Chromatographic analysis of triple cough therapy; bromhexine, guaiafenesin and salbutamol and pharmaceutical impurity: in-silico toxicity profile of drug impurity.

Bromhexine (BR), guaiafenesin (GUF) and salbutamol (SAL) are formulated as Ventocough syrup® (with and without sugar), labeled to contain propyl paraben and sodium benzoate as inactive ingredients. They are used to make coughing more productive and easier. A crucial element and a major issue in the pharmaceutical industry is the control of organic related impurities to obtain safe and effective treatment. Guaiacol (GUL) is reported to be GUF related impurity that was proved to be extremely toxic (toxic rating class 5), and its use should be banned. In this work, In-Silico study and ADMET estimation were conducted to predict GUL pharmacokinetic properties and its toxicity profile. Additionally, two chromatographic methods were conducted to analyze the studied components along with GUF impurity in the presence of the labeled dosage form excipients. The In-Silico study assured that GUL has oral rat acute toxicity and it is considered to be skin sensitizer. On the other hand, the developed TLC- densitometeric method depended on using a mobile phase mixture of hexane: methylene chloride: triethylamine (5.0:6.0:0.3, by volume) as a developing system. UV-Scanning was performed immediately at 275 nm for SAL, GUF and GUL, while scanning at 310 nm was used for scanning BR. Linearity was established in the ranges of 0.25-4.0, 0.25-4.0, 0.5-8.0 and 0.1-1.6 µg/band for BR, SAL, GUF and GUL, respectively. In the developed HPLC method, separation was performed on X-Bridge® C18 column (250 × 4.6 mm, 5 µm) using a solvent mixture of 0.05M disodium hydrogen phosphate pH 3 with aqueous phosphoric acid: methanol (containing 0.3%, v/v triethylamine) (40:60, v/v). Detection was done at 225 nm and separation was achieved within 10 min. Linearity was proved in the range of 2-50 µg/mL for the proposed drugs. Validation of the developed methods was done and all the calculated parameters were within the acceptable limits recommended by ICH guidelines. After that, methods were used to examine the potency of the selected marketed dosage forms and concentrations of all drugs were within the acceptable limits. Additionally, complete separation between the studied drugs and the additives were observed. The developed methods can be used during routine quality control analysis of the proposed drugs when the required issues concern on sensitivity, selectivity and analysis time.

The improvement of routine drug quality control analysis of some antihypertensive drugs using high-performance thin-layer chromatography densitometry method with greenness profile evaluation.

The common antihypertensive drugs are B-blockers and diuretics. For the determination of beta-blocker medicines (bisoprolol fumarate and carvedilol) and diuretic drug (Furosemide), new and accurate chromatographic method has been developed. The separation was achieved using a developing system that includes chloroform:methanol:ethyl acetate:ammonia (6:2:2:0.2 by volume) as a mobile phase and the bands were detected at 240 nm. The concentration ranges were 5-25, 1-7, and 1-3.5 µg/band for bisoprolol fumarate, carvedilol, and furosemide, respectively. This chromatographic approach is the first methodology for simultaneously determining bisoprolol fumarate, carvedilol, and furosemide in their pure forms and in their pharmaceutical dosage forms. The advantages of using known analytical procedures are their simplicity, speed, cost effectiveness, lack of laboriousness, and ability to save time as the three tablets are determined in one step and can be used for routine analysis of the investigated combinations in quality control laboratories. According to International Conference of Harmonization guidelines, the established procedures have been validated, and the results were statistically compared to those obtained by the reported reversed-phase-high-performance liquid chromatography methods using Student's t-test and F-test, with no significant difference between them, indicating that the proposed methods can be used for routine drug quality control analysis.

Ecofriendly HPLC Method for Simultaneous Determination of the Co-Prescribed Drugs in Chemotherapy Omeprazole, Ondansetron and Deflazacort in Spiked Human Plasma.

