Whiteness and greenness assessments of a sensitive HPLC method with fluorimetric detection for dapagliflozin quantitation in human plasma: Application to a healthy human volunteer.

The evident ecological impact of human actions, like air pollution, global warming, and ozone depletion, underscores the need for environmentally friendly approaches across various domains, including analytical chemistry. This study aimed to establish a validated, eco-friendly, and sustainable approach utilizing a fluorescence detector coupled with high-performance liquid chromatography for quantifying the antihyperglycemic agent dapagliflozin (DAPA), in human plasma. This method employed a C18 Microsorb MV (4.5 × 250 mm, 5 µm [particle size]) column at 40°C, with 40:60% v/v isocratic elution of acetonitrile and (0.1%) orthophosphoric acid as the mobile phase at 1 mL/min flow rate. DAPA and the internal standard demonstrated their greatest response by performing excitation at 225 nm (λex) and recording chromatograms at an emission wavelength (λem) equal to 305 nm. The presented approach demonstrated high linearity between 50 and 2000 ng/mL and full adherence to the guidelines of the US Food and Drug Administration regarding the validation of bioanalytical methods. The described technique was effectively used for quantification of DAPA in human plasma samples from a healthy male participant who received a tablet of 10 mg DAPA. Analytical Eco-Scale, Analytical GREEnness metric, and the recently created ChlorTox Scale were utilized for greenness assessment. Additionally, the "Red, Green, and Blue 12" model was used in whiteness evaluation.

In-lab synthesized turn-off fluorescence sensor for estimation of Gemigliptin and Rosuvastatin polypill appraised by Spider diagram, AGREE and whiteness metrics.

Gemigliptin-Rosuvastatin single-pill combination is a promising therapeutic tool in the effective control of hyperglycemia and hypercholesterolemia. Organic sensors with high quantum yields have profoundly significant applications in the pharmaceutical industry, such as routine quality control of marketed formulations. Herein, the fluorescence sensor, 2-Morpholino-4,6-dimethyl nicotinonitrile 3, (λex; 226 nm, λem; 406 nm), was synthesized with a fluorescence quantum yield of 56.86% and fully characterized in our laboratory. This sensor showed high efficiency for the determination of Gemigliptin (GEM) and Rosuvastatin (RSV) traces through their stoichiometric interactions and simultaneously fractionated by selective solvation. The interaction between the stated analytes and sensor 3 was a quenching effect. Various experimental parameters and the turn-off mechanism were addressed. The adopted approach fulfilled the ICH validation criteria and showed linear satisfactory ranges, 0.2-2 and 0.1-1 µg/mL for GEM and RSV, respectively with nano-limits of detection less than 30 ng/mL for both analytes. The synthesized sensor has been successfully applied for GEM and RSV co-assessment in their synthetic polypill with excellent % recoveries of 98.83 ± 0.86 and 100.19 ± 0.64, respectively. No statistically significant difference between the results of the proposed and reported spectrophotometric methods in terms of the F- and t-tests. Ecological and whiteness appraisals of the proposed study were conducted via three novel approaches: the Greenness Index via Spider Diagram, the Analytical Greenness Metric, and the Red-Green-Blue 12 model. The aforementioned metrics proved the superiority of the adopted approach over the previously published one regarding eco-friendliness and sustainability. Our devised fluorimetric turn-off sensing method showed high sensitivity, selectivity, feasibility, and rapidity with minimal cost and environmental burden over other sophisticated techniques, making it reliable in quality control labs.

Development, validation and greenness assessment of a new electro-driven separation method for simultaneous analysis of cefixime trihydrate and linezolid in their fixed dose combination.

