Utility of Green Chemistry for Sustainable Fluorescence Derivatization Approach for Spectrofluorimetric Quantification of Pemetrexed as Antineoplastic Drug in Pharmaceutical Formulation and Spiked Human Plasma.

Pemetrexed is a chemotherapeutic medicine, under the trade name Alimta. Malignant pleural mesothelioma patients. Its application in lung cancer has been studied. Here in, a second spectrofluorimetric method was advanced for quantifying of Pemetrexed in pharmaceutical formulation and spiked human plasma. That method depends on fluorescence derivatization of Pemetrexed 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 which can be detected at 520 nm afterwards excitation at 460 nm. The method has been validated using ICH criteria, and it demonstrated linearity in a range of 2-120 ngmL-1. The proposed method was applied precisely and accurately for quantifying Pemetrexed within pharmaceutical formulation and spiking human plasma without any interferences. Moreover, the method's sustainability was evaluated and compared to the published method using two greenness assessment tools termed analytical eco-scale and Analytical GREENness (AGREE). That findings suggest that the method is more sustainable than the published method.

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.

Dual benefits of NBD-Cl fluorogenic action and sample pretreatment (SALLE) technology in the assessment of anticoagulant medication: Dabigatran in pharmaceutical capsules and plasma samples.

Dabigatran (DBG), marketed as Pradaxa, is an anticoagulant medication prescribed for the treatment and mitigation of blood clots and to lower the risk of stroke in individuals with the heart condition known as atrial fibrillation. This medication is specifically indicated for preventing blood clots post hip or knee replacement surgeries and in patients with a prior history of clots. Compared to warfarin, dabigatran serves as a viable alternative that does not necessitate routine blood monitoring tests. The complimentary benefits associated with SALL (salting-out assisted liquid-liquid extraction) and the fluorogenic capabilities of benzofurazan. These methods were combined to provide an affordable and sensitive DBG assaying method. The spectral strength of the yellow luminous product was examined at 533.8 nm and by adjustment of a wavelength of 474.7 nm for excitation. To assess its linearity, the calibration chart was tested across a DBG concentration range of 30-500 ng/ml. Via accurate computation based on ICH, the detection limit (LD) was determined to be 9.5 ng/ml, and the strategy can quantify the DBG to a limit of 28 ng/ml. To ensure success, various crucial parameters for method implementation have been extensively studied and adapted. The validation of the strategy adhered to the policies outlined by ICH, affirming its precision in quantifying DBG in capsules. Furthermore, the inclusion of SALLE steps facilitated accurate monitoring of DBG in plasma samples, introducing a unique and advanced methodology for analyzing this compound in biological samples.

Ultra-Sensitive Fluorimetric Method for the First Estimation of Vonoprazan in Real Human Plasma and Content Uniformity Test.

Vonoprazan (VON) has been approved recently via US-FDA in 2015 as the first in class of potassium competitive acid blocker group. VON is used for management of GIT ulcer, reflux esophagitis and for eradication of Helicobacter pylori. So, the first spectrofluorimetric method was developed for estimation of VON in real human plasma and content uniformity test. The fluorimetric methodology based on reaction of secondary amine group in VON with benzofurazan (0.05% w/v NBD-Cl) reagent as nucleophilic substitution reaction in alkaline medium (0.1 M borate buffer pH 8.2) to produce highly fluorescent product measure at 530 nm after excitation at 465 nm. The linear calibration range was found 15 to 200 ng mL-1 with lower limit of quantitation (LOQ) equal to 8.57 ng mL-1. The method was successfully applied for estimation of VON in pharmacokinetic (PK) and content uniformity studies. The maximum plasma concentration was found to be (Cmax) 71.03 ng mL-1 after maximum time (tmax) equal to 1.5 ± 0.15 h. The presented strategy also applied to ensure concentration of drug in each tablet using content uniformity test with high percent of recovery 100.05 ± 0.66. The proposed method was established for clinical laboratories and therapeutic drug monitoring studies.

Development of An HPLC Method for the Determination of Mesalazine in Human Plasma by Fluorimetric Derivatization and Application to A Prototype Pharmacokinetic Study.