Green, selective and accurate high-performance liquid chromatography (HPLC) chromatographic method is presented for simultaneous separation and quantitation of the co-prescribed drugs in chemotherapy omeprazole, ondansetron and deflazacort in spiked human plasma. An isocratic HPLC separation was performed on X Bridge C18 (4.6 × 250 mm) column with 5 µm particle size using mobile phase consisting of methanol: ammonium acetate buffer pH 4 adjusted by acetic acid (60: 40, v/v). The injection volume was 20µL with UV detection wavelength at 237 nm at room temperature. Flow rate of the mobile phase was adjusted to be 2.0 ml/min. Dexamethasone was used as internal standard to correct the variation during sample pretreatment. FDA guidelines were followed to validate the developed method. Successful application of the developed method was revealed by simultaneous determination of omeprazole, ondansetron and deflazacort in spiked human plasma in ranges of 1-20, 0.1-8 and 0.2-8 µg mL-1 for omeprazole, ondansetron and deflazacort, respectively. Four greenness assessment tools were used to evaluate the greenness of the developed method and the results were accepted. This method permitted the accurate simultaneous determination of the studied drugs, thus it can be used during therapeutic drug monitoring in daily clinical practice.

Ecofriendly Chromatographic Assay of Metformin Combinations; Content Uniformity Tests and Toxicity Profiling.

Accurate, sensitive and green HPTLC chromatographic method was proposed for simultaneous determination of metformin, glipizide and sitagliptin in the presence of metformin potential toxic impurities melamine and cyanoguanidine. The separation was completed on silica gel HPTLC F254 plates using a mixture of ethyl acetate: methanol: ammonia: formic acid (7: 2: 0.2: 0.2, by volume) as a developing system with UV scanning for the developed bands at 235 nm. The Rf values for metformin, glipizide, sitagliptin, melamine and cyanoguanidine were 0.17, 0.84, 0.67, 0.47 and 0.75, respectively. Linear responses were observed in the ranges of 0.2-3, 0.07-1.5, 1.5-5, 0.8-4 and 0.4-2 µg/band with correlation coefficients of 0.9999, 0.9998, 0.9997, 0.9996 and 0.9998 for metformin, glipizide, sitagliptin, melamine and cyanoguanidine, respectively. The proposed method was validated as per ICH criteria with respect to linearity, accuracy, precision, specificity and robustness. The validated method was successfully applied for determination of the studied drugs in Janumet® and Engilor® tablets; also, the results were statistically compared to those obtained by the reported spectrophotometric method and no significant difference was found between them. This method permitted the accurate simultaneous determination of the studied drugs, indicating its ability to be used for routine quality control assays of these drugs.

Diosmin mitigates dexamethasone-induced osteoporosis in vivo: Role of Runx2, RANKL/OPG, and oxidative stress.

Secondary osteoporosis is commonly caused by long-term intake of glucocorticoids (GCs), such as dexamethasone (DEX). Diosmin, a natural substance with potent antioxidant and anti-inflammatory properties, is clinically used for treating some vascular disorders. The current work targeted exploring the protective properties of diosmin to counteract DEX-induced osteoporosis in vivo. Rats were administered DEX (7 mg/kg) once weekly for 5 weeks, and in the second week, vehicle or diosmin (50 or 100 mg/kg/day) for the next four weeks. Femur bone tissues were collected and processed for histological and biochemical examinations. The study findings showed that diosmin alleviated the histological bone impairments caused by DEX. In addition, diosmin upregulated the expression of Runt-related transcription factor 2 (Runx2) and phosphorylated protein kinase B (p-AKT) and the mRNA transcripts of Wingless (Wnt) and osteocalcin. Furthermore, diosmin counteracted the rise in the mRNA levels of receptor activator of nuclear factor-kB ligand (RANKL) and the reduction in osteoprotegerin (OPG), both were induced by DEX. Diosmin restored the oxidant/antioxidant equilibrium and exerted significant antiapoptotic activity. The aforementioned effects were more pronounced at the dose level of 100 mg/kg. Collectively, diosmin has proven to protect rats against DEX-induced osteoporosis by augmenting osteoblast and bone development while hindering osteoclast and bone resorption. Our findings could be used as a stand for recommending supplementation of diosmin for patients chronically using GCs.

Chromatographic analysis of bromhexine and oxytetracycline residues in milk as a drug analysis medium with greenness profile appraisal.

