Urszula Hubicka

Separation and characterization of ciprofloxacin, difloxacin, lomefloxacin, norfloxacin, and ofloxacin oxidation products under potassium permanganate treatment in acidic medium by UPLC-MS/MS.

A simple, sensitive and reproducible ultra-performance liquid chromatography method for determination of ciprofloxacin, difloxacin, lomefloxacin, norfloxacin and ofloxacin oxidation stability under permanganate treatment in acidic conditions at pH from 3.0 to 6.0, was developed. Chromatographic separations were carried out using the Acquity UPLC BEH C18 column; (2.1×100 mm, 1.7 μm particle size). The column was maintained at 40°C, and eluted under isocratic conditions using 83% of eluent A and 17% of eluent B over 6.5 min, at a flow rate of 0.3 mL min(-1). Eluent A: water/formic acid (0.1 v/v%); eluent B: acetonitrile/formic acid (0.1 v/v%). An oxidation process followed kinetic of the second order reaction and depended upon solution acidity. Oxidation of fluoroquinolones proceeded at piperazine moiety yielding respective hydroxy and oxo analogs, and remaining the quinolone fragment intact. Structures of products formed were assigned on a basis of UPLC/MS/MS fragmentation pathways.

THIN-LAYER CHROMATOGRAPHY WITH DENSITOMETRY FOR THE DETERMINATION OF DIFLOXACIN AND ITS PHOTODEGRADATION PRODUCTS. KINETIC EVALUATION OF THE DEGRADATION PROCESS AND IDENTIFICATION OF PHOTOPRODUCTS BY MASS SPECTROMETRY

TLC-densitometric method was developed for determination of difloxacin (DIF) in the presence its photodegradation products. Silica gel TLC F254 plates were used as the stationary phase and methylene chloride:methanol:2-propanol:ammonia 25% (4:4:5:2, v/v/v/v) as the mobile phase. The elaborated method meets the acceptance criteria for specificity, linearity, sensitivity, accuracy, and precision. The photodegradation process of DIF followed kinetics of the first order reaction for the substrate. Potential photodegradation products of DIF identified by UPLC-MS/MS are: 7-(2-aminoethylamino)-6-fluoro-1-(4-fluorophenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid, 7-amino-6-fluoro-1-(4-fluorophenyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid, 6-fluoro-1-(4-fuorophenyl)-7-(3-hydroxypiperazin-1-yl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid, 6-fluoro-1-(4-fuorophenyl)-4-oxo-7-(piperazin-1-yl)-1,4-dihydroquinoline-3-carboxylic acid.

Determination of sparfloxacin and its photodegradation products by thin-layer chromatography with densitometry detection. Kinetic evaluation of the degradation process and identification of photoproduct by mass spectrometry

A TLC-densitometric method was developed for the determination of sparfloxacin in the presence of its photodegradation products. Silica gel TLC F254 plates were used as the stationary phase and methylene chloride : methanol : 2-propanol : ammonia 25% (4 : 4 : 5 : 2 v/v/v/v) as the mobile phase. The developed method meets the acceptance criteria for specificity, linearity, sensitivity, accuracy, and precision. The linear regression analysis for the calibration curve showed a good linear correlation over the concentration range 1.00–3.00 μg per band, with the determination coefficient, R2, exceeding 0.9958. The method was shown to have good precision and intermediate precision, as reflected by the relative standard deviation values, lower than 2.72% and characterized by a recovery rate at three concentration levels from 93.60% to 103.90%. The limits of detection and quantification were respectively 0.16 and 0.48 μg per band. The photodegradation process of sparfloxacin followed the kinetics of a first order reaction for the substrate. Nine products of photodegradation were identified by UPLC-MS/MS. The structure of a product previously not identified was determined.

Lipophilicity study of fifteen fluoroquinolones by reversed-phase thin-layer chromatography

The lipophilicity of fifteen fluoroquinolones has been investigated. Reversed-phase thin layer chromatography with densitometric detection was applied to determine the RM0 factor. The RP-TLC investigations were performed in mixtures of organic modifier–water. The partition coefficients of examined fluoroquinolones were also calculated with computational programs. All the obtained data, from experimental methods and theoretical calculations, were compared and a suitable conclusion was reached. The received results demonstrate that the RP-TLC method may be the only reliable technique for fluoroquinolones in describing their lipophilic nature as well as the activity.

Determination of danofloxacin and its photodegradation products by HPLC-DAD. Kinetic evaluation of the degradation process and identification of photoproducts by mass spectrometry

A HPLC method was developed for determination of danofloxacin (DAN) in the presence of its photodegradation products. Chromatography was performed on a Gemini-NX C18 110A, 150 mm × 4.60 mm, 3 μm particle size column with 0.025 M phosphate buffer (pH = 5.00)–acetonitrile–methanol (95 : 10 : 30 v/v/v) as the mobile phase at a flow rate of 1.2 mL min−1. UV detection was performed at 280 nm. The column was thermostatted at 25 °C. The elaborated method meets the acceptance criteria for specificity, linearity, sensitivity, accuracy, and precision. The linear regression analysis for the calibration curve showed a good linear correlation over the concentration range 0.20–0.80 mg mL−1, with determination coefficient, R2, exceeding 0.9966. The method was shown to have good and intermediate precision, as reflected by the relative standard deviation values, lower than 2.21% and characterized by a recovery rate at three concentration levels from 98.0% to 101.70%. The limits of detection and quantification were respectively 0.0055 and 0.0167 mg mL−1. The photodegradation process of DAN followed kinetics of the first order reaction for the substrate. Ten products of photodegradation were identified by UPLC/MS/MS.