In this study, a new, fast and sensitive HPLC method with fluorometric detection was developed for the determination of mesalazine in human plasma and applied to a pharmacokinetic study. Mesalazine was precolumn derivatized with NBD-Cl and the fluorescent derivative was separated on a C18 (150 × 4.6 mm × 2.6 µm) analytical column at 30 ºC using a mobile phase composed of acetonitrile-0.1% o-phosphoric acid in water (70:30, v/v) by isocratic elution with flow rate of 1.0 mL min-1. The method was based on the measurement of the derivative using fluorescence detection (λex = 280 nm, λem = 325 nm). The retention time of mesalazine is 3.08 ± 0.06 min. Nortriptiline was used as internal standard. This currently developed method was validated according to ICH criteria by evaluating the specificity, linearity, precision, accuracy and robustness. The method was determined to be linear in a concentration range of 0.25-1.5 µg mL-1 with the correlation coefficient of 0.9997. LOD and LOQ were found to be 0.075 and 0.25 µg mL-1, respectively. Intraday and interday RSD values were less than 5.92%. The plasma concentration-time profile and pharmacokinetic parameters such as AUC0-t, AUC0-∞, Cmax, tmax, t1/2, were calculated according to the assays. The presented method can certainly be used for bioequivalence and bioavailability investigations and routine analysis of the drug in plasma.

Two spectroscopic methods for estimation of anti-migraine medications: Sumatriptan and Zolmitriptan based on nucleophilic substitution reaction.

Sensitive and selective spectrophotometric and spectrofluorimetric methods have been developed for the estimation of two anti-migraine drugs, namely sumatriptan succinate (SUM) and zolmitriptan (ZOL). These methods depend on producing a yellow-coloured product after the reaction of the two drugs with 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD-Cl). The reaction products exhibited maximum absorbance at 481 nm in borate buffer of pH 9 and fluorescence emission peak at 540 nm after excitation at 470 nm for the two drugs. The linear ranges were 5-60 µg/ml for SUM and 5-50 µg/ml for ZOL in the spectrophotometric method (Method I), whereas this was 0.4-4 µg/ml for SUM and 0.5-5 µg/ml for ZOL in the spectrofluorimetric method (Method II). The method validity was assessed according to International Council for Harmonisation (ICH) guidelines. Statistical analysis of the results obtained from the proposed and comparison methods confirmed that the proposed methods were highly accurate and precise. The suggested methods could be used for the determination of the mentioned drugs in both pure form and in tablets.

Bio-analytically fluorimetric method for estimation of ertapenem in real human plasma and commercial samples; application to pharmacokinetics study.

Ertapenem (EPM) has been recently approved by the United States Food and Drug Administration (US-FDA) as an antimicrobial drug. EPM has a broad spectrum of action against different bacterial strains and is most commonly prescribed in Egypt for the treatment of Klebsiella pneumonia. In this study, EPM was estimated using a sensitive and selective spectrofluorimetric method for human plasma and pharmaceutical vials. The measured fluorescence (at 540 nm) was obtained from reaction of EPM with 0.05% w/v benzofurazan (NBD-Cl) using 0.1 M borate buffer pH 8.8 after excitation at 460 nm. The fluorometric linear range was stable from 10 to 350 ng ml-1 . The lower limit of detection and the lower limit of quantitation were found to be 2.13 and 6.47 ng ml-1 respectively. Many factors such as pH, temperature, heating time, and NBD-Cl concentration were optimized. The presented work was validated according to International Council for Harmonisation guidelines and bio-analytically validated using FDA recommendations. The significant finding of this study, sensitivity, was successfully applied in Egypt for a pharmacokinetic application and commercial vials. Pharmacokinetic parameters were studied and the result, recorded as Cmax of EPM, was found to be 83.60 µg ml-1 after infusion of 0.5 g of Invanz® for 30 min. AUC0-∞ was found to be 320 ± 30.2 µ.h ml-1 .

Novel spectrofluorimetric quantification of alogliptin benzoate in biofluids exploiting its interaction with 4-chloro-7-nitrobenzofurazan.

The direct determination of alogliptin benzoate (ALO) using fluorescence has not yet been accomplished because ALO cannot fluoresce naturally. Accordingly, it should be derivatized first with a fluorogenic reagent to enhance the sensitivity required for its bioanalysis. This method is the first spectrofluorimetric assay for ALO quantification exploiting the nucleophilic nature of its amino group to react with 4-chloro-7-nitrobenzofurazan (NBD-Cl) in borate buffer at pH 8.5 to produce a strong fluorescent compound that is excited at and emits at wavelengths 470 and 527 nm, respectively. Experimental variables concerning the conditions of reaction and fluorogenic intensity were carefully investigated and optimized. Linearity was from 1-250 ng ml-1 with a lower detection limit of 0.29 ng ml-1 and a lower quantification limit of 0.88 ng ml-1 . Validation of the current study was accomplished with mean per cent recovery of 100.62 ± 1.59 in tablets and 99.86 ± 0.82 in human plasma. Furthermore, the current method has been utilized in the bioanalysis of ALO in real rat plasma after oral administration with a simple specimen preparation. The developed method has proven to be a promising alternative method for ALO analysis in bioequivalence studies.