The establishment and validation of a straightforward, accurate, and eco-friendly capillary zone electrophoretic-diode array detection (CZE-DAD) procedure has been presented for concurrent measurement of two common antibiotics, namely, linezolid (LIN) and cefixime trihydrate (CEF), in their binary mixture or combined dosage form. The selected fused silica capillary has total and effective lengths equal to 58.5 cm and 50 cm, respectively, with a 50 µm internal diameter. Injections were performed utilizing 100 mM borate buffer at pH 10.2 as the background electrolyte (BGE) with a 15.0 s injection time. The finally utilized voltage was 30 kV. DAD was programmed to measure LIN at 250 nm and CEF at 285 nm. In less than 6 min, the two cited drugs were resolved at 2.51 and 5.47 min for LIN and CEF respectively. The introduced procedure had a linear response in the concentration range of 5-50 µg/mL for both analytes with correlation coefficients > 0.9999. Detection and quantification limits were 1.213 and 4.042 µg/mL, respectively, for LIN and 0.301 and 1.004 µg/mL, respectively, for CEF. Validation was conducted according to the International Council for Harmonization (ICH), concerning linearity, detection and quantitation limits, range, accuracy, precision, selectivity, and robustness. Precision was found acceptable due to the low relative standard deviation (RSD%) values that did not exceed 1.86% either for repeatability or for intermediate precision. Additionally, the adequately recovered concentrations and the low values of percentage relative error (Er%) provide evidence of the accuracy of the proposed method. On the other hand, the robustness of the introduced method was affirmed by the acceptable RSD% values that did not exceed 0.6% after deliberate changes in the following procedure parameters: buffer concentration, buffer pH, and wavelength. Finally, the ability of the presented method to quantify the two tested drugs in laboratory-prepared tablets was confirmed by the adequate recoveries (≥ 99%) utilizing the standard-addition procedure, along with the absence of any significant difference between the proposed method and the reference method as proven by the student's t-test and the variance-ratio F-test values that did not exceed the theoretical ones. The analytical Eco-Scale and the analytical GREEness metric (AGREE) were the tools utilized for greenness assessment. This CZE procedure is the first electro-driven separation method that was utilized for the analysis of both antibiotics in their combined laboratory-prepared tablets with no interference from the co-formulated adjuvants.

A New Avenue for Enhanced Treatment of Hyperuricemia and Oxidative Stress: Design, Synthesis and Biological Evaluation of Some Novel Mutual Prodrugs Involving Febuxostat Conjugated with Different Antioxidants.

Febuxostat (FEB) is the first non-purine xanthine oxidase inhibitor (XOI) used for the treatment of hyperuricemia and gout. The oxidative stress induced by reactive oxygen species (ROS) which accompany purine metabolism by XO, could contribute to cellular damage and several pathological conditions. In this view, the present work addresses the evaluation of combining the hypouricemic effect of FEB and the free radical scavenging potential of various natural antioxidants in a single chemical entity by implementing the "mutual prodrug" strategy. Accordingly, a series of five ester prodrugs containing FEB together with different naturally occurring antioxidants namely, thioctic acid (4), thymol (5), menthol (6), vanillin (7), and guaiacol (8) was synthesized. Prominently, all the chemically conjugated prodrugs (4 - 8) revealed an obvious increase in the hypouricemic and antioxidant potentials when compared with their corresponding promoieties and physical mixtures. Moreover, they showed a potential protective effect against CCl4-induced hepatotoxicity and oxidative stress, together with no cytotoxicity on normal breast cells (MCF10A). Furthermore, the in vitro chemical and enzymatic stability studies of the prodrugs (4 - 8) using a developed HPLC method, verified their stability in different pHs, and rapid hydrolysis in rabbit plasma and liver homogenate to their parent metabolites. Moreover, the prodrugs (4 - 8) showed higher lipophilicity and lower aqueous solubility when compared to the parent drugs. Finally, the obtained merits from the implementation of the mutual prodrug strategy would encourage further application in the development of promising candidates with high therapeutic efficacy and improved safety profiles.

Development of a sustainable multianalyte MEKC method for quantitation of the antihyperlipidemic drugs ezetimibe together with three statins. Greenness and whiteness appraisal studies.