Tetracyclines are frequently employed in animal husbandry. Bromhexine is a mucolytic drug that improves the efficacy of tetracyclines. It has been reported that residues of tetracyclines in milk may have negative effects on humans. Two versatile and accurate methods were developed for concurrent analysis of oxytetracycline (OTC) and bromhexine (BR) residues in spiked milk samples. Sample preparation was carefully considered for extraction and cleanup using the ecofriendly chemicals acetic acid, 0.1 N EDTA and ethanol or methanol. The first method was a TLC-densitometric method in which TLC plates previously treated with 10% EDTA (pH 9 with 40% sodium hydroxide) were used as a stationary phase. A solvent mixture of methanol : methylene chloride : 2% aqueous acetic acid (8 : 2 : 0.5, by volume) was the developing system, and detection was carried out at 254 nm. Metformin was used as the internal standard, and linearity was achieved in the ranges of 0.2-10 and 0.04-2 µg per band for OTC and BR, respectively. The second method was a RP-HPLC method; separation was performed on a C18 column using an isocratic mixture of ethanol : 7.5% aqueous acetic acid (70 : 30, v/v). Separation was achieved within 10 minutes, and linearity was proven in the ranges of 0.05-50 and 0.05-30 µg mL-1 for OTC and BR, respectively. Diclofenac sodium was used as an internal standard. The proposed methods were validated in accordance with the FDA Center for Veterinary Medicine guidelines. Moreover, the performance and health and environmental impacts of the methods were evaluated using several greenness metrics, namely, the National Environmental Methods Index (NEMI), modified NEMI, Green Analytical Procedure Index (GAPI), Analytical Eco-Scale and Analytical GREEnness (AGREE) metric approaches. All the obtained results proved the validity of the developed methods concerning its performance and ecological effects. The methods can be used to investigate the presence of OTC residues in various marketed milk samples to maintain public health.

Sensitive and selective bioscreening of the most commonly used coronavirus disease drug, Favipiravir, and its co-administered therapeutic, Meropenem, in human plasma.

Favipiravir and Meropenem have been concurrently used as directly acting antiviral and antibiotic agents for the treatment of coronavirus disease in human plasma. Accurate and specific reversed phase ultra-performance liquid chromatographic and high-performance thin-layer chromatographic methods were developed and validated for the first time analysis of this combination in spiked human plasma using Cefepime as an internal standard. In the developed ultra-high-performance liquid chromatography method, separation was performed on a BEH C18 column with a mixture of ACN and 0.05 M potassium dihydrogen orthophosphate (pH = 3) in a ratio of 10:90 (v/v) as an eluate. Scanning of the separated peaks was at 298 nm. The developed method showed high sensitivity, and the drugs showed linearity in the range of 5-70 µg/ml for Favipiravir and 2-50 µg/ml for Meropenem. The proposed high-performance thin-layer chromatographic method included the separation using a mixture of ethyl acetate:methanol:deionized water:formic acid (5:4:1.5:0.3, by volume), then spots detection at 300 nm. Methods were investigated for greenness using the eco-scale and national environmental method index tools and were validated according to food and drug administration guidelines. Methods can be applied for bio-analysis and therapeutic drug monitoring studies.

A new green approach for the reduction of consumed solvents and simultaneous quality control analysis of several pharmaceuticals using a fast and economic RP-HPLC method; a case study for a mixture of piracetam, ketoprofen and omeprazole drugs.

