Insights of different analytical approaches for estimation of budesonide as COVID-19 replication inhibitor in its novel combinations: green assessment with AGREE and GAPI approaches.

Simple, direct, rapid, and sensitive HPLC and spectrophotometric methods were established for simultaneous estimation of a novel combination of budesonide and azelastine (BUD/AZL) in their laboratory-prepared mixture and dosage form according to the medicinally recommended ratio 1:4.28. Budesonide is an important inhalation corticosteroid that plays a vital role in the inhibition of COVID-19 replication and cytokine production. The first chromatographic method was created for the simultaneous estimation of BUD epimers in the presence of AZL with excellent efficiency in a relatively short chromatographic run (< 9 min). The separation of BUD epimers with AZL was carried out on a C18 column using acetonitrile: phosphate buffer of pH 3.5 adjusted by 0.2 M orthophosphoric acid (40:60, v/v) as a mobile phase, UV detection at 230 nm and a flow rate of regulated at 2 mL/min. Besides, three spectrophotometric methods were applied for the simultaneous determination of the provided mixture adopting zero order, first order derivative, and ratio first derivative approaches. The Zero-order spectrophotometry was used for the determination of AZL in presence of BUD, where BUD shows no absorbance at 290 nm. The first derivative amplitude at 265 nm (1D265) (zero-crossing of AZL) and the ratio of first derivative amplitudes at 270 nm (1DD270) using 10.0 µg mL-1 AZL as divisor was chosen for the simultaneous determination of BUD in the presence of AZL in the binary mixture. The proposed methods were found to be rectilinear in the concentration range of (0.4-40.0 µg mL-1) and (0.05-40.0 µg mL-1) for BUD and AZL, respectively in the HPLC method. Whereas the concentration range for AZL in the zero-order method was (1.0-35.0 µg mL-1) and for BUD in the first derivative and ratio derivative method was (6.0-20.0 µg mL-1). Validation of the suggested approaches according to the ICH criteria was performed. Furthermore, to ensure the proposed approaches' greenness, The AGREE and GAPI metrics were utilized, and the afforded results revealed an excellent greenness of the proposed approaches.

Isocratic ion pair chromatography for estimation of novel combined inhalation therapy that blocks coronavirus replication in chronic asthmatic patients.

A rapid and sensitive isocratic ion-pair chromatographic method was developed for the accurate analysis of ternary mixtures of formoterol, tiotropium, and ciclesonide in their novel combined inhalation that is widely used for the symptomatic treatment of patients with chronic obstructive disease. Analytical separation was performed using a C8 column and ion pair mobile phase composed of acetonitrile: acidified deionized water (55: 45% v/v) containing 0.025% sodium dodecyl sulfate. The pH was adjusted to 3.0 using orthophosphoric acid and eluted isocratically at 2.0 mL/min and 40 °C applying UV detection at 237 nm. The calibration ranges were found to be 0.3-9.0 µg/mL for formoterol, 0.45-13.5 µg/mL for tiotropium, and 10.0-300.0 µg/mL concerning ciclesonide. The proposed method exhibited good repeatability, accuracy, and sensitivity (R.S.D. < 2.0%). The approach is rapid (run time does not exceed 15 min) and achieves satisfactory resolution (resolution factors = 7.45 and 5.3 between formoterol and tiotropium and tiotropium and ciclesonide respectively). The sensitivity and the efficiency of the proposed method permit their successful estimation with a recovery percentage ± SD of 99.33% ± 0.43 for formoterol, 99.15% ± 0.60 for tiotropium, and 99.90% ± 0.41 for ciclesonide.

Multiple analytical methods for determination of formoterol and glycopyrronium simultaneously in their novel combined metered dose inhaler.