Implementing powerful and sustainable research that complies with green analytical chemistry (GAC) and white analytical chemistry (WAC) fundamentals can downsize the environmental compliance costs and fruitfully affects practical and economic issues. Within this framework, rapid and white analytical micellar electrokinetic capillary chromatography (MEKC) methodology was developed for the synchronized estimation of the antihyperlipidemic drugs Ezetimibe (EZE), Atorvastatin (ATO), Rosuvastatin (ROS) and Simvastatin (SIM). The technique was established using fused silica capillary (50 cm, 50 µm id) and the background electrolyte was 0.025 M borate buffer pH 9.2 containing 0.025 M sodium dodecyl sulfate (SDS) and 10% v/v acetonitrile as the organic modifier. Diode array detector was adjusted at 243 nm for ATO and ROS and 237 nm for EZE and SIM. Separation was accomplished within 10 min with migration times of 4.12, 5.42, 8.23 and 8.74 min for ROS, ATO, EZE and SIM respectively. The 4 drugs were quantitated in the concentration range of 10-100 µg/mL and the correlation coefficients were not less than 0.9993. The high sensitivity was illustrated by values of the detection and quantitation limits. The limits of detection for ROS, ATO, EZE and SIM were 0.52, 0.75, 0.42 and 0.64 µg/mL, respectively, whereas, the limits of quantitation values were 1.73, 2.50, 1.40 and 2.13 µg/mL for the studied drugs, respectively. In addition to validation, as reported by the ICH guidelines, greenness and whiteness assessment using the novel AGREE calculator and the holistic functionality model RGB12 were performed. The results proved the efficiency and whiteness of the suggested technique to be routinely implemented in quality control laboratories for the assay of the four drugs and the binary mixtures of EZE with either ATO, ROS or SIM in fixed-dose combined tablets.

Towards the Development of Dual Hypouricemic and Anti-inflammatory Candidates: Design, Synthesis, Stability Studies and Biological Evaluation of Some Mutual Ester Prodrugs of Febuxostat-NSAIDs.

Treatment of gout involves two basic approaches: reducing the serum uric acid mainly by xanthine oxidase inhibitors (XOIs) and alleviating the intensity of the accompanying acute arthritic inflammation using non-steroidal anti-inflammatory drugs (NSAIDs). Febuxostat (FEB) is the first non-purine XOI approved for the treatment of hyperuricemia and gout. The present study aims at combining the hypouricemic effect of FEB and the anti-inflammatory (AI) properties of NSAIDs in a single entity by adopting the "mutual prodrug" approach. Accordingly, a series of seven ester prodrugs comprising basically FEB together with different NSAIDs namely, diclofenac (4), ibuprofen (5), ketoprofen (6), indomethacin (7), naproxen (8), ketorolac (9) and etodolac (10) was synthesized. All the investigated seven prodrugs (4-10) were equipotent or even superior to their corresponding parent drugs in the hypouricemic and AI activities, together with a gastrointestinal (GI) safety profile. Among this series, the prodrug FEB-DIC (4) showed excellent dual in vivo hypouricemic and anti-inflammatory activity (43.60 % and 15.96 %, respectively) when compared to the parent drugs FEB and diclofenac (36.82 % and 12.10 %, respectively) and its physical mixture (37.28 % and 12.41 %, respectively). Investigation of the in vitro chemical stability and hydrolysis of the prodrug (4) in aqueous and biological samples using a developed HPLC method confirmed its stability in various pHs, whereas rapid hydrolysis to the parent drugs in liver homogenate and human plasma was proven. Finally, it is concluded that the mutual prodrug approach could be successfully used in drug design and development for overcoming undesirable difficulties without losing the desired activities of the parent drugs.

Rational Design and Synthesis of New Selective COX-2 Inhibitors with In Vivo PGE2-Lowering Activity by Tethering Benzenesulfonamide and 1,2,3-Triazole Pharmacophores to Some NSAIDs.