Novel spectrofluorimetric quantification of linagliptin in biological fluids exploiting its interaction with 4-chloro-7-nitrobenzofurazan.

Direct determination of linagliptin (LIN) using fluorimetry has been limited because LIN releases a very weak fluorescence signal. Accordingly, it should be derivatized first with a fluorogenic reagent to enhance its fluorescence and consequentially the sensitivity required for its bioanalysis. This is the first description of a spectrofluorimetric method for LIN quantification in human plasma. The suggested method exploits the nucleophilic nature of its amino group to react with 4-chloro-7-nitrobenzofurazan (NBD-Cl) in borate buffer at pH 8.5 to yield a strong fluorescent product with excitation and emission wavelengths of 459 and 529 nm, respectively. Experimental variables concerning the conditions of reaction and fluorescence intensity were carefully investigated and optimized. The linearity range was from 1.0-100 ng ml-1 with a lower detection limit and a lower quantification limit of 0.60 ng ml-1 and 1.82 ng ml-1 , respectively. Validation of the suggested method has been accomplished and the application to LIN analysis in commercial tablets as well as in human plasma resulted in a mean per cent recovery of 100.12 ± 1.57 and 99.65 ± 1.22, respectively. The developed method was proven to be a promising, simple and fast alternative bioanalytical method for LIN quantification in clinical and bioequivalence research.

Utility of the fluorogenic characters of benzofurazan for analysis of tigecycline using spectrometric technique; application to pharmacokinetic study, urine and pharmaceutical formulations.

Tigecycline (TIGE) is the newest tetracycline derivative antibiotic with low toxicity, it is used for management of infectious diseases caused by Gram-positive and Gram-negative bacteria. Hence, an efficient, selective and sensitive method was developed for analysis of TIGE in commercial formulations, human plasma and urine. The spectrofluorimetric technique based on the reaction of secondary amine moiety in TIGE with 4-chloro-7-nitrobenzofurazan (NBD-Cl) in slightly alkaline medium producing a highly fluorescent product measured at 540 nm (λex at 470 nm) after heating for 15 min at 75°C. The proposed strategy was upgraded and approved by ICH rules and bio-analytical validated using US-FDA recommendations. A linear relationship between fluorescence intensity and TIGE concentration was observed over the concentration range 40-500 ng mL-1 with limit of quantification (LOQ) 21.09 ng mL-1 and limit of detection (LOD) 6.96 ng mL-1 .The ultra-affectability and high selectivity of the proposed strategy permits analysis of TIGE in dosage form, human plasma and urine samples with good recovery ranged from 97.23% to 98.72% and from 99.36% to 99.80% respectively, without any interfering from matrix components. Also, the developed strategy was used to examine the stability of TIGE in human plasma and applied for pharmacokinetic investigation of TIGE.

Development of sensitive benzofurazan-based spectrometric methods for analysis of spectinomycin in vials and human biological samples.

Spectinomycin hydrochloride (SPEC) is an aminoglycoside antibiotic that has a broad spectrum against several bacterial strains of humans and veterinary infections. However, SPEC lacks a fluorophore or chromophore and this lack makes its analysis a challenge. Our study provides a simple, sensitive and low-cost spectrofluorimetric/spectrophotometric method based on the reaction between secondary amine groups and a benzofurazan reagent using borate buffer (pH 9.2). The yielding compound was measured fluorimetrically at 530 nm (excitation at 460 nm) with colorimetry at 410 nm. The calibration curves ranged from 30 to 400 ng ml-1 and from 0.2 to 6 µg ml-1 for spectrofluorimetric and spectrophotometric analyses, respectively. The limits of detection were calculated to be 4.15 ng ml-1 and 0.05 µg ml-1 for spectrofluorimetric and spectrophotometric processes, respectively. The ultra-affectability and high selectivity of the proposed method permitted analysis of SPEC in the dosage form and in human plasma samples, with good recoveries of about 101.19 and 97.11%, respectively, without any matrix interference. The proposed strategy was validated using International Conference on Harmonization standards and validated bioanalytically using USFDA recommendations.

Benzofurazan-based fluorophore for the selective determination of flupentixol dihydrochloride: Application to content uniformity testing.

