Highly sensitive HPLC method for the determination of galantamine in human plasma and urine through derivatization with dansyl chloride using fluorescence detector

A new method based on fluorescence derivatization with 5‐(dimethylamino) naphthalene‐1‐sulfonyl chloride (dansyl chloride) was developed for the quantitative determination of galantamine in human plasma and urine using high‐performance liquid chromatography. The reaction between galantamine and dansyl chloride was optimally realized in 30 min at room temperature and pH 10.5, with a reagent to galantamine molar ratio of 2.13. The derivative was extracted with dichloromethane, and the extract was dried under a nitrogen stream and dissolved in the mobile phase. Chromatographic analysis was performed with an Inertsil C₁₈ column and a mobile phase comprising 40% acetonitrile and 60% 10 mM o‐phosphoric acid, 1.2 ml/min. The injection volume was 20 μl. The derivatives were detected with a fluorescence detector (excitation 375 nm/emission 537 nm). The retention time for the dansyl derivative of galantamine was 16.8 min. Linearity was observed between 125 and 2000 ng/ml in water, urine and plasma. The limit of detection and limit of quantification for the developed method were 6.27–70.99 and 18.81–212.97 ng/ml, respectively. Per cent recovery was calculated as 95.15 for urine and 95.78 for plasma. Interday repeatability values for urine and plasma samples (n = 6) at three different concentrations were calculated as a per cent relative standard deviation of 0.24–0.59 and 0.35–0.56. The corresponding per cent relative standard deviation values for intraday repeatability were 0.13–0.51 and 0.04–0.15, respectively.     

Validated spectrofluorimetric method for the determination of tamsulosin in spiked human urine,pure and pharmaceutical preparations

A novel, sensitive and selective spectrofluorimetric method was developed for the determination of tamsulosin in spiked human urine and pharmaceutical preparations. The proposed method is based on the reaction of tamsulosin with 1‐dimethylaminonaphthalene‐5‐sulfonyl chloride in carbonate buffer pH 10.5 to yield a highly fluorescent derivative. The described method was validated and the analytical parameters of linearity, limit of detection (LOD), limit of quantification (LOQ), accuracy, precision, recovery and robustness were evaluated. The proposed method showed a linear dependence of the fluorescence intensity on drug concentration over the range 1.22 × 10^‐7 to 7.35 × 10^‐6 M. LOD and LOQ were calculated as 1.07 × 10^‐7 and 3.23 × 10^‐7 M, respectively. The proposed method was successfully applied for the determination of tamsulosin in pharmaceutical preparations and the obtained results were in good agreement with those obtained using the reference method. Copyright © 2013 John Wiley & Sons, Ltd.     

Development and validation of a sensitive spectrofluorimetric method for the determination of amoxapine in human plasma and urine

A selective and sensitive spectrofluorimetric method was developed and validated for the determination of amoxapine in human plasma and urine. The developed method is based on labeling with 5‐dimethylaminonaphthalene‐1‐sulfonyl chloride (dansyl chloride) and monitoring at 397 nm (excitation)/514 nm (emission). The method was validated for linearity, limit of detection (LOD), limit of quantification (LOQ), precision, accuracy, recovery and robustness. The calibration curves were linear over a concentration range of 250–2500 and 50–1250 ng/mL for plasma and urine, respectively. The LOD values were calculated to be 13.31 and 13.17 ng/mL for plasma and urine, respectively. The proposed method was applied to study of amoxapine in human plasma and urine. Copyright © 2013 John Wiley & Sons, Ltd.     

Spectrofluorimetric determination of nomifensine in human plasma and urine by derivatization with fluorescamine

A sensitive, simple and rapid spectrofluorimetric method was developed for the determination of nomifensine in human plasma and urine. The present method was based on the derivatization by fluorescamine in phosphate buffer at pH 4.0 to produce a highly fluorescent product which was measured at 488 nm (excitation at 339 nm). The method was validated according to the ICH guidelines with respect to linearity, limit of detection, limit of quantification, accuracy, precision, recovery and robustness. The assay was linear over the concentration ranges 100–2,000 and 50–2,000 ng/mL for plasma and urine, respectively. The limits of detection were calculated to be 13.9 and 7.5 ng/mL for plasma and urine, respectively. The method was successfully applied to the analysis of the drug in human plasma and urine. Copyright © 2011 John Wiley & Sons, Ltd.     