New selective COX-2 inhibitors were designed and synthesized by tethering 1,2,3-triazole and benzenesulfonamide pharmacophores to some NSAIDs. Compounds 6b and 6j showed higher in vitro COX-2 selectivity and inhibitory activity (IC50 = 0.04 µM and S.I. = 329 and 312, respectively) than celecoxib (IC50 = 0.05 µM and S.I. = 294). Compound 6e revealed equipotent in vitro COX-2 inhibitory activity to celecoxib. Furthermore, 6b and 6j expressed more potent relief of carrageenan-induced paw edema thickness in mice than celecoxib, with ED50 values of 11.74 µmol/kg and 13.38 µmol/kg vs. 16.24 µmol/kg, respectively. Compounds 6b and 6j inhibited the production of PGE2 with a % inhibition of PGE2 production of 90.70% and 86.34%, respectively, exceeding celecoxib's percentage (78.62%). Moreover, 6b and 6j demonstrated a gastric safety profile comparable to celecoxib. In conclusion, compounds 6b and 6j better achieved the target goal as more potent and selective COX-2 inhibitors than celecoxib in vitro and in vivo.

Quantification of 16 cephalosporins in the aquatic environment by liquid chromatography-tandem mass spectrometry.

Antimicrobial agents are essential to protect human and animal health. During the coronavirus disease 2019 pandemic, antimicrobials such as cephalosporins were widely used as prophylactics and to prevent bacterial co-infection. Undoubtedly, the prevalence of antibiotics in the aquatic environment will ultimately affect the degree of resistance against these bacteria in animals and the environmental systems. In order to monitor 16 cephalosporins in the aquatic environment, we developed a new liquid chromatography-tandem mass spectrometry method that functioned simultaneously under positive and negative electrospray ionization switching modes. The chromatographic separation has been implemented using a pentafluorophenyl propyl column kept at 40°C. The limits of detection and quantitation for the studied cephalosporins ranged from (8 × 10-4 ) to (7.11 × 10-2 ) ng/ml and from (2.61 × 10-3 ) to (2.37 × 10-1 ) ng/ml, respectively. The percent extraction efficiency (apparent recovery) and relative standard deviations for the analyzed cephalosporins ranged from 61.69% to 167.67% and 2.45% to 13.48%, respectively. The overall findings showed that the effluent from the wastewater treatment plants that receive wastewater from pharmaceutical factories had a higher detected amount of cephalosporins than that of domestic sewage. Moreover, seven cephalosporins, including cefuroxime, ceftazidime, cefradine, cefprozil, cefixime, cefalexin, and cefadroxil (0.68-105.45 ng/L) were determined in the aquatic environment.

Febuxostat-based amides and some derived heterocycles targeting xanthine oxidase and COX inhibition. Synthesis, in vitro and in vivo biological evaluation, molecular modeling and in silico ADMET studies.

Various febuxostat derivatives comprising carboxamide functionalities and different substituted heterocycles were synthesized and evaluated for their biological activities as xanthine oxidase (XO) and cyclooxygenase (COX) inhibitors. All the tested compounds exhibited variable in vitro XO inhibitory activities (IC50 values 0.009-0.077 µM), among which the analog 17 has emerged as the most potent derivative (IC50 0.009 µM), representing nearly 3-times the potency of febuxostat (IC50 0.026 µM). The same analogs were further investigated for their in vitro COX-1 and COX-2 inhibitory activity, where fifteen analogs demonstrated recognizable COX-2 inhibitory potential (IC50 values range 0.04 - 0.1 µM), when correlated with celecoxib (IC50 0.05 µM), together with appreciable selectivity indices. Compounds 5a, 14b, 17, 19c, 19e and 21b that showed significant in vitro XO and/ or COX inhibitory potentials were further investigated for their in vivo hypouricemic as well as anti-inflammatory activities. Interestingly, the in vivo results were concordant with the collected in vitro data. Docking of compounds 5a, 14b, 17, 19c, 19e and 21b with the active sites of XO and COX-2 isozymes demonstrated superior binding profile compared with the reported ligands (febuxostat and celecoxib, respectively). Their docking scores were reasonable and cohering to a great extent with their corresponding in vitro IC50 values. Moreover, in silico computation of the predicted pharmacokinetic and toxicity properties (ADMET), together with the ligand efficiency (LE) of the same six compounds suggesting their liability to act as new orally active drug candidates with a predicted high safety profile.

A microfabricated potentiometric sensor for metoclopramide determination utilizing a graphene nanocomposite transducer layer.