One of the main aims of green analytical chemistry (GAC) is the reduction of solvents and chemicals consumed. Recycling the mobile phase in chromatographic techniques provides an efficient way to implement GAC principles. However, this is not an easy job, particularly in the case of the gradient mode. Analysis of multi-pharmaceuticals for the same manufacturer using one mobile phase system dramatically reduces consumed solvents, time, and cost for pharmaceuticals analysis in quality control laboratories. This work is an attempt to reduce time, cost and effort needed for quality control analysis of several dosage forms produced by the same manufacturer. Our novel and green RP-HPLC method is able to separate and quantify a tertiary mixture of piracetam, ketoprofen and omeprazole produced by the same manufacturers. The analyst can easily quantify the three drugs in the three dosage forms in one run using the gradient elution mode of methanol and water (from 50% methanol to 85% methanol in ten minutes) with a flow rate 1.5 mL min-1 on a non-polar C18 column. Suitable dilutions were done for the working solution of the mixed pharmaceutical formulations prior to chromatographic analysis. This procedure will dramatically reduce the consumed solvents and save time and money during pharmaceutical analysis. The calibration ranges are (5-25), (5-25) and (3-20) µg mL-1 for the three studied drugs. The International Council for Harmonization (ICH) procedures were followed in the validation process and the results were evaluated in comparison with official HPLC methods, where no noteworthy differences were found. The green profile of the method and pictograms of AGREE and Green Analytical Procedure Index (GAPI) approaches proved the eco-friendly character for the studied drugs. The simultaneous quantitative analysis for Stimulan® and Hyposec® capsules, and Ketolgin® tablets from the Amoun Pharmaceutical Company, Egypt, can be accomplished via the novel method. Also, Memoral® ampoules, Topfam® tablets, and Gastroloc® capsules from Sigma Pharmaceutical Industries, Egypt, could be analyzed simultaneously. Omez® capsules and Ketogesic® tablets from the Pharaonia Pharmaceuticals, Egypt, could be determined simultaneously too. Applying this RP-HPLC method, a significant reduction of the total cost is assured as the required amount of solvent is noticeably decreased when performing multi-analyses in comparison to single component analysis.

Rapid and ecofriendly UPLC quantification of Remdesivir, Favipiravir and Dexamethasone for accurate therapeutic drug monitoring in Covid-19 Patient's plasma.

Innovative therapeutic protocols to the rapidly spreading coronavirus disease (COVID19) epidemic is highly required all across the world. As demonstrated by clinical studies, Favipiravir (FVP) and Remdesivir (REM) are new antiviral medicines that are effective against COVID-19. REM is the first FDA approved antiviral medicine against COVID-19. In addition to antivirals, corticosteroids such as dexamethasone (DEX), and anticoagulants such as apixaban (PX) are used in multidrug combinations protocols. This work develops and validates simple and selective screening of the four medicines of COVID -19 therapeutic protocol. FVP, REM, DEX, and PX as internal standard in human plasma using UPLC method by C18 column and methanol, acetonitrile, and water acidified by orthophosphate (pH = 4) in a ratio of (15: 35: 50, by volume) as an eluate flowing at 0.3 mL/min. The eluent was detected at 240 nm. The method was linear over (0.1-10 µg/mL) for each of FVP, REM, and DEX. The validation of the UPLC method was assessed in accordance with FDA guidelines. The method can detect as low as down to 0.1 µg/mL for all. The recoveries of the drugs in spiked human plasma ranged from 97.67 to 102.98 percent. Method accuracy and precision were assessed and the drugs showed good stability. The method was proven to be green to the environment after greenness checking by greenness profile and Eco-Scale tool.

Spectrophotometric Determination of Metformin in Different Pharmaceutical Combinations With Glipizide or Sitagliptin in Presence of Toxic Impurities of Metformin.

BACKGROUND: The presented quadruple divisor spectrophotometric method was able to resolve and analyze a complex quintuple drug matrix with severe overlapped spectra without previous separation or extraction steps or need of complicated apparatus like chromatographic methods and had the advantage of being green as the solvent used was water. METHOD: A simple, sensitive, and precise quadruple devisor spectrophotometric method was developed for simultaneous determination of metformin, glipizide, and sitagliptin in the presence of metformin potential impurities melamine and cyanoguanidine. The proposed method was applied for analysis of metformin, glipizide, and sitagliptin in pure form and pharmaceutical formulation (tablets). The developed method was validated and met the requirements for ICH guidelines with respect to linearity, accuracy, precision, specificity, and robustness. RESULTS: A linear response was observed in the range of 2-27, 2-20, 1-20, 0.5-10, and 1-10 µg/mL for metformin, glipizide, sitagliptin, melamine, and cyanoguanidine, respectively, with a correlation coefficient of 0.9996, 0.9998, 0.9997, 0.9997, and 0.9996 for metformin, glipizide, sitagliptin, melamine, and cyanoguanidine, respectively. CONCLUSIONS: The validated method was successfully applied for determination of the studied drugs in Janumet® and Engilor® tablets; moreover, the results were statistically compared to those obtained by the reported RP-HPLC method and no significant difference was found between them, indicating the ability of the proposed method to be used for routine quality-control analysis of these drugs. HIGHLIGHTS: Novel application of quadruple divisor spectrophotometric technique is introduced for resolving mixtures containing five components with severely overlapping spectra. A simple and sensitive spectrophotometric method was developed for simultaneous determination of metformin, glipizide, and sitagliptin in the presence of metformin potential impurities melamine and cyanoguanidine. The validity of the proposed method was revealed as per ICH guidelines.