One of the major causes of mortality all over the world is chronic obstructive pulmonary disease (COPD). Recently approved combined inhaler of formoterol fumarate (FF) and glycopyrronium bromide (GLY) has been used in very low concentrations (µg level/actuation) doses in COPD patients. The first spectrophotometric and advanced highly sensitive liquid chromatography has been achieved successfully throughout this study, permitting validated analysis of dual combined inhaler in raw material as well as pharmaceutical inhaled dosage form. Three sensitive analytical methods were carried out for the simultaneous assay of FF and GLY in their novel combined Metered dose inhaler (MDI). The first method depends on measuring the first derivative amplitudes at 208.27 nm for FF and at 213.27 nm and 239.86 nm for GLY, respectively. The second method depends on measurement of the first derivative of the ratio spectra at 214 or 229 nm for FF and 240 or 259 nm for GLY, respectively. For the spectrophotometric methods, the linearity ranges were 0.48-9.6 µg/mL for FF and 0.9-18 µg/mL for GLY. For the third method, valid ion-pairing chromatographic method was carried out applying C18 column and isocratic mobile phase of 60% v/v acetonitrile and 40% v/v deionized waster (pH 3.0) enclosing 0.025% sodium dodecyl sulfate, using UV detection adjusted to 210 nm and flow rate of 1.2 mL/min. For the ion-pairing chromatographic method, the linearity ranges were 0.048-4.8 µg/mL for FF and 0.09-9.0 µg/mL for GLY. The developed methods are reproducible, valid and offer efficient resolution between formoterol and glycopyrronium using spectrophotometric methods and highly sensitive and precise chromatographic method. The percent recoveries of the inhaled drugs in their MDI were good. The method was successfully established for the quantitative analysis of FF and GLY in their combined pharmaceutical inhaler capsules to validate the therapeutic efficiency of the combined drugs in quality control labs.

Application of derivative emission fluorescence spectroscopy for determination of ibuprofen and phenylephrine simultaneously in tablets and biological fluids.

Two sensitive, rapid, and accurate derivative emission spectrofluorimetric methods applying zero crossing techniques were developed for simultaneous determination of binary mixtures of ibuprofen (IBU) and phenylephrine hydrochloride (PHE) in pure powder, synthetic mixture and combined tablets. The proposed methods were performed via measuring the intersected drug derivative amplitude of one drug at the zero crossing points for the other one and vice versa. The two methods rely on the measurement of the combined drugs native fluorescence after excitation at 270 nm in methanol directly, followed by differentiation using first (D1) and second derivative (D2) techniques. Applying the D1, IBU was measured quantitatively at 293.1 nm at zero crossing of PHE, on the other side; PHE was measured quantitatively at 300.7 nm at zero crossing of IBU. By the same way, applying the D2, the wavelengths selected were 303.5 nm for IBU and 312.9 nm for PHE. The concentration plots of derivative fluorescence intensity were rectilinear over the range of 0.5-10 µg/mL and 0.025-0.5 µg/mL for IBU and PHE, respectively. The results obtained with average % recoveries ±â€¯RSD are 99.73 ±â€¯0.72 (IBU, D1), 99.49 ±â€¯0.95 (PHE, D1), 99.79 ±â€¯0.47 (IBU, D2), and 99.88 ±â€¯0.34 (PHE, D2) were in good agreement with the comparison method. The proposed methods offer high sensitivity that enable direct analysis of IBU and PHE in spiked human plasma. The proposed methods were entirely validated in terms of ICH guidelines.

Eco-Friendly Green Liquid Chromatographic Determination of Azelastine in the Presence of its Degradation Products: Applications to Degradation Kinetics.

Background: Green solvents such as microemulsion were used in the proposed method because they play a vital role in the analytical method's influence on the environment. Objective: A highly sensitive, specific, and validated stability-indicating eco-friendly green microemulsion liquid chromatography (MELC) method was developed for separation of the antihistaminic drug Azelastine HCl (AZL) from its degradation products with application to degradation kinetics. Methods: Chromatographic separation was operated on a C18 column with a microemulsion mobile phase, which consists of 0.1 M sodium dodecyl sulphate, 10% n-propanol, 1% n-octanol, and 0.3% triethylamine, by using 0.02 M phosphoric acid at pH 3.5 and irbesartan as internal standard. The eluted compounds were monitored at 210 nm with flow rate 1 mL/min at ambient temperature. Results: A linear dependence of the peak area on drug concentration over the concentration range of 0.1 to 25 µg/mL was achieved with an LOD of 0.04 µg/mL and an LOQ of 0.10 µg/mL. Moreover, the proposed method was successfully applied for determination of AZL in eye drops and metered dose nasal inhaler as well as to study the kinetics of alkaline, acidic, neutral, oxidative, and photolytic degradation processes of AZL according to the International Council for Harmonization guidelines. Conclusions: The proposed method could be used as a harmless alternative for quality control analysis of the mentioned drug, without interference from dosage form additives or decomposition products. Highlights: A highly sensitive stability-indicating eco-friendly green MELC method was developed for the separation of the antihistaminic drug AZL from its degradation products.