Spectrofluorimetric determination of tobramycin in human serum and pharmaceutical preparations by derivatization with fluorescamine

A simple, sensitive and selective spectrofluorimetric method has been developed for the determination of tobramycin (TOB) in human serum and pharmaceutical preparations. The method is based on the reaction between the primary amino group of TOB and fluorescamine in borate buffer, pH 8.5, to give a highly fluorescent derivative which is measured at 469 nm after excitation at 388 nm. The fluorescence intensity was directly proportional to the concentration over the range 300–1500 ng/mL, with a limit of detection of 65 ng/mL and limit of quantitation of 215 ng/mL. All variables were investigated to optimize the reaction conditions. The method was validated according to International Conference on Harmonization guidelines in terms of specificity, linearity, limit of detection, limit of quantification, accuracy, precision and robustness. Good recoveries were obtained ranging from 97.4 to 100.64%, indicating that no interference was observed from concomitants usually present in pharmaceutical dosage forms. The method was successfully, applied for the analysis of the drug substance in its pharmaceutical preparations and spiked serum samples. Copyright © 2013 John Wiley & Sons, Ltd.     

Spectrofluorimetric method for the determination of sulpiride in pharmaceutical preparations and human plasma through derivatization with 2‐cyanoacetamide

A sensitive and accurate spectrofluorimetric method has been developed for the determination of sulpiride in pharmaceutical preparations and human plasma. The developed method is based on the derivatization reaction of 2‐cyanoacetamide with sulpiride in 30% ammonical solution. The fluorescent derivatized reaction product exhibited maximum fluorescence intensity at 379 nm after excitation at 330 nm. The optimum conditions for derivatization reactions were studied and the fluorescence intensity versus concentration plot was found to be linear over the concentration range 0.2–20.0 µg/mL with a correlation coefficient of 0.9985. The limit of detection (LOD) and limit of quantification (LOQ) were found to be 0.82 and 2.73 ng/mL, respectively. The proposed method was validated according to ICH guidelines. The effects of common excipients and co‐administered drugs were also studied. The accuracy of the method was checked using the standard addition method and percent recoveries were found to be in the range of 99.00–101.25% for pharmaceutical preparations and 97.00–97.80% for spiked human plasma. The method was successfully applied to commercial formulations and the results obtained for the proposed method were compared with a high‐performance liquid chromatography reference method and statistically evaluated using the Student's t‐test for accuracy and the variance ratio F‐test for precision. A reaction pathway was also proposed. Copyright © 2012 John Wiley & Sons, Ltd.     

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. Copyright © 2012 John Wiley & Sons, Ltd.     

Validated spectrofluorimetric method for the determination of clonazepam in pharmaceutical preparations

A simple, highly sensitive and validated spectrofluorimetric method was applied in the determination of clonazepam (CLZ). The method is based on reduction of the nitro group of clonazepam with zinc/CaCl₂, and the product is then reacted with 2‐cyanoacetamide (2‐CNA) in the presence of ammonia (25%) yielding a highly fluorescent product. The produced fluorophore exhibits strong fluorescence intensity at ?_em = 383 nm after excitation at ?_ex = 333 nm. The method was rectilinear over a concentration range of 0.1–0.5 ng/mL with a limit of detection (LOD) of 0.0057 ng/mL and a limit of quantification (LOQ) of 0.017 ng/mL. The method was fully validated and successfully applied to the determination of CLZ in its tablets with a mean percentage recovery of 100.10 ± 0.75%. Method validation according to ICH Guidelines was evaluated. Statistical analysis of the results obtained using the proposed method was successfully compared with those obtained using a reference method, and there was no significance difference between the two methods in terms of accuracy and precision. Copyright © 2015 John Wiley & Sons, Ltd.     

A novel spectrofluorimetric method for the determination of calcitonin in ampules through derivatization with fluorescamine

In this study, a simple, sensitive and selective spectroflourimetric method has been developed for the determination of salmon calcitonin (sCT) in ampules. The method is based on the reaction between sCT and fluorescamine at pH 8.5 in borate buffer, resulting in a highly fluorescent derivative. Fluorescence of derivatized sCT solutions was measured by setting the excitation and emission monochromators and slit widths to 390, 484 and 10 nm, respectively. Sevaral derivatization parameters were optimized. A calibration graph was constructed using standard solutions of the derivatized calcitonin in the range 0.5–6.0 µg/mL. Limit of detection and limit of quantification values were determined to be 0.124 and 0.372 µg/mL, respectively. The proposed method was successfully applied for the determination of sCT in commercially available ampules. High recovery values (101.0–102.0 %), and a low relative standard deviation (RSD %) value (5.3–5.4) proved the accuracy and precision of the proposed method. An isocratic reversed‐phase high‐performance liquid chromatographic (HPLC) method, as a reference, was also developed for the determination of sCT. A reversed‐phase Nucleosil® C₁₈ column (250 mm × 4.6 mm i.d., 10 µm particle size, 120 Å pore size) was used and the detector was set at 210 nm. Statistical comparison of the results of the two methods showed clearly that there was no significant difference between them. Copyright © 2012 John Wiley & Sons, Ltd.     

