCL (NAL) does not have a ceiling dose, it is used to manage the distasteful sensory in leukaemia. Consequently, green methods for synchronous spectrofluorimetric quantification of a ternary mixture of ETO, MOX and NAL were developed. The first approach relies simply on the estimation of MOX at 371 nm by conventional synchronous fluorimetric technique (Δλ of 60 nm). The second approach depends on applying the first derivative synchronous fluorimetric technique (Δλ of 60 nm) for simultaneous estimation of ETO and NAL at 257 and 273 nm, respectively. A good linear correlation was obtained in the ranges of 0.04-0.40, 0.10-1.00 and 0.50-5.00 µg ml-1 for MOX, ETO and NAL, respectively. Moreover, the proposed approaches were successfully applied for the estimation of the studied drugs in the pharmaceutical dosage forms. Additionally, the synchronous assessment of ETO, MOX and NAL in the spiked human urine was successfully attained by the facile protein precipitation technique. The mean % recoveries in spiked human urine were 99.49, 98.07 and 98.48 for MOX, ETO and NAL, respectively.

Green and sensitive spectrofluorimetric method for the determination of two cephalosporins in dosage forms.

Using two green and sensitive spectrofluorimetric methods, we quantified two cephalosporins, cefepime (CFM) and cefazolin (CFZ), in raw and pharmaceutical formulations. The first method is based on the reaction between CFM and fluorescamine (borate buffer, pH 8), which yields a highly fluorescent product. After excitation at 384 nm, the fluorescent product emits light at 484 nm. At concentration ranges from 12.0 to 120.0 ng ml-1, the relative fluorescence intensity/concentration curve was linear with a limit of quantification (LOQ) of 2.46 ng ml-1. The second method relied on measuring the CFZ quenching action on acriflavine fluorescence through formation of an ion-associate complex using Britton-Robinson buffer at pH 8. We measured acriflavine fluorescence at 505 nm after excitation at 265 nm. The decrease in acriflavine fluorescence intensity was CFZ concentration-dependent. Using this method, we quantified CFZ in concentrations ranging from 1 to 10 µg ml-1 with a LOQ of 0.48 µg ml-1. We studied and optimized the factors influencing reaction product formation. Moreover, we adapted our methods to the investigation of the mentioned drugs in raw and pharmaceutical formulations with greatly satisfying results. We statistically validated our methods according to International Council on Harmonisation Guidelines. The obtained results were consistent with those obtained with the official high-performance liquid chromatography methods.

Insights for applying erythrosine B as a green fluorescence probe for estimation of anticancer tamoxifen and its analog; clomiphene in nanogram concentration.

The growth of tumor tissue is extremely pervasive among post-menopausal women. Commonly, from the clinical application, adjuvant selective estrogen receptor modulators such as tamoxifen are prescribed for prohibiting metastatic breast cancer, while its analog, clomiphene, is used to treat infertility in women. Lately, the significance of green chemistry on our environment was through reducing the influence of hazardous chemicals. Consequently, efforts were screened to perform a fast and simple eco-friendly green method for the determination of two aromatase inhibitors. In this study, a sensitive green spectrofluorimetric approach was developed to detect and characterize tamoxifen citrate (TAM) and clomiphene citrate (CLO) via complex formation with erythrosine B. The reaction between erythrosine B dye (EB) and the two aromatase inhibitors results in quenching the fluorescence activity of the dye by the formation of ion-pair in Britton-Robinson buffer (BRB) solution (pH 4.3) at 554 nm (λex = 527 nm). The approach outcome confirmed that the solvent's inherent nature has a critical impact on the approachs' sensitivity and reproducibility. An approved linear correlation was achieved between the reduction in the emission value of EB's fluorescence and the concentration in the ranges of 40.0-600.0 ng/mL for both TAM and CLO with mean % recoveries 100.20 ± 0.93 and 100.07 ± 1.09, respectively. The approach was validated regarding ICH protocols, and the outcomes were acceptable. The changes in Gibb's free energy (ΔG°) by the obtained ion-pair between EB and TAM or CLO were -36.65 or -37.03 kJ mol-1, respectively, which indicates the reaction feasibility at ambient temperature. Commercial dosage forms for TAM and CLO were simply analyzed, and good recoveries were achieved within the range. The National Environmental Methods Index, Analytical Eco-Scale, and Green Analytical Procedure Index applications to our illustrated approach present additional eligibility to this study.