Validated ion pair chromatographic method for simultaneous determination and in vitro dissolution studies of indacaterol and glycopyrronium from inhaled capsule dosage form.

A rapid, and highly sensitive analytical method were developed for the simultaneous determination of indacaterol maleate (IND) and glycopyrronium bromide (GLY) in their inhaler capsules. Valid ion-pairing chromatographic (IPC) method was performed for separation of GLY in presence of IND using C18 column and mobile phase consisting of acetonitrile: acidified deionized water (60:40% v/v) containing 0.02% sodium dodecyl sulfate (SDS) adjusted to pH 3.0 using OPA (orthophosphoric acid) isocratically eluted at 2.0 mL/min. Quantitation was achieved with UV detection at 210 nm. Cyproheptadine was used as an internal standard. The retention times were 1.9 and 2.5 min for IND, and GLY, respectively. For the IPC method, the calibration graphs were linear in the range of 0.66-66.0 µg/mL for IND and 0.3-30.0 µg/mL for GLY. The proposed method are rapid, reproducible (R.S.D. <2.0%) and achieves satisfactory resolution between IND and GLY (resolution factor = 4.23). The mean recoveries of the analytes in their inhaler capsule were satisfactory. It was applied successfully to in vitro dissolution testing using Franz diffusion cell and extended to a content uniformity test consistent with the United States Pharmacopoeia (USP) guidelines and were found to be precise and accurate for the capsules studied with acceptance value of 4.53 and 1.39 for IND and GLY, respectively.

Simultaneous Determination of Gatifloxacin and Prednisolone in their Bulk Powder, Synthetic Mixture and Their Combined Ophthalmic Preparation Using Micellar Liquid Chromatography.

A simple rapid and accurate micellar high performance liquid chromatographic method was improved and validated for the analysis of mixture containing gatifloxacin sesquihydrate (GTF) and prednisolone acetate (PRED) in their synthetic mixture and their combined preparation. The separation was achieved using a C18 column, micellar mobile phase consisted of 0.2 M sodium dodecyl sulfate, 12.5% n-propanol and 0.3% triethylamine in 0.02 M orthophosphoric acid at pH 7.0 at a flow rate of 1 ml/min with UV detection at 270 nm. The proposed method was found to be rectilinear over the concentration ranges of 5.0-45 µg ml-1 and 10-50 µg ml-1 with recovery percentage of 99.95 ± 0.82 and 100.07 ± 0.84 for GTF and PRED, respectively. The separation of both drugs was accomplished in a very short chromatographic run (<5 min), the method is reproducible (R.S.D. < 1.0%) and show satisfactory resolution between GTF and PRED (Rs) = 1.67. The developed method was validated according to International Conference on Harmonization (ICH) guidelines. The limit of detection of the proposed method was 0.33 and 0.21 µg ml-1, and the limit of quantitation was 0.99 and 0.64 µg ml-1 for GTF and PRED, respectively.

Validated spectroscopic methods for determination of anti-histaminic drug azelastine in pure form: Analytical application for quality control of its pharmaceutical preparations.

Two simple, sensitive, rapid, validated and cost effective spectroscopic methods were established for quantification of antihistaminic drug azelastine (AZL) in bulk powder as well as in pharmaceutical dosage forms. In the first method (A) the absorbance difference between acidic and basic solutions was measured at 228nm, whereas in the second investigated method (B) the binary complex formed between AZL and Eosin Y in acetate buffer solution (pH3) was measured at 550nm. Different criteria that have critical influence on the intensity of absorption were deeply studied and optimized so as to achieve the highest absorption. The proposed methods obeyed Beer's low in the concentration range of (2.0-20.0µg·mL-1) and (0.5-15.0µg·mL-1) with % recovery±S.D. of (99.84±0.87), (100.02±0.78) for methods (A) and (B), respectively. Furthermore, the proposed methods were easily applied for quality control of pharmaceutical preparations without any conflict with its co-formulated additives, and the analytical results were compatible with those obtained by the comparison one with no significant difference as insured by student's t-test and the variance ratio F-test. Validation of the proposed methods was performed according the ICH guidelines in terms of linearity, limit of quantification, limit of detection, accuracy, precision and specificity, where the analytical results were persuasive.