Development and Validation of Ecofriendly HPLC-MS Method for Quantitative Assay of Amoxicillin, Dicloxacillin, and Their Official Impurity in Pure and Dosage Forms.

Novel, accurate, selective, and rapid high-performance liquid chromatography mass spectrometry method was developed for simultaneous analysis of amoxicillin trihydrate, dicloxacillin sodium, and their official impurity 6-aminopenicillanic acid. The chromatographic separation was carried out by applying the mixture on a C18 column (3.5 µm ps, 100 mm × 4.6 mm id) using acetonitrile:water (65 : 35 by volume) as a mobile phase within only 4 min. The quantitative analysis was executed using single quadrupole mass spectrometer in which electrospray ionization, selected ion monitoring, and negative mode were operated. The retention times were 1.61, 2.54, and 3.50 mins for amoxicillin, 6-aminopenicillanic acid, and dicloxacillin, respectively. The method was validated in linear ranges of 2-28 µg mL-1, 2-35 µg mL-1, and 1-10 µg mL-1 for amoxicillin, dicloxacillin, and 6-aminopenicillanic acid, respectively. The results obtained from the suggested HPLC/MS were statistically compared with those obtained from the reported HPLC method, where no significant difference appeared respecting accuracy and precision. According to the analytical eco-scale assessment method, the proposed method was proved to be greener than the reported one, where the analysis time and the amount of the wasted effluent decreased.

Cinnamaldehyde and hesperetin attenuate TNBS-induced ulcerative colitis in rats through modulation of the JAk2/STAT3/SOCS3 pathway.

Ulcerative colitis is an autoimmune inflammatory disorder with a negative impact on the life quality of patients. Cinnamaldehyde and hesperetin were chosen due to their antioxidants and anti-inflammatory effects. This study explored the protective effects of cinnamaldehyde (40 and 90 mg/kg, po) and hesperetin (50 and 100 mg/kg, po) on 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced ulcerative colitis in rats. Cinnamaldehyde and hesperetin significantly improved macroscopic and histopathological examinations with a significant reduction in myeloperoxidase and intracellular adhesion molecule-1 expression. They significantly reduced colon oxidative stress by a significant elevation in both reduced glutathione content and superoxide dismutase activity with a significant reduction of NO content. Furthermore, cinnamaldehyde and hesperetin alleviated the inflammatory injury by a significant reduction in interleukin-6 along with suppression of nuclear factor-κB, receptor for advanced glycation end products, and tumor necrosis factor-α expression. Moreover, cinnamaldehyde and hesperetin significantly decreased p-JAK2 and p-STAT3 while significantly increased suppressors of cytokine signaling 3 (SOCS3) protein expression. In conclusion, cinnamaldehyde and hesperetin counteracted TNBS-induced ulcerative colitis through antioxidant, anti-inflammatory properties as well as modulation of the JAk2/STAT3/SOCS3 pathway.

High-Performance Liquid Chromatography Method for Simultaneous Determination of Guaifenesin, Salbutamol Sulfate and Guaifenesin Impurity (Guaiacol).

A simple and reliable HPLC method is developed for isocratic separation of a ternary mixture of Salbutamol Sulfate (SAL), Guaifenesin (GUI) and its impurity Guaiacol (GUA) either in pure powder or in pharmaceutical formulation. Chromatographic separation was applied on a Hypersil GOLD CN column with a mobile phase consisting of 0.05 M KH2PO4 (containing 0.1% triethylamine, pH adjusted to 3.7 by phosphoric acid): methanol (60: 40 by volume) using 0.6 mL min-1 flow rate and detection of peaks at 275 nm at 25°C with run time around 6 min. The calibration plots were linear over the concentration ranges of 0.5-20, 0.5-30 and 0.1-10 µg mL-1 for SAL, GUI and GUA, respectively. ICH guidelines were used for the validation of presented method. The obtained results were compared with the results obtained from the reported HPLC method and no significant difference was found regarding accuracy and precision.