CdS nanoparticles‐ enhanced chemiluminescence and determination of baicalin in pharmaceutical preparations

CdS nanoparticles (CdS NPs) of different sizes were synthesized by the citrate reduction method. It was found that CdS NPs could enhance the chemiluminescence (CL) of the luminol‐potassium ferricyanide system and baicalin could inhibit CdS NPs‐enhanced luminol‐potassium ferricyanide CL signals in alkaline solution. Based on this inhibition, a flow‐injection CL method was established for determination of baicalin in pharmaceutical preparations and human urine samples. Under optimized conditions, the linear range for determination of baicalin was 5.0 x 10^?6 to 1.0 x 10^?3 g/L. The detection limit at a signal‐to‐noise ratio of 3 was 1.7 x 10 ^?6 g/L. CL spectra, UV‐visible spectra and transmission electron microscopy (TEM) were used to investigate the CL mechanism. The method described is simple, selective and obviates the need of extensive sample pretreatment. Copyright © 2012 John Wiley & Sons, Ltd.     

Utility of Von Pechman synthesis of coumarin reaction for development of spectrofluorimetric method for quantitation of salmeterol xinafoate in pharmaceutical preparations and human plasma

Simple, precise and selective spectrofluorimetric technique was evolved for quantitation of selective β₂ agonist drug namely salmeterol xinafoate (SAL). Utilizing its phenolic nature, a method was described based on the reaction of the studied drug with ethyl acetoacetate (EAA) to yield extremely fluorescent coumarin product which can be detected at 480 nm (λ_ex = 420 nm). The procedure obeys Beer's law with a correlation coefficient of r = 0.9999 in the concentration range between 500 and 5000 ng ml^?1 with and 177 ng ml^?1 for limit of detection (LOD) and limit of quantification (LOQ), respectively. Diverse reaction variables influencing the firmness and formation of the coumarin product were accurately examined and modified to ensure greatest sensitivity of the procedure. The proposed technique was performed and examined according to the US Food and Drug Administration (FDA) guidelines for bio‐analytical methods and was efficiently applied for quantitation of SAL in both pharmaceutical preparations (% recovery = 100.06 ± 1.07) and spiked human plasma (% recovery = 96.64–97.14 ± 1.01–1.52).     

Validated spectrofluorimetric method for determination of two phosphodiesterase inhibitors tadalafil and vardenafil in pharmaceutical preparations and spiked human plasma

A valid, sensitive and rapid spectrofluorimetric method has been developed and validated for determination of both tadalafil (TAD) and vardenafil (VAR) either in their pure form, in their tablet dosage forms or spiked in human plasma. This method is based on measurement of the native fluorescence of both drugs in acetonitrile at λ_em 330 and 470 nm after excitation at 280 and 275 nm for tadalafil and vardenafil, respectively. Linear relationships were obtained over the concentration range 4–40 and 10–250 ng/mL with a minimum detection of 1 and 3 ng/mL for tadalafil and vardenafil, respectively. Various experimental parameters affecting the fluorescence intensity were carefully studied and optimized. The developed method was applied successfully for the determination of tadalafil and vardenafil in bulk drugs and tablet dosage forms. Moreover, the high sensitivity of the proposed method permitted their determination in spiked human plasma. The developed method was validated in terms of specificity, linearity, lower limit of quantification (LOQ), lower limit of detection (LOD), precision and accuracy. The mean recoveries of the analytes in pharmaceutical preparations were in agreement with those obtained from the comparison methods, as revealed by statistical analysis of the obtained results using Student's t‐test and the variance ratio F‐test. Copyright © 2015 John Wiley & Sons, Ltd.     