Simultaneous estimation of troxerutin and calcium dobesilate in presence of the carcinogenic hydroquinone using green spectrofluorimetric method.

In the present study, we conducted two facile and highly sensitive spectrofluorimetric approaches in order to quantify the vasoprotective agents; troxerutin (TROX) and calcium dobesilate (DOB) in the presence of hydroquinone (HQ) (as a highly toxic impurity and potential degradation product of DOB) in commercial formulations and human plasma. The first approach relies simply on using ethanol as an eco-friendly solvent for the estimation of DOB at 345 nm after being excited at 305 nm. The linearity was carefully investigated between DOB concentration and the relative fluorescence intensity in the range of 0.05-0.8 µg ml-1. Due to the high method simplicity and sensitivity, applying the first approach to quality control analysis and spiked human plasma samples with mean % recoveries 100.74 ± 3.71 adds another merit. The second approach involved rapid conventional fluorimetric estimation of ethanolic TROX solution in TROX/DOB combined dosage forms at 455/350 nm (emission/excitation) with a linear calibration chart covering the range of 0.1-1.2 µg ml-1. Moreover, the second approach involved a comprehensive study in a trial to solve the problem of superposition of DOB and HQ graph adopting the first derivative synchronous fluorimetric mechanism in ethanol at Δλ = 60 nm. Therefore, DOB was measured at 286 and 323 nm, while HQ could be quantitated at 301 nm. The Beer-Lambert Law has complied over the ranges of 0.1-1.0 and 0.02-0.4 µg ml-1 for DOB and HQ, respectively. Guidelines adopted by the International Council of Harmonization (ICH) were used to validate the target approaches. The developed methods are more convenient for routine quality control laboratory instead of the time-consuming and sophisticated reported techniques. Moreover, different aspects of evaluating the greenness of the proposed approaches were conducted to have a complete image of their environmental impact.

Investigation of some univariate and multivariate spectrophotometric methods for concurrent estimation of Vancomycin and Ciprofloxacin in their laboratory prepared mixture and application to biological fluids.

Four simple, rapid, accurate and precise spectrophotometric methods were established and validated in accordance with ICH Q2 (R1) guidelines for the simultaneous determination of Vancomycin (VNC) and Ciprofloxacin (CPR) in their raw materials, laboratory prepared mixtures and pharmaceutics. Method A depends on using first derivative spectrophotometry (D1) where VNC and CPR were resolved at 243.6 and 262.0 nm, respectively. Concerning method B, it is based on utilizing first derivative of ratio spectra (DD1) where determination was performed at the peak maxima at 244.0 nm and 258.0 nm for VNC and CPR, respectively. Two chemometric models were applied for the quantitative analysis of both drugs in their laboratory prepared mixtures, namely, partial least squares (PLS) (method C) and artificial neural network (ANN) (method D). For univariate methods linearity range for both drugs was in the range of 3-30 and 1-10 µg/mL for VNC and CPR, respectively. Multivariate calibration methods using five level, two factor calibration model for the development of 25 mixtures were also applied for the simultaneous estimation of the two drugs in their laboratory prepared mixture using spectral region from 200.0 to 300.0 nm using interval 1 nm. The suggested methods have been successfully extended to the assay of the two studied drugs in laboratory-prepared mixtures and pharmaceuticals with excellent recovery. First derivative spectrophotometry (D1) was also applied for the assay of both analytes in spiked human plasma with good recovery. No interaction with common pharmaceutical additives has been observed which indicate the selectivity of the method. The results obtained were favourably compared with those obtained applying the reported methods. The methods are validated in compliance with the ICH Q2 (R1) guidelines and the measured accuracy and precision are assessed to be within the accepted limits.