Validated spectral manipulations for determination of an anti-neoplastic drug and its related impurities including its hazardous degradation product.

Innovative and specific double dual wavelength, dual ratio subtraction spectrophotometric methods were carried out along with a successive ratio subtraction spectrophotometric method for determination of dacarbazine and its related impurities including toxic and hazardous ones. For determination of dacarbazine by the double dual wavelength method, the absorbance differences between 323 and 350 nm of the zero order absorption spectra of dacarbazine were used. The values of absorbance difference between 267.2 and 286.2 nm of the zero order spectra of 5-amino-imidazole-4 carboxamide were used for its determination by the dual ratio subtraction method. The zero order absorption spectrum of 2-azahypoxanthine at 235 nm was used for its determination after applying the successive ratio subtraction method. ICH guidelines were followed for validation of the developed methods, where linear relationships were obtained in the range of 4-20, 1-16, and 2-20 µg mL-1 for dacarbazine, 5-amino imidazole-4-carboxamide and 2-azahypoxanthine, respectively. Accurate, precise, and specific results were obtained upon applying the proposed methods according to ICH guidelines. Furthermore, the developed methods were successfully applied for determination of dacarbazine in its pharmaceutical formulation. Comparing the results of the developed methods with those of the official USP spectrophotometric method statistically showed no significant difference. The developed methods don't need any sophisticated techniques so they are considered cost effective methods. Moreover, the introduced methods have the advantages of being green where water was used as a solvent. The methods proved to be more economic, fast and simple than other reported HPLC methods.

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

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

Green analysis of newly approved binary omeprazole/aspirin mixture in presence of aspirin impurity using ultra-high-performance liquid chromatography and thin-layer chromatography methods.

Two green, simple, and accurate chromatographic methods were developed and validated for the simultaneous determination of omeprazole and aspirin mixture in the presence of salicylic acid, a major impurity of aspirin. Method A is a reversed-phase ultra-high-performance liquid chromatography; the separation was performed on a C18 column, with a mobile phase composed of ethanol:0.1% aqueous solution of triethylamine acidified with orthophosphoric acid (pH 3) (30:70, v/v) at 0.15 mL/min flow rate and 230 nm. Omeprazole, aspirin, and aspirin impurity retention times were 7.47, 4.40, and 5.13 min, respectively. Good linearity was achieved in the concentration ranges of 5-80, 5-85, and 3-50 µg/mL for the three mentioned components, respectively. Method B is thin-layer chromatography (TLC) where silica gel TLC F254 plates were utilized to achieve separation using ethanol:ethyl acetate (2:8, v/v) as a developing system at 240 nm. The resulted Rf values were 0.83, 0.65, and 0.23 for omeprazole, aspirin, and impurity, respectively. The concentration ranges of 0.1-3 µg/band for the three drugs showed good linearity. The proposed methods are eco-friendly and greener when compared to the already reported method (Microchemical Journal, 152, 104350). This is the first use of TLC method for the determination of the three drugs. International Council for Harmonization (ICH) guidelines were followed to ensure the validity of developed methods.

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

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

Development and Validation of RP-HPLC and an Ecofriendly HPTLC Method for Simultaneous Determination of Felodipine and Metoprolol Succinate, and their Major Metabolites in Human Spiked Plasma.