A novel spectrofluorimetric method for determination of imatinib in pure,pharmaceutical preparation,human plasma,and human urine

The following paper represents a simple, highly sensitive, responsive validated and developed spectrofluorimetric method for estimation of imatinib (IMB) in its pure, commercial preparation, human urine and human blood plasma. The calibration curve was in the range 4–900 ng ml^?1 for pure form and urine and 8–900 ng ml^?1 for plasma in a medium contains carboxymethyl cellulose (CMC) and acetate buffer (pH 5) with excitation wavelength (λ_ex) 230 nm and emission wavelength (λ_em) 307 nm. The limit of detection (LOD) was 0.37 ng ml^?1 for the pure form, 0.64 ng ml^?1 for human urine, and 0.70 ng ml^?1 for human plasma, while the limit of quantitation (LOQ) was 1.2 for pure form, 1.91 for urine and 2.1 for plasma. The suggested method was successfully applied for evaluation of IMB in tablets within 99% mean percentage recovery. The excipients that are usually used as additives in pharmaceutical dosage form did not interfere with the suggested method. The method was efficiently used for estimation of IMB in human urine and human plasma. The effect of some cations that might be present in urine and plasma was also studied. The method was also focused on human volunteers and in vitro drug release.     

Synchronous spectrofluorometric methods for simultaneous determination of diphenhydramine and ibuprofen or phenylephrine in combined pharmaceutical preparations

Simple and rapid synchronous fluorometric methods were adopted and validated for the simultaneous analysis of a binary mixture of diphenhydramine (DIP) and ibuprofen (IBU) ( Mix I) or DIP and phenylephrine (PHE) (Mix II) in their co‐formulated pharmaceuticals without prior separation. Analysis of Mix I is based on the measurement of the peak amplitudes (D¹) of synchronous fluorescence intensities at 265.1 nm for DIP and 260 nm for IBU. The relationship between the concentration and the amplitude of the first‐derivative synchronous fluorescence spectra showed good linearity over the concentration ranges 0.50–10.00 μg ml^?1 and 0.50–7.90 μg ml^?1 for DIP and IBU, respectively. Analysis of Mix II was based on measurement of the peak amplitude (D¹) synchronous fluorescence intensities at 230 nm for DIP and at 253.9 nm for PHE. Moreover, for Mix II, the peak amplitude (D²) synchronous fluorescence intensities were measured at 227.9 nm for DIP and at 264.9 nm for PHE. Calibration plots were rectilinear over the concentration range 0.30–3.50 μg ml^?1 and 0.03–0.75 μg ml^?1 for DIP and PHE, respectively. The proposed methods were successfully applied to determine the studied compounds in pure form and in pharmaceutical preparations.     

Utility of green chemistry for sustainable fluorescence derivatization approach for spectrofluorimetric quantification of Darolutamide as antineoplastic drug in pharmaceutical formulation and spiked human plasma

Darolutamide is an oral nonsteroidal androgen receptor antagonist used to delay the process of prostate cancer to metastatic disease and to increase the quality of life for people with advanced prostate cancer. Here, a second spectrofluorimetric method was advanced for quantifying Darolutamide in pharmaceutical formulation and spiked human plasma. This method depends on the fluorescence derivatization of Darolutamide with 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) at 75°C in a (pH 9) of borate buffer to produce a fluorescent derivative that can be detected at 520 nm after excitation at 460 nm. The method has been validated using ICH criteria, and it demonstrated linearity in the range 5–200 ng ml⁻¹_. The limit of detection (LOD) and limit of quantitation (LOQ) were 1.15 and 3.84 nm, respectively. The proposed method was applied precisely and accurately for quantifying Darolutamide within the pharmaceutical formulation and spiking human plasma without any interferences. Moreover, the method's sustainability was evaluated and compared with the published method using two greenness assessment tools termed analytical eco-scale and Analytical GREEnness (AGREE). These findings suggest that the method is more sustainable than the published method.     

Development and validation of a spectrofluorimetric method for the quantification of ceftriaxone in pharmaceutical formulations and plasma

We describe the development and validation of a new, simple, sensitive and cost‐effective method for the determination of ceftriaxone in commercial formulations and spiked human plasma. The method proposes the conversion of ceftriaxone into a fluorescent product by reacting with ortho‐phthalaldehyde (OPA) in the presence of sulfite at room temperature. The reaction medium is buffered to pH 10 using borate buffer. The derivatized reaction product is highly fluorescent and exhibits maximum fluorescence intensity at λ_em = 386 nm after excitation at λ_ex = 324 nm. The experimental parameters affecting progress of the derivatization reaction were carefully studied and optimized. Under optimum experimental conditions, the method has an excellent correlation coefficient of 0.9984 with a broad linear range of 0.4?20 µg/mL. The limit of detection (LOD) and limit of quantification (LOQ) were found to be 1.30 × 10^?3 and 3.90 × 10^?3 µg/mL, respectively. The interference effects of common excipients on the quantification of drug were investigated and no interference effect was observed. The proposed method has been successfully applied to the determination of ceftriaxone in pharmaceutical formulations and spiked human plasma samples. The method has been validated statistically through percent recovery studies using standard addition and by comparison with a reference HPLC method. The developed method exhibits excellent inter‐ and intraday precision. Copyright © 2013 John Wiley & Sons, Ltd.     