Synthesis, antimicrobial, antiquorum-sensing, antitumor and cytotoxic activities of new series of cyclopenta(hepta)[b]thiophene and fused cyclohepta[b]thiophene analogs.

New series of cyclopenta(hepta)[b]thiophene and fused cyclohepta[b]thiophene analogs were synthesized. The new analogs were assessed for antibacterial efficacy toward Escherichia coli ATCC 12435, Bacillus cereus UW 85 and Staphylococcus aureus. Compounds 5a, 6b and 12 showed eminent activity toward all selected bacterial strains compared to ampicillin. The antifungal efficacy of the same analogs was also examined toward Candida albicans and Aspergillus fumigatus 293, whereas 5a,b and 12 showed excellent efficacy toward both of the tested fungi. Moreover, 4b, 6a, 14a and 17 demonstrated interesting antifungal efficacy toward A. fumigatus. The same analogs were assessed for antiquorum-sensing efficacy toward Chromobacterium violacium ATCC 12472, whereas 5a, 12 and 15a demonstrated moderate activity. The new analogs were also esteemed for in vitro antitumor activity over HepG2, MCF-7 and HT-29 cancer cell lines. Results indicated that 6b and 10 are the most potent analogs against the three tested cell lines. In addition, 5a, 6a, 7 and 15a displayed interesting activity toward all tested cell lines. The active in vitro antitumor analogs were screened for in vivo antitumor activity over EAC in mice as well as in vitro cytotoxicity toward W138 human normal cell line. Results demonstrated that 6a,b and 10 have the highest in vivo activity, and that all tested compounds were found to be less cytotoxic than 5-FU toward W138 normal cell line. The DNA-binding affinity of the active antimicrobial and/or antitumor analogs was also assessed, whereas 4a, 5b, 10 and 15a exhibited the highest affinity. In silico studies affirmed that the inspected compounds are compatible with Lipinski's rule of five with expected good oral absorption.

Validated sensitive spectrofluorimetric method for determination of antihistaminic drug azelastine HCl in pure form and in pharmaceutical dosage forms: application to stability study.

A highly sensitive, simple and rapid spectrofluorimetric method was developed for the determination of azelastine HCl (AZL) in either its pure state or pharmaceutical dosage form. The proposed method was based on measuring the native fluorescence of the studied drug in 0.2 M H2 SO4 at λem  = 364 nm after excitation at λex  = 275 nm. Different experimental parameters were studied and optimized carefully to obtain the highest fluorescence intensity. The proposed method showed a linear dependence of the fluorescence intensity on drug concentration over a concentration range of 10-250 ng/mL, with a limit of detection of 1.52 ng/mL and limit of quantitation of 4.61 ng/mL. Moreover, the method was successfully applied to pharmaceutical preparations, with percent recovery values (± SD) of 99.96 (± 0.4) and 100.1 (± 0.52) for nasal spray and eye drops, respectively. The results were in good agreement with those obtained by the comparison method, as revealed by Student's t-test and the variance ratio F-test. The method was extended to study the stability of AZL under stress conditions, where the drug was exposed to neutral, acidic, alkaline, oxidative and photolytic degradation according to International Conference on Harmonization (ICH) guidelines. Copyright © 2016 John Wiley & Sons, Ltd.

Simultaneous determination of enalapril and hydrochlorothiazide in pharmaceutical preparations using microemulsion liquid chromatography.

