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Development and validation of HPLC method for separation and determination of active substances in pharmaceutical preparation Valetol
Ullrichová, Jana ; Šatínský, Dalibor (advisor) ; Matysová, Ludmila (referee)
Development and validation of HPLC method for separation and determination of active substances in pharmaceutical preparation Valetol Jana Ullrichová Abstract A method for the determination of paracetamol, propyphenazone and caffeine by the high- performance liquid chromatographic method with ultraviolet detection has been developed and validated. The analysis was performed at the room temperature in an isocratic mode on the reversed phase ODS Hypersil 5 µm C-18 column (250x4,6mm). A mobile phase (water : 2-propanol : diethylamine : methanol (50 + 15 + 3 + 32) adjusted to pH 7.5 by means of phosphoric acid) was suitable for the separation and the determination of paracetamol, caffeine and propyphenazone. UV detection was applied at 273 nm. Injected volume was 5 µl, flow rate 0,5 ml.min-1 . The developed method is sensitive and selective and can be applied for the routine studies of pharmaceuticals in the tablet form.
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Using of sequential injection chromatography method for separation and determination of salicylic acid and triamcinolone acetonide in pharmaceutical preparation Triamcinolon-IVAX
Holík, Pavel ; Šatínský, Dalibor (advisor) ; Solich, Petr (referee)
Using of sequential injection chromatography method for separation and determination of salicylic acid and triamcinolone acetonide in pharmaceutical preparation Triamcinolon-IVAX Pavel Holík Abstract A novel and fast simultaneous determination of triamcinolone acetonide (TCA) and salicylic acid (SA) in topical pharmaceutical formulations by sequential injection chromatography (SIC) as an alternative to classical high performance liquid chromatography (HPLC) has been developed. A recently introduced Onyx™ monolithic C18 (50 mm × 4,6 mm, Phenomenex® ) with 5 mm monolithic precolumn were used for the first time for creating sequential injection chromatography system based on a FIAlab® 3000 with a six-port selection valve and 5,0 ml syringe pump in study. The mobile phase used was acetonitrile/water (35:65, v/v), pH 3.3 adjusted with acetic acid at flow rate 15 µl.s-1 . UV detection provided by fibre-optic DAD detector was set up at 240 nm. Propylparaben was chosen as suitable internal standard (IS). There is only simple pre-adjustment of the sample of topical solution (dilution with mobile phase) so the analysis is not uselessly elongated. Parameters of the method showed good linearity in wide range, correlation coefficient >0,999; system precision (relative standard deviation, R.S.D.) in the range 0,45-1,95%...
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Determination of bopindolol by the technique of sequential injection analysis with chemiluminescence detection
Holá, Petra ; Polášek, Miroslav (advisor) ; Šatínský, Dalibor (referee)
The oxidation of bopindolol [ 4-(2-benzoyloxy-3-tert-butylaminopropyl)-2-methylindole (I)], by permanganate in the medium of aqueous H2SO4 is accompanied by the emission of chemiluminescence (CL) radiation. The CL signal is enhanced by hexametaphosphate. This CL reaction was used for devising automated sequential injection analysis (SIA) assay of I in pharmaceutical preparations. The PC-controlled SIA setup consisted of a Cavro XL 3000 2.5- ml syringe pump, Vici-Valco 10-port selection valve and Spectra-Physics FS970 flow- through fluorescence detector equipped with a lab-made CL detection module. The net CL signal of I increased by a factor of 3 (compared to purely aqueous test solution of I injected) if the test solution contained 60% (v/v) of methanol. Optimal order, concentrations and volumes of aspirated zones of reactants were: 61l of 80mM Na hexametaphosphate, 60 l of I in 60% (v/v) methanol, 40 l of 30mM H2SO4 and 9 l of 0.5mM KMnO4. Calibration curve relating the intensity of CL (peak height) to the concentration of I was linear in the range 1 - 8 M I; the limit of detection (S/N = 3) was 0.2 M I. The sample throughput was 100 h-1 . The repeatability of the peak heights was characterised by RSD 1.8% for 15 replicate injections of 2 M I. The SIA-CL method was used for the assay of I...