DETERMINATION OF DANOFLOXACIN AND ITS PHOTODEGRADATION PRODUCTS BY THIN-LAYER CHROMATOGRAPHY: KINETIC EVALUATION OF DEGRADATION PROCESS

TLC-densitometric method was developed for determination of danofloxacin (DAN) in presence of its photodegradation products. Silica gel HPTLC Lichrospher 60 F254 plates were used as the stationary phase and methanol:acetone:1 M citric acid:triethylamine (2.8:2:0.2:0.5, v/v/v/v) as the mobile phase. The elaborated method meets the acceptance criteria for specificity, linearity, sensitivity, accuracy, and precision. The linear regression analysis for the calibration curve showed a good linear correlation over the concentration range 1.25– 3.75 µg per band, with determination coefficient, exceeding 0.9806. Accuracy of the method expressed as % recovery at three concentration levels was from 95.56% to 97.30%. Good precision and intermediate precision with % RSD less than 0.92% was also observed. The limits of detection and quantification were respectively 1.10 and 3.40 ng per band. Satisfactory results of validation of the method were also confirmed by determination of DAN in veterinary commercial preparation. The photodegradation process of DAN followed kinetics of the first order reaction for the substrate.

TLC-DENSITOMETRIC DETERMINATION OF TOLPERISONE AND ITS IMPURITIES 4-METHYLPROPIOPHENONE AND PIPERIDINE IN PHARMACEUTICAL PREPARATIONS

A chromatographic-densitometric method was developed for determination of tolperisone hydrochloride and its impurities. Silica gel TLCF 254 plates were used as the stationery phase and cyclohexane-1,4 dioxane-isopropanol–ethanol-glacial acetic acid (16:0.5:1:4:0.6 v/v/v) and n-butanol-isopropanol-water-glacial acetic acid (10:7:8:2 v/v/v/v) as the mobile phases. Densitometric measurements were done at 260 nm for tolperisone hydrochloride and 4-methylpropiophenone and at 570 nm for piperidine hydrochloride. The method is specific, sensitive, precise, and has a broad range of linearity, with potential application in pharmaceutical analysis.

The impact of ZnO and TiO2 on the stability of clotrimazole under UVA irradiation: Identification of photocatalytic degradation products and in vitro cytotoxicity assessment

Graphical abstract Figure. No Caption available. HighlightsPhotocatalytic degradation of clotrimazole by TiO2/ZnO under UVA irradiation.UPLC method development for determination of clotrimazole in the presence of its photocatalytic degradation products.Structural characterization of fourteen presumable photodegradation products of clotrimazole using UPLC–MS/MS.In vitro cytotoxic assessment using the Human skin fibroblast cell line. Abstract In order to ensure the safe and effective use of pharmaceutical products especially for topical administration photostability testing is necessary. The current paper presents an in‐depth analysis of the stability of one of the most common antifungal agents, namely clotrimazole. Clotrimazole has proven to be stable under UVA irradiation in applied experimental conditions, but the presence of catalysts such as ZnO and TiO2 has contributed significantly to the degradation of this compound. The findings indicate that its photocatalytic degradation reactions followed the pseudo first‐order kinetics with rate constant depending on the pH and the used solvent. Using LC–MS/MS, 14 presumable degradation products of clotrimazole were identified and the plausible transformation pathways were proposed. The in vitro cytotoxicity risk evaluation based on photostability of clotrimazole was also performed using the Human skin fibroblast cell line (BJ) ATCC™ CRL‐2522. There was no statistically significant difference between cells viability in all analyzed combinations of clotrimazole, TiO2/ZnO, and UVA irradiation (p < 0.05).

Chromatographic-Densitometric Analysis of Chosen Fluoroquinolones on TLC Plates Using Mobile Phases with Different Viscosity

TLC-densitometric method was developed for the identification and quantitative determination of selected fluoroquinolones. Silica gel TLC 60 F254 plates were used as the stationary phase. For the separation of examined compounds the suitability of the following four mobile phases with different dynamic viscosity was checked: (I) methanol:ammonia:ethyl acetate:acetonitrile 1:1:2:1 v/v/v/v (1.70 mPa · s), (II) dichloromethane:methanol:isopropanol:ammonia 4:4:5:2 v/v/v/v (2.08 mPa · s), (III) dichloromethane:ethanol:toluene:ammonia:n-butanol:water 6:6:2:1.8:3:0.3 v/v/v/v/v/v (2.27 mPa · s), (IV) 1,4- dioxane:ammonia:tetrahydrofuran 6:3:2 v/v/v (3.24 mPa · s). The calculated values of retardation factors RF, capacity factors k, selectivity factors α, resolution factors Rs, and also asymmetry factors As for the evaluation of separation conditions were used. The obtained results confirmed that dynamic viscosity of the mobile phases is essential and influences the retention parameters of the studied fluoroquinolone chemotherapeutics. The validation results for the mobile phase IV indicate its suitability for the identification and also quantitative analysis of selected drugs in pharmaceutical analysis.