A rapid high-performance liquid chromatography procedure for analytical quality control of mixture containing enalapril maleate (ENM) and hydrochlorothiazide (HCT) in their pharmaceutical preparations was developed using a microemulsion as an eluent. The separation was performed on a column packed with cyano-bonded stationary phase adopting UV detection at 210 nm using a flow rate of 1 mL/min. The optimized microemulsion mobile phase consisted of 0.2 M sodium dodecyl sulfate, 1% octanol, 10% n-propanol and 0.3% triethylamine in 0.02 M phosphoric acid, and pH was adjusted at 3.5. The proposed method was found to be linear over the concentration ranges 1-100 and 0.05-5 µg/mL for ENM and HCT, respectively with a correlation coefficient of 0.9999 for both drugs. The developed method was validated in terms of specificity, linearity, lower limit of quantification, lower limit of detection, precision and accuracy. The proposed method is rapid (5 min), reproducible (relative standard deviation <2.0%) and achieves a satisfactory resolution between ENM and HCT (resolution factor = 3.62). The mean recoveries of the analytes in tablets were in agreement with those obtained from a comparison method, as revealed by statistical analysis of the obtained results using Student's t-test and the variance ratio F-test.

Microemulsion liquid chromatographic method for simultaneous determination of simvastatin and ezetimibe in their combined dosage forms.

A rapid HPLC procedure using a microemulsion as an eluent was developed and validated for analytical quality control of antihyperlipidemic mixture containing simvastatin (SIM) and ezetimibe (EZT) in their pharmaceutical preparations. The separation was performed on a column packed with cyano bonded stationary phase adopting UV detection at 238 nm using a flow rate of 1 mL/min. The optimized microemulsion mobile phase consisted of 0.2 M sodium dodecyl sulphate, 1% octanol, 10% n-propanol, and 0.3% triethylamine in 0.02 M phosphoric acid at pH 5.0. The developed method was validated in terms of specificity, linearity, lower limit of quantification (LOQ), lower limit of detection (LOD), precision, and accuracy. The proposed method is rapid (8.5 min), reproducible (RSD < 2.0%) and achieves satisfactory resolution between SIM and EZT (resolution factor = 2.57). The mean recoveries of the analytes in pharmaceutical preparations were in agreement with those obtained from a reference method, as revealed by statistical analysis of the obtained results using Student's t-test and the variance ratio F-test.

Synthesis and antitumor activity of new sulfonamide derivatives of thiadiazolo[3,2-a]pyrimidines.

New series of sulfonamide derivatives of [1,3,4]thiadiazolo[3,2-a]pyrimidine were synthesized and investigated as antitumor agents. Some of the newly prepared compounds were tested for their in vitro and in vivo antitumor activities. Preliminary biological studies revealed that compounds 4c, 4f, and 4j exhibited the highest affinity to DNA, while compounds 4h,i, 6a-c, 8 and 12-14 exhibited moderate activity. Also, compounds 4j, 4f and 4c showed the highest percentage increase in lifespan of mice inoculated with Ehrlich ascites cells over 5-flurouracil (positive control). The detailed synthesis, spectroscopic and biological data are reported.

Simultaneous determination of norfloxacin and tinidazole binary mixture by difference spectroscopy.

A simple and rapid difference spectroscopic method was developed for the simultaneous determination of binary mixture of norfloxacin (NF) and tinidazole (TZ) without prior separation. The proposed method depends upon measuring the absorbance of NF at 291.6 nm which is the zero crossing point on the difference spectra of TZ in 0.1 N NaOH vs. 0.1 N HCl. Similarly, the absorbance of TZ was measured at 344.4 nm which is the zero crossing point on the difference spectra of NF. Beer's law was obeyed in the concentration range of 2-20 and 5-50 µg/mL for NF and TZ, respectively. The lower limits of detection (LOD) of NF and TZ are 0.23 and 0.36 µg/mL, respectively, while the lower limits of quantification (LOQ) of NF and TZ were 0.70 and 1.08 µg/mL, respectively. The precision of the method was satisfactory; the maximum value of relative standard deviations did not exceed 1.5% (n=10). The accuracy, expressed as recovery is between 98.25 and 101.8% with relative error of 0.29 and 0.23 for NF and TZ, respectively. The proposed method was successfully applied for the determination of both drugs in bulk powder, laboratory prepared mixture and commercial dosage forms such as tablets without interference from the commonly encountered excipients and additives. The results obtained are in good agreement with those obtained by the reference methods.

Synthesis and anti-inflammatory activity of novel (substituted)benzylidene acetone oxime ether derivatives: molecular modeling study.