One of the most commonly used drugs in treatment of schizophrenia is flupentixol dihydrochloride, therefore it is important to develop a simple, low cost and sensitive spectrofluorimetric method for the estimation of flupentixol dihydrochloride. The yellow fluorescent product that is generated from the nucleophilic substitution reaction of the free lone pair of the alcoholic hydroxyl group of the drug and 4-chloro-7-nitrobenzofurazan (NBD-Cl) in Mcllvaine buffer pH 7.0 was estimated at 510 nm (λex 460 nm). The variables that affect the development of the reaction product were explored and optimized. The linear range of this method was 0.5-2.5 µg ml-1 with a limit of quantitation equal to 0.29 µg ml-1 . Our method was successfully applied for the assurance of flupentixol in tablet form with average percentage recovery of 99.08 ± 1.01% without obstruction from the basic excipients exhibits. Furthermore, our strategy was extended to study the content uniformity testing of flupentixol in Fluaxnol® tablets.

Validated spectroscopic methods for the estimation of triamterene in bulk powder and capsule form via derivatization reactions.

Sensitive, simple and specific spectrophotometric and spectrofluorimetric methods were developed for the determination of triamterene in bulk powder and pharmaceutical dosage form. The spectrophotometric method (Method A) is based on the formation of an ion-pair complex with eosin Y in acetate buffered solution of pH 3.7 followed by measuring the absorbance at 545 nm. The absorbance-concentration plot is rectilinear over the range 3.0-15.0 µg/mL with a limit of detection (LOD) of 0.429 µg/mL and a limit of quantitation (LOQ) of 1.031 µg/mL. The spectrofluorimetric method (Method B) is based on the reaction of triamterene with 7- chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) in basic solution (pH 10) to form a product with high fluorescence measured at 546 nm after being excited at 438 nm. The plot of fluorescence versus concentration is linear within the range 2.0-10.0 µg/mL. The suggested methods were applied for the analysis of commercial capsules containing the studied drug with successful results. The results obtained from the proposed method were statistically compared with those of a reported one and revealed good agreement. The presented methods are useful in routine analysis of triamterene in laboratories of quality control.

Application of 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole in analysis: Fluorescent dyes and unexpected reaction with tertiary amines.

4-Chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) is widely applied as a fluorescent tagging reagent in biochemistry, as a derivatization agent in analytical chemistry, and as a component for design of fluorescent nanoparticles. Four new 7-nitrobenzo-2-oxa-1,3-diazole (NBD)-tagged polyamines containing two to four amine moieties were synthesized and used as an effective tool for staining of siliceous frustules of the diatom algae and spicules of the siliceous sponges, including fossilized samples. An unexpected reaction between NBD-Cl and tertiary amine groups was found, giving rise to NBD-tagged amines with elimination of an alkyl group. The reaction proceeds through the Meisenheimer complex and quaternary salt, which transform to the product by Hofmann reaction (alkene elimination) or nucleophilic substitution (halogenated compound formation). In the case of polyamines, NBD-Cl causes chain scissoring, giving a set of NBD-tagged amines. The found NBD-Cl reaction with tertiary amines must be taken into account when using NBD-Cl and similar activated aromatic systems for amine derivatization in analytical and biochemistry applications. The reaction with polyamines opens the way to libraries of NBD-tagged compounds.

Validated spectrofluorimetric and spectrophotometric methods for the determination of brimonidine tartrate in ophthalmic solutions via derivatization with NBD-Cl. Application to stability study.

Two simple, selective and accurate methods were developed and validated for the determination of brimonidine tartrate (BT) in pure state and pharmaceutical formulations. Both methods are based on the coupling of the drug with 4-chloro-7-nitro-2,1,3-benzoxadiazole in borate buffer (pH 8.5) at 70 °C and measurement of the reaction product spectrophotometrically at 407 nm (method I) or spectrofluorimetrically at 528 nm upon excitation at 460 nm (method II). The calibration graphs were rectilinear over the concentration ranges of 1.0-16.0 and 0.1-4.0 µg/mL with lower detection limits of 0.21 and 0.03, and lower quantification limits of 0.65 and 0.09 µg/mL for methods I and II, respectively. Both methods were successfully applied to the analysis of commercial ophthalmic solution with mean recovery of 99.50 ± 1.00 and 100.13 ± 0.71%, respectively. Statistical analysis of the results obtained by the proposed methods revealed good agreement with those obtained using a comparison method. The proposed spectrofluorimetric method was extended to a stability study of BT under different ICH-outlined conditions such as alkaline, acidic, oxidative and photolytic degradation. Furthermore, the kinetics of oxidative degradation of the drug was investigated and the apparent first-order reaction rate constants, half-life times and Arrhenius equation were estimated. The proposed methods are practical and valuable for routine applications in quality control laboratories for the analysis of BT.

Validated spectrofluorimetric method for the determination of carbamazepine in pharmaceutical dosage forms after reaction with 4-chloro-7--nitrobenzo-2-oxa-1,3-diazole (NBD-Cl).