Selective spectrofluorimetric method for determination of Lisinopril in pharmaceutical preparations and in presence of hydrochlorothiazide: Application to content uniformity testing

A novel sensitive and cost‐effective spectrofluorimetric method has been developed and validated for determination of lisinopril (an angiotensin converting enzyme inhibitor) in its pure form and pharmaceutical preparations. The method is based on the reaction of the drug with ninhydrin and phenylacetaldehyde in buffered medium (pH 7.0) to form a highly fluorescent product measured at 460 nm after excitation at 390 nm. Different experimental parameters were optimized and calibration curve was constructed. The fluorescence‐concentration relationship was linear in the range of 0.15–4.0 μg mL^?1. The calculated Limit of detection (LOD) and Limit of quantitation (LOQ) were 0.04 and 0.12 μg mL^?1, respectively. The method was successfully applied for the analysis of pharmaceutical preparations containing the studied drug either alone or co‐formulated with hydrochlorothiazide. The obtained results were in agreement with those of the reported method in respect to accuracy and precession. Moreover, the method was applied content uniformity testing according to United States Pharmacopeia (USP) guidelines.     

Green sensitive spectrofluorimetric determination of pemetrexed as antineoplastic drug: Application in pharmaceutical dosage forms and spiked human plasma

A recent antineoplastic medication is pemetrexed, this medicine is now being developed and produced on a large scale, thus approaches for quality control are urgently needed. Spectrofluorimetric guidelines for the simple estimation of pemetrexed were validated. Pemetrexed's assay depends on observations of its native fluorescence at wavelengths 275/450 nm and pH 4. The proposed approach was also used to identify the examined drug in both its formulation and in human plasma that had been spiked.     

A novel green spectrofluorimetric method for simultaneous determination of antazoline and tetryzoline in their ophthalmic formulation

A novel spectrofluorimetric method has been developed for determination of antazoline (ANT) and tetryzoline (TET) in their pharmaceutical formulation. A combined application of synchronous spectrofluorimetry and second derivative mathematical treatment was developed. The proposed method depends on reacting the cited drugs with dansyl chloride (DNS-Cl) being a suitable derivatizing agent generating highly fluorescent derivatives measured at emission wavelengths of 703.0 and 642.0 nm after excitation wavelengths of 350.0 and 320.0 nm for ANT and TET, respectively. The joint use of synchronous spectrofluorimetry with second derivative mathematical treatment is for the first time to be developed and optimized in aid of using fluorescence data manager software generating second derivative peak amplitudes at 556.5 nm for ANT and 516.7 nm for TET. Linear responses have been represented over a wide range of concentration (0.5–12.0 μg/mL for ANT and 0.5–10.0 μg/mL for TET). Additionally, statistical comparison of the developed method with the official ones has been carried out where no significant difference was found. Additionally, greenness profile assessment was accomplished by means of four metric tools. Indeed, the method developed is found to be precise, sensitive, and discriminating to assess the cited drugs for regular analysis.     

Microenvironment improvement protocol for the sensitive spectrofluorimetric determination of an hepatitis C virus antiviral (Simeprevir): application to human plasma

Simeprevir (SPV) is a powerful antihepatitis C virus agent that was newly introduced into the pharmaceutical market. We here established and validated an easy, simple, and sensitive spectrofluorimetric method for its estimation at λ_em 427 nm (λ_ex 337 nm). The suggested procedure was based on two times enhancement in the original emission of SPV through modifying its microenvironment in buffered aqueous solution by adding Triton X‐100. The relationship between the concentration of SPV and the observed fluorescence intensity was linear in the range 0.06–1.0 μg ml^?1 with a correlation coefficient of 0.9997. The limits of detection and quantitation were 21 and 64 ng ml^?1, respectively. The present method was effectively applied to quantify SPV content in pharmaceutical tablets and human plasma spiked with the drug with no interference from tablet excipients or plasma components.