Two liquid chromatographic approaches for the simultaneous determination of xipamide and its degradation product (2,6-xylidine) using time-programmed fluorescence detection.

A study of the performance of reversed-phase chromatography with a programmable multiwavelength fluorimetric technique using either conventional hydro-organic or micellar eluent is established for the determination of xipamide (XIP) in the presence of its degradation product, 2,6-xylidine (XY). In conventional liquid chromatography (CLC), the analyses were carried out on a Promosil ODS 100 Å column (250 mm × 4.6 mm i.d., 5 µm) using a mobile phase consisting of methanol/0.1 M phosphate buffer (65: 35, v/v) at pH 4.0. For micellar liquid chromatography (MLC), a short Spherisorb column (150 mm × 4.6 mm i.d., 5 µm) was employed in conjunction with a greener mobile phase (pH 5.0) containing 0.1 M sodium dodecyl sulfate and 15% n-propanol. CLC proved to be superior to MLC in terms of sensitivity for the determination of the degradation product because it could detect trace amounts down to 10.0 ng/ml of XY as a degradation product in XIP. However, MLC represents an eco-friendly approach for the simultaneous determination of XIP and XY. In addition, the opportunity for the direct introduction of biological matrices into the chromatographic system is one of the distinctive benefits of MLC. The proposed methods were applied for the determination of XIP in its tablets, human urine and content uniformity testing. The results of the proposed methods were statistically compared with those obtained using the comparison fluorimetric method, revealing no significant differences in the performance of the methods regarding accuracy and precision.

Determination of adapalene in gel formulation by conventional and derivative synchronous fluorimetric approaches. Application to stability studies and in vitro diffusion test.

BACKGROUND: Adapalene is a retinoid analogue with actions similar to those of tretinoin. It is used in topical treatment of mild to moderate acne. A survey of the literature reveals that no spectrofluorimetric method has been reported yet for determination of ADP, so it was thought necessary to develop a highly sensitive stability indicating spectrofluorimetric method. RESULTS: Two highly sensitive spectrofluorimetric approaches were conducted for the assay of adapalene (ADP) in its gel. In the first approach, ADP exhibits an intense native fluorescence at 389 nm after excitation at 312 nm using borate buffer (pH 7.0)/ethanol system. This approach was successfully applied for routine analysis of ADP in its gel and ideally suited to the in vitro diffusion test. To elucidate the inherent stability of ADP, bulk sample was subjected to different stress conditions as specified by ICH guidelines. The acidic and oxidative degradation products were resolved from the intact drug using second and first derivative synchronous fluorimetry at 346 and 312.45 nm, respectively (the second approach). The synchronous fluorescence was scanned at Δ λ of 80 nm in case of acidic degradation and at Δ λ of 100 nm in case of oxidative degradation. Good linearity was obtained for ADP over the range 2.0-14.0 ng/mL with good correlation coefficient   0.999 in each approach. The approaches were carefully examined in terms of linearity, accuracy and precision. They were suitable for routine quality control laboratory. Moreover, the stability-indicating power of the second approach was ascertained via forced degradation studies. CONCLUSIONS: The proposed approaches were validated and successfully applied for the quantitative assay of a small concentration of ADP in its pharmaceutical gel. The conventional spectrofluorimetry was ideally suited for in vitro diffusion test. Stability studies were also conducted using different forced degradation condition according to ICH recommendation.Graphical abstractSimultaneous determination of ADP and its degradation products.

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.

Micelle-enhanced spectrofluorimetric method for determination of lacidipine in tablet form; application to content uniformity testing.