In the recent drug analysis arena, optimizing a green, eco-friendly, and cost-effective technique is the main target. In order to cope with green analytical chemistry principles and the trending development of miniaturized portable and handheld devices, an innovative microfabricated ion-selective electrode for the analysis of metoclopramide (MTP) was developed. The fabricated electrode adopted a two-step optimization process. The first step of optimization depended on screening different ionophores in order to enhance the sensor selectivity. Calix-4-arene showed the maximal selectivity towards MTP. The second step was utilizing a graphene nanocomposite as an ion-to-electron transducer layer between the calix-4-arene polymeric membrane and the microfabricated copper solid-contact ion-selective electrode. The graphene nanocomposite layer added more stability to electrode potential drift and short response times (10 s), probably due to the hydrophobic behavior of the graphene nanocomposite, which precludes the formation of a water layer at the Cu electrode/polymeric membrane interface. The proposed MTP sensor has been characterized according to IUPAC recommendations and the linear dynamic range estimated to be 1 × 10-6 to 1 × 10-2 M with LOD of 3 × 10-7 M. The proposed sensor has been successfully employed in the selective determination of MTP in bulk powder, pharmaceutical formulation, and biological fluid. No statistical significant difference was observed upon comparing the results with those of the official method. The Eco-score of the method was assessed using the Eco-Scale tool and was compared with that of the official method. Graphical abstract.

Quick Test for Determination of N-Bombs (Phenethylamine Derivatives, NBOMe) Using High-Performance Liquid Chromatography: A Comparison between Photodiode Array and Amperometric Detection.

The emergence of a new class of novel psychoactive substances, N-benzyl-substituted phenethylamine derivatives so-called "NBOMes" or "Smiles", in the recreational drug market has forced the development of new sensitive analytical methodologies for their detection and quantitation. NBOMes' hallucinogenic effects mimic those of the illegal psychedelic drug lysergic acid diethylamide (LSD) and are typically sold as LSD on blotter papers, resulting in a remarkable number of fatalities worldwide. In this article, four halide derivatives of NBOMe, namely, 2-(4-fluoro-2,5-dimethoxyphenyl)-N-(2-methoxybenzyl)ethan-1-amine, 2-(4-chloro-2,5-dimethoxyphenyl)-N-(2-methoxybenzyl)ethan-1-amine, 2-(4-bromo-2,5-dimethoxyphenyl)-N-(2-methoxybenzyl)ethan-1-amine, and 2-(4-iodo-2,5-dimethoxyphenyl)-N-(2-methoxybenzyl)ethan-1-amine, were detected and quantified simultaneously using a high-performance liquid chromatographic method, and two detection systems were compared: photodiode array detection (detection system I) and amperometric detection via a commercially available impinging jet flow-cell system incorporating embedded graphite screen-printed macroelectrodes (detection system II). Under optimized experimental conditions, linear calibration plots were obtained in the concentration range of 10-300 and 20-300 µg mL-1, for detection systems I and II, respectively. Detection limit (limit of detection) values were between 4.6-6.7 and 9.7-18 µg mL-1, for detection systems I and II, respectively. Both detectors were employed for the analysis of the four NBOMe derivatives in the bulk form, in the presence of LSD and adulterants commonly found in street samples (e.g. paracetamol, caffeine, and benzocaine). Furthermore, the method was applied for the analysis of simulated blotter papers, and the obtained percentage recoveries were satisfactory, emphasizing its advantageous applicability for the routine analysis of NBOMes in forensic laboratories.

High-performance liquid chromatography with diode array detection method for the simultaneous determination of seven selected phosphodiesterase-5 inhibitors and serotonin reuptake inhibitors used as male sexual enhancers.