Herein, we report the design, synthesis, and pharmacological properties of a series of substituted benzylidene acetone oxime ether derivatives from the corresponding oxime derivatives. All the newly synthesized compounds were investigated in vivo for their anti-inflammatory activities using carrageenin-induced rat paw oedema model. Among the compounds examined, compounds 5b and 7a showed the highest activity, nearly equivalent to that of the standard drug diclofenac sodium. Hence, they were screened for their analgesic activities using acetic acid-induced writhing model in mice and also, their ulcerogenic effects were studied. Compound 7a was found to possess significant anti-inflammatory and analgesic activities with negligible ulcerogenic effect. Docking study of the synthesized compound 7a into the active site of COX-1 and COX-2 revealed a similar binding mode to SC-558, a selective COX-2 inhibitor.

Simultaneous determination of ritodrine and isoxsuprine using coupling technique of synchronous fluorimetry and H-point standard addition method.

Simultaneous determination of two structurally related ss(2) adrenergic receptor agonists namely, Ritodrine HCl (RTH) and Isoxsuprine HCl (ISP) was performed using coupling technique of synchronous fluorimetry and H-point standard addition method. Under optimum conditions, linear determination ranges were 1.48 - 14.80 x 10(-6) mol L(-1) and 1.54 - 15.44 x 10(-6) mol L(-1) for ISP and RTH respectively. RTH and ISP could be determined simultaneously without interference from each other when their concentration ratio varies from 5:1 to 1:5 in the mixed sample. The proposed method was applied to the determination of RTH and ISP in synthetic mixture of pharmaceutical samples, the accuracy and precision of the results were satisfactory.

Determination of lisinopril in dosage forms and spiked human plasma through derivatization with 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) followed by spectrophotometry or HPLC with fluorimetric detection.

Two sensitive, simple and specific methods based on spectrophotometry and reversed-phase HPLC with fluorimetric detection are described for the determination of lisinopril in dosage forms as well as in spiked human plasma using solid phase extraction (SPE) procedures. Both methods are based on the derivatization of lisinopril with 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) in borate buffer of pH 9 to yield a yellow, fluorescent product. The spectrophotometric method depends on measuring the formed yellow color at 470 nm after optimization of the reaction conditions. The HPLC method is based on measurement of the derivatized product using fluorescence detection at 540 nm (excitation at 470 nm). The separation of the derivatized drug, the excess reagent and the internal standard (bumetanide) was performed on a reversed-phase ODS column using isocratic elution with methanol-0.02 M sodium dihydrogen phosphate, pH 3.0 (55:45, v/v) at a flow rate of 1.0 ml/min. The calibration graphs were linear over the concentration ranges 2-20 or 0.02-3.2 microg/ml of lisinopril with minimum detectability of 0.3 and 0.008 microg/ml (6.1 x 10(-7) and 1.7 x 10(-8)M) for the spectrophotometric and the HPLC methods, respectively. The proposed methods were applied without any interference from the tablet excipients for the determination of lisinopril in dosage forms, either alone or co-formulated with hydrochlorothiazide. Furthermore, the use of the HPLC method was extended to the in vitro determination of the drug in spiked human plasma. Interference from endogenous amino acids has been overcomed by using the solid phase extraction technique, the percentage recovery (n=6) was 101.6+/-3.35.

Spectrophotometric determination of propranolol in formulations via oxidative coupling with 3-methylbenzothiazoline-2-one hydrazone.

A simple spectrophotometric method has been developed for the determination of propranolol hydrochloride in pure as well as in dosage forms. The method is based on the oxidative coupling reaction with 3-methylbenzothiazoline-2-one hydrazone. A mixture of an acidic solution of the chromogenic agent and the drug upon treatment with ceric ammonium sulfate produces an orange color peaking at 496 nm. The absorbance-calibration plot was linear over the range 1-10 microg/ml with minimum detectability (S/N=2) of 0.1 microg/ml (3.38x10(-7) M). The molar absorbitivity was 3.195x10(3) l/M/cm with correlation coefficient (n=10) of 0.9999. The different experimental parameters affecting the development and stability of the color were carefully studied and optimized. The proposed method was applied successfully to the determination of propranolol in its dosage forms. A proposal of the reaction pathway was presented.