BACKGROUND: Felodipine is a calcium channel blocker used together with metoprolol succinate for treatment of hypertension. OBJECTIVE: Two chromatographic methods were developed for simultaneous determination of felodipine (FEL) and metoprolol succinate (MET), and their major metabolites, dehydrofelodipine and metoprolol acid, respectively. METHODS: The first method was RP-HPLC which comprised separation of the studied components by gradient elution using a Phenomenex C8 column and a mobile phase composed of water (adjusted to pH 3.5 with o-phosphoric acid)-acetonitrile - methanol (45:40:15, by volume) for the first 6 min and (30:60:10, by volume) for the next 4 min at a flow rate of 1 mL/min followed by UV detection of the eluted peaks at 225 nm. The second method was an HPTLC method where separation was achieved using a mobile phase consisting of toluene-ethyl acetate-methanol-ammonia-formic acid (10:5:2.5:0.3:0.1, by volume) and scanning of the separated bands at 225 nm. RESULTS: Validation of the developed methods was done according to ICH guidelines. Successful application of the developed methods was carried out for determination of the studied drugs in human spiked plasma and in Logimax® tablets. CONCLUSIONS: The developed RP-HPLC and HPTLC methods can be further applied for quality control testing of the studied drugs. HIGHLIGHTS: RP-HPLC and HPTLC methods for determination of FEL, MET and their major metabolites. The developed methods were successfully applied for determination of FEL and MET in Logimax® tablets.

Development and Validation of Two Chromatographic Methods for Simultaneous Determination and Quantification of Amiloride Hydrochloride, Hydrochlorothiazide, and Their Related Substances, in Pure and Tablet Forms.

BACKGROUND: Amiloride hydrochloride (AM) is a potassium sparing diuretic, while hydrochlorothiazide (HCZ) is the protype of thiazide diuretics. The combining of the studied drugs exhibits a synergistic effect. Moreover, HCZ prevents the potassium depletion side effect caused by AM. OBJECTIVE: Two accurate and precise simultaneous chromatographic separation methods were promoted and investigated to quantify AM, HCZ, official impurities of HCZ (cholorothiazide and salamide), and the official impurities of AM (methyl 3, 5-diamino-6-chloropyrazine-2-carboxylate). METHODS: The components of the quintuple mixture were quantified by two methods. The first method was high-performance thin layer chromatography (HPTLC), where exemplary separation was achieved on silica gel HPTLC F254 plates at the stationary phase using ethyl acetate-ethanol-ammonia solution (8 + 2 + 0.2, v/v) as a developing system. Scanning of bands at 273 nm was done. The second method was a reversed-phase chromatography (RP-HPLC) method using C18 (4.6 × 100 mm) column and mobile phase comprising 0.1% phosphoric acid solution-acetonitrile (90 + 10, v/v) with UV determination at 273 nm. Adjustment of the flow rate at 1 mL/min and pH at 3.6 was performed. RESULTS: Regarding RP-HPLC, optimum separation of the quintuple mixture was achieved within just five minutes. According to HPTLC, symmetric and sharp peaks were separated on the resulted chromatogram. Validity of the introduced methods was investigated by applying international conference on harmonization (ICH) guidelines. CONCLUSIONS: The methods were successfully applied for assays of the studied drugs in their pure and tablet forms. No significant difference was revealed through application of statistical comparison between results of the suggested methods and those of the reported method regarding both accuracy and precision.

Capacitive sensor based on molecularly imprinted polymers for detection of the insecticide imidacloprid in water.

This manuscript reports on the development of a capacitive sensor for the detection of imidacloprid (IMD) in water samples based on molecularly imprinted polymers (MIPs). MIPs used as recognition elements were synthesized via a photo-initiated emulsion polymerization. The particles were carefully washed using a methanol (MeOH) /acetic acid mixture to ensure complete template removal and were then dried. The average size of the obtained particles was less than 1 µm. The imprinting factor (IF) for IMD was 6 and the selectivity factor (α) for acetamiprid, clothianidin, thiacloprid and thiamethoxam were 14.8, 6.8, 7.1 and 8.2, respectively. The particles were immobilized on the surface of a gold electrode by electropolymerization. The immobilized electrode could be spontaneously regenerated using a mixture of MeOH/10 mM of phosphate buffer (pH = 7.2)/triethylamine before each measurement and could be reused for 32 times. This is the first-time that automated regeneration was introduced as part of a sensing platform for IMD detection. The developed sensor was validated by the analysis of artificially spiked water samples. Under the optimal conditions, the linearity was in the range of 5-100 µM, with a limit of detection (LOD) of 4.61 µM.