This work presents a simple, sensitive and generic high-performance liquid chromatography with diode array detection method for the simultaneous determination of seven drugs prescribed for the treatment of erectile dysfunction and premature ejaculation. Investigated drugs include the phosphodiesterase-5 inhibitors: sildenafil, tadalafil, and vardenafil, in addition to the selective serotonin reuptake inhibitors: dapoxetine, duloxetine, fluoxetine, and paroxetine. The drugs were separated using a Waters C8 column (4.6 × 250 mm, 5 µm) with the mobile phase consisting of phosphate buffer pH 3, acetonitrile and methanol in the ratio 60:33:7. The flow rate was 1.2 mL/min, and quantification was based on measuring peak areas at 225 nm. Peaks were perfectly resolved with retention times 3.3, 3.9, 6.4, 7.5, 9.5, 10.7, and 13.4 min for vardenafil, sildenafil, paroxetine, duloxetine, dapoxetine, fluoxetine, and tadalafil, respectively. The developed method was validated with respect to system suitability, linearity, ranges, accuracy, precision, robustness, and limits of detection and quantification. The proposed method showed good linearity in the ranges 5-500, 2-200, 2-200, 3-300, 1.5-150, 2-200, and 2-200 µg/mL for sildenafil, tadalafil, vardenafil, dapoxetine, duloxetine fluoxetine, and paroxetine, respectively. The limits of detection were 0.18-0.38 µg/mL for the analyzed compounds. The applicability of the proposed method to real life situations was assessed through the analysis of commercial tablets, and satisfactory results were obtained.

Validated spectrofluorimetric determination of two pharmaceutical antihypertensive mixtures containing amlodipine besylate together with either candesartan cilexetil or telmisartan.

Amlodipine besylate (AML) is available in fixed-dose combination tablets with either candesartan cilexetil (CAN) or telmisartan (TEL). This work describes a simple, selective and sensitive spectrofluorimetric method for analysis of AML/CAN and AML/TEL binary mixtures without prior separation. The method involves measurement of the native fluorescence of AML at excitation and emission wavelengths of 367 and 454 nm, respectively, in water without interference from either of the two drugs. By contrast, the intrinsic fluorescence of CAN was measured at excitation and emission wavelengths of 265 and 392 nm, respectively, in ethanol, while TEL was measured at 366 nm in 0.05 M sodium hydroxide solution using 294 nm as the excitation wavelength. The proposed spectrofluorimetric procedure was validated with respect to linearity, ranges, precision, accuracy, selectivity, robustness, detection and quantification limits. Regression analysis showed a good correlation between fluorescence intensity and concentration over the ranges 0.1-1.4, 0.025-0.25 and 0.0025-0.05 µg/mL for AML, CAN and TEL, respectively. Limits of detection were 0.034, 0.0063 and 0.0007 µg/mL for AML, CAN and TEL, respectively. The proposed method was successfully applied for the analysis of several synthetic binary mixtures of different ratios and laboratory-prepared tablets with good recoveries, and no interference from common pharmaceutical additives was observed.

New simple spectrophotometric method for determination of the binary mixtures (atorvastatin calcium and ezetimibe; candesartan cilexetil and hydrochlorothiazide) in tablets.

A new simple spectrophotometric method was developed for the determination of binary mixtures without prior separation. The method is based on the generation of ratio spectra of compound X by using a standard spectrum of compound Y as a divisor. The peak to trough amplitudes between two selected wavelengths in the ratio spectra are proportional to concentration of X without interference from Y. The method was demonstrated by determination of two drug combinations. The first consists of the two antihyperlipidemics: atorvastatin calcium (ATV) and ezetimibe (EZE), and the second comprises the antihypertensives: candesartan cilexetil (CAN) and hydrochlorothiazide (HCT). For mixture 1, ATV was determined using 10 µg/mL EZE as the divisor to generate the ratio spectra, and the peak to trough amplitudes between 231 and 276 nm were plotted against ATV concentration. Similarly, by using 10 µg/mL ATV as divisor, the peak to trough amplitudes between 231 and 276 nm were found proportional to EZE concentration. Calibration curves were linear in the range 2.5-40 µg/mL for both drugs. For mixture 2, divisor concentration was 7.5 µg/mL for both drugs. CAN was determined using its peak to trough amplitudes at 251 and 277 nm, while HCT was estimated using the amplitudes between 251 and 276 nm. The measured amplitudes were linearly correlated to concentration in the ranges 2.5-50 and 1-30 µg/mL for CAN and HCT, respectively. The proposed spectrophotometric method was validated and successfully applied for the assay of both drug combinations in several laboratory-prepared mixtures and commercial tablets.