A sensitive and simple spectrofluorimetric method has been developed and validated for the determination of the anti-epileptic drug carbamazepine (CBZ) in its dosage forms. The method was based on a nucleophilic substitution reaction of CBZ with 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) in borate buffer (pH 9) to form a highly fluorescent derivative that was measured at 530 nm after excitation at 460 nm. Factors affecting the formation of the reaction product were studied and optimized, and the reaction mechanism was postulated. The fluorescence-concentration plot is rectilinear over the range of 0.6-8 µg/mL with limit of detection of 0.06 µg/mL and limit of quantitation of 0.19 µg/mL. The method was applied to the analysis of commercial tablets and the results were in good agreement with those obtained using the reference method. Validation of the analytical procedures was evaluated according to ICH guidelines.

Development of sensitive spectrofluorimetric and spectrophotometric methods for the determination of duloxetine in capsule and spiked human plasma.

A new, sensitive and selective spectrofluorimetric method has been developed for the determination of duloxetine (DLX) in capsule and spiked human plasma. DLX, as a secondary amine compound, reacts with 7-chloro-4-nitrobenzofurazon (NBD-Cl), a highly sensitive fluorogenic and chromogenic reagent used in many investigations. The method is based on the reaction between the drug and NBD-Cl in borate buffer at pH 8.5 to yield a highly fluorescent derivative that is measured at 523 nm after excitation at 478 nm. The fluorescence intensity was directly proportional to the concentration over the range 50-250 ng/mL. The reaction product was also measured spectrophotometrically. The relation between the absorbance at 478 nm and the concentration is rectilinear over the range 1.0-12.0 µg/mL. The methods were successfully applied for the determination of this drug in pharmaceutical dosage form. The spectrofluorimetric method was also successfully applied to the determination of duloxetine in spiked human plasma. The suggested procedures could be used for the determination of DLX in pure form, capsules and human plasma being sensitive, simple and selective.

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.

Application of a nucleophilic substitution reaction for spectrofluorimetric determination of aripiprazole in pharmaceutical dosage form and plasma matrix; greenness assessment.

Aripiprazole is an antipsychotic medicine used to treat a variety of mental disorders, including irritability linked with autism disorder in children. Herein, a green and highly sensitive spectrofluorimetric method was developed for the determination of aripiprazole in pharmaceutical dosage form and plasma matrix. The method based on the formation of a fluorescent adduct from the nucleophilic substitution reaction of 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-chloride) with aripiprazole, which can be detected at 542 nm following excitation at 481 nm. Factors that affect the development and fluorescence sensitivity of the reaction product were investigated and optimized. The reaction yielded the most optimal fluorescence responses when it was performed using 1.5 mL of 0.2 % w/v NBD-chloride, 1.5 mL of borate buffer pH 9, heating at 80 °C for 20 min, and ethanol as a diluting solvent. The method was validated as per ICH guidelines for analytical and bioanalytical procedures. Good linearity was established between the fluorescence responses of the reaction product and aripiprazole concentrations in the range of 100-1200 ng/mL with adequate accuracy and precision results. The applied method was very sensitive and selectively determined aripiprazole in pharmaceutical and plasma matrices with no interferences. Furthermore, the compliance of the proposed method with the principles of green analytical chemistry was evaluated in comparison with the reported method using analytical eco-scale and AGREE metrics. The outputs proved that the proposed method complied more with the principles of green analytical chemistry than the reported method.

Benzofurazan -based fluorophore for the spectrofluorimetric determination of 6-Aminocaproic acid: Application to spiked human plasma and urine.

6-Aminocaproic acid is one of the most widely used antihemorrhagic and antifibrinolytic agent, therefore, it is essential to create a novel, sensitive, low cost and straightforward spectrofluorimetric method for its determination. The nucleophilic substitution interaction between the primary amine of 6-aminocaproic acid with 4-chloro-7-nitro benzofurazan (NBD-Cl) generated a yellow product. The reaction proceeded in borate buffer (pH 9) and its fluorescence has been measured at 525 nm after excitation at 472 nm. All of the parameters that have impact on the performance of the developed method were investigated and optimized. The range of linearity was 0.1-0.7 µg/mL while, the quantitation limit was down to 0.101 µg/mL and limit of detection was 0.033 µg/mL. This approach was effectively employed to evaluate the content of 6-aminocaproic acid in laboratory prepared dosage form with average percentage recovery of 100.19 ± 0.72% without any interference from basic excipients. Moreover, the proposed method was extended to determine 6-aminocaproic acid in spiked human plasma and urine.