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Determination of fluoride in toothpaste using SIA system
Barvínková, Zuzana ; Sklenářová, Hana (advisor) ; Šatínský, Dalibor (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Mgr. Zuzana Barvínková Consultant: Doc. PharmDr. Hana Sklenářová, Ph.D. Title of Thesis: Determination of fluoride in toothpaste using SIA system Determination of fluorides was tested in the toothpastes using the SIA system. For determination of fluorides reaction of aluminium ions with aluminon that forms a red complex was used. After addition of a fluoride solution, aluminium ions were released from this complex. Spectrophotometry was used for the detection. The calibration range for sodium fluoride was linear over the range of 3x10-3 to 1x10-2 mol.l-1 . The repeatabitily of the measurement was tested for blank solution (water) and for the solutions of sodium fluoride 5x10-3 mol.l-1 and 6.25x10-3 mol.l-1 . The repeatability was in the range of 1.19 - 3.70 %, that was found acceptable. Then, potencially interfering ions were identified, which may effect determination of fluorides. Following the composition of usual toothpastes interferents could be: Cl- , Br- , I- , Ca2+ , PO4 3- , Zn2+ , CO3 2- , Al3+ , CH3COO- , Sn2+ , tartrate anion, citrate anion, lauryl sulphate anion, sorbitol and methylparaben. Among interfering ions, which influence the determination in low concentration, belong: Ca2+ ,...
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Development and applications of affinity carrier for isolation of human carbonyl-reducing enzymes
Andrýs, Rudolf ; Wsól, Vladimír (advisor) ; Šebela, Marek (referee) ; Šatínský, Dalibor (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Mgr. Rudolf Andrýs Supervisor: Prof. Ing. Vladimír Wsól, Ph.D Title of dissertation thesis: DEVELOPMENT AND APPLICATION OF AFFINITY CARRIER FOR ISOLATION OF HUMAN CARBONYL-REDUCING ENZYMES For several millennia the human medicine is based on application of small bioactive molecules that are administered in the form of plant extracts or synthetic compounds. However, their use in modern medicine is not possible without a detailed understanding of their biochemical effects and identification of their molecular targets. Chemical proteomics based on the specific recognition between the bioactive molecule and the target molecule is currently the most widely used techniques for identification of molecular targets of small molecules. Compared to conventional biochemical methods (e.g. 2D electrophoresis), chemical proteomics represents particularly sensitive and very selective technique that enable successful identification of biomolecules from complex biological samples that are naturally presented in very small concentrations. Carbonyl- reducing enzymes, which play an important role in physiology due to their involvement in metabolism of various endogenous (e.g. prostaglandins, steroid...
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Development and validation of HPLC method for determination of retinyl esters in human serum using monolithic columns.
Krčmová, Lenka ; Solich, Petr (advisor) ; Šatínský, Dalibor (referee)
DEVELOPMENT AND VALIDATION OF HPLC METHOD FOR DETERMINATION OF RETINYL ESTERS IN HUMAN SERUM USING MONOLITHIC COLUMN Absorption test of vitamin A is used for monitoring of intestinal permeability and may represent a sensitive indicator of intestinal damage. In this study, a simple and rapid reversed- phase high-performance liquid chromatography (RP-HPLC) procedure for selective and sensitive determination of retinol, -tocopherol, retinyl-palmitate and retinyl-stearate in blood serum has been developed and used for the monitoring of intestinal mucosal damage in cancer patients treated with cytotoxic drugs after absorption test of vitamin A. The HPLC instrumentation Series 200 LC from Perkin Elmer (Norwalk, USA) with diode- array detector (DAD) was used for the analysis. Separation of retinol, -tocopherol, retinyl- palmitate and retinyl-stearate were performed using the monolithic column Chromolith Performance RP-18e, 100 x 4.6 mm (Merck, Darmstadt, Germany). The gradient elution was used at the flow rate 3 ml min-1 ; mobile phase methanol:water (95:5) in 0-2.1 min and methanol:2-propanol (60:40) in 2.1-4.9 min. The total time of analysis was 6 min. The injection volume of sample was 20 μl and analysis was done at ambient temperature. The detection of retinol, -tocopherol and retinyl esters was carried out...
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