A highly sensitive, simple and rapid spectrofluorimetric method was developed for the determination of lacidipine (LCP) in tablets. The proposed method is based on the investigation of the fluorescence spectral behavior of LCP in both sodium dodecyl sulphate (SDS) and the tween-80 micellar system. In aqueous solutions of acetate buffer (pH 4.5), the fluorescence intensities of LCP were greatly enhanced (ca. 2.4 and 4.3 folds) in the presence of either SDS or tween-80, respectively. The fluorescence intensity was measured at 444 nm after excitation at 277 nm using either SDS or tween-80 as a surfactant. The fluorescence-concentration plots were rectilinear over the ranges of 50.0-500.0 ng/ml and 5.0-200.0 ng/ml with lower detection limits of 5.11 and 2.06 ng/ml and lower quantification limits of 17 and 6.87 ng/ml using SDS and tween-80, respectively. The method was successfully applied to the analysis of LCP in commercial tablets and the results were in good agreement with those obtained with the comparison method. Furthermore, content uniformity testing of pharmaceutical tablets was also conducted.

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.

Simultaneous determination of some dihydroxybenzenesulphonic acid derivatives and their degradation product and main impurity (hydroquinone) by ion-pair liquid chromatography.

A simple and sensitive high-performance liquid chromatography method was developed and validated for the determination of calcium dobesilate (DOB) or ethamsylate (ETM) in the presence of their degradation product, hydroquinone (HQ). The analyses were carried out on Promosil C18 column (4.6 mM × 250 mM, 5 µm particle size) using an ion-pair mobile phase consisting of methanol-1.5 mM tetra-butyl ammonium bromide in 0.06 M phosphate buffer (25 : 75, v/v) at pH 6.0 with fluorescence detection at 286/333 nm. Pindolol was used as an internal standard. The proposed method was found to be rectilinear over the concentration ranges of 0.05-0.5 µg/mL for DOB, 0.1-0.8 µg/mL for ETM and 0.005-0.1 µg/mL for HQ. The method was applied for the determination of the studied drugs in different dosage forms and biological fluids. The results of the proposed method were statistically compared with those obtained by the comparison methods revealing no significance differences in the performance of the methods regarding accuracy and precision. Moreover, applying a time-programmed fluorescence technique was valuable for the detection of trace amounts of HQ as an impurity and allowed purity testing of ETM or DOB within the BP pharmacopeial limit (0.1%).

Micelle-enhanced spectrofluorimetric method for determination of cyproheptadine hydrochloride in tablets: application to in-vitro drug release and content uniformity test.

A highly sensitive and simple spectrofluorimetric method was developed for the determination of cyproheptadine hydrochloride (CYP) in its pharmaceutical formulations. The proposed method is based on the investigation of the fluorescence spectral behaviour of CYP in a sodium dodecyl sulphate (SDS) micellar system. In aqueous solution, the fluorescence intensity of CYP was greatly enhanced (150 %) in the presence of SDS. The fluorescence intensity was measured at 410 nm after excitation at 280 nm. The fluorescence-concentration plot was rectilinear over the range 0.2-2.0 µg/mL, with lower detection limit of 0.06 µg/mL. The proposed method was successfully applied to the assay of commercial tablets as well as content uniformity testing. The application of the proposed method was extended to test the in-vitro drug release of CYP tablets, according to USP guidelines. The results were statistically compared with those obtained by official USP method and were found to be in good agreement.

Kinetic determination of acarbose and miglitol in bulk and pharmaceutical formulations using alkaline potassium permanganate.

A simple and sensitive kinetic spectrophotometric method was established for the determination of acarbose and miglitol in bulk and in their pharmaceutical preparations using alkaline potassium permanganate as an oxidizing agent. The method involves determination of acarbose and miglitol by kinetic studies of their oxidation at room temperature for a fixed time of 15 minutes for acarbose and 25 minutes for miglitol. The absorbance of the colored manganate ion was measured at 610 nm. Alternatively, the kinetic decrease in the absorbance of permanganate upon addition of the studied drugs at 525 nm was also used. The absorbance concentration plot was rectilinear over the concentration range of 4-20 and 1-10 µg/ml for acarbose and miglitol, respectively. The detection limits were 0.189 and 0.089 µg/ml at 610 nm and 0.081 and 0.179 µg/ml at 525 nm for acarbose and miglitol respectively. The method was successfully applied for the determination of these drugs in their dosage forms. The results obtained were in good agreement with those obtained with the reference methods.