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Analytical and bioanalytical evaluation of drug candidates from the group of aroylhydrazones
Kresová, Jana ; Kovaříková, Petra (advisor) ; Stariat, Ján (referee)
High-performance liquid chromatography (HPLC) is a progressive analytical method used for qualitative and quantitative drug analysis. This work describes various modifications of the chromatographic method for determining of BSIH (isonicotinic acid [2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)- benzylidene]-hydrazide) and SIH (salicylaldehyde isonicotinoyl hydrazone) in rabbit plasma. BSIH belongs to prochelators which convert to iron chelator SIH in the presence of hydrogen peroxide. Increased stability in plasma and lower toxicity even during repetitive administration is the advantage of BSIH. Separation of BSIH, SIH and internal standard (o-108) was tested on various stationary phases using different chromatographic conditions. However, none of them provided better results than the initial one. Tha analysis was performed on chromatographic column Zorbax Bonus-RP (150 mm x 3,0 mm, particle size 3,5 µm,) with guard column using identical sorbent at a flow rate of 0,3 ml/min. Phosphate buffer (10 mM monosodium phosphate, pH6, adjusted with sodium hydroxide) : acetonitrile with methanol (60:40) (v/v) with the ratio of 60:40 (v/v)) as the mobile phase. Detector response was registered at 297 nm. Linearity of the method was verified in the concentration range of 10 µM to 100 µM for both BSIH and...
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Metabolomic analysis of urine samples using ultra performance liquid chromatography and high resolution mass spectrometry
Mrvová, Andrea ; Mokrý, Milan (advisor) ; Stariat, Ján (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Drug Control Candidate: Andrea Mrvová Consultants: Prof. Seppo Auriola; Department of Pharmaceutical Chemistry, Faculty of Health Sciences, Kuopio, University of Eastern Finland RNDr. Milan Mokrý, CSc. Department of Pharmaceutical Chemistry and Drug Control, Faculty of Pharmacy in Hradec Králové, Charles University in Prague Title of Thesis: Metabolomic analysis of urine samples using ultra performance liquid chromatography and high resolution mass spectrometry Metabolomics is inquiring into the analysis of small-molecule metabolites within biologic system. This branch promises great progress not only as a diagnostic tool for disease. As suitable objects for the analysis are bio-fluids (e.g. urine, serum, saliva) and as common analytical techniques that are employed for the metabolomic analysis are NMR, GC/MS and LC/MS. Metabolomic analysis provides a great number of data and it is a problematic task to go through it. There is an effort to design system of effective identification. The aim of this work is to find optimal conditions for metabolomic analysis and to test reliability of computer-assisted workflow for the detection and identification of compounds. This should be as the first...
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HPLC analysis of drug candidates from the group of aroylhydrazones II.
Stariat, Ján ; Kovaříková, Petra (advisor) ; Kučera, Radim (referee)
1. ABSTRACT High performance liquid chromatography (HPLC) is one of the most frequently used analytical techniques for the analysis of drugs. Although iron is a vital element, excessive amounts in the body are highly toxic. The search for highly selective and effective iron chelating agents has been mainly inspired by the need to mobilize iron from tissues that are chronically overloaded with iron. However, recent investigations focused on the possibility to use iron chelators for the treatment of many other pathologies. Salicylaldehyde isonicotinoyl hydrazone (SIH), a biocompatible iron chelator derived from aroylhydrazone, is under extensive investigation as a promising drug candidate. Besides ability to bind iron, it shows interesting pharmacological effects: antioxidative, antiproliferative, cardioprotective, antimalarical and antimicrobic. The aim of this study was to develop optimal HPLC conditions for the separation of SIH and its potential metabolites (isoniazide, acetylisoniazide, salicylaldehyde) and to apply the method to the study focused on the isolation of analytes from rabbit urine using SPE. The best chromatographic analysis was achieved on a HPLC column (Phenomenex 250 4.6 mm I. D.) packed with Prodigy 5u ODS3 100A (5 μm) as a stationary phase. The mobile phase was composed of methanol :...
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Development of a bioanalytical method for the determination of novel drug candidates derived from thiosemicarbazone in a biological material
Suprunová, Vlasta ; Stariat, Ján (advisor) ; Kučera, Radim (referee)
Nowadays, high-performance liquid chromatography is widely used analytical method for separation and quantification of drugs in biological material due to its high sensitivity and selectivity. HPLC is suitable for quantitative as well as qualitative analysis. 2-benzoylpyridine-4-ethyl-3-thiosemicarbazone (Bp4eT) is a potential drug from the group of thiosemicarbazones, which are currently intensively studied and developed as anticancer agents. Their specific mechanism of action, based on chelation of iron, might overcome resistance to standard chemotherapy. The aim of this study was to develop and assess essential validation parameters of an HPLC-MS/MS method for determination of Bp4eT and its main phase I metabolites in rat plasma and to utilise the method for the analysis of samples from a pilot pharmacokinetic study in rats. The separation of Bp4eT and its metabolites was achieved on chromatographic column Discovery HSC18 (75 x 4.6 mm, 3 µm) protected by the same type of guard column using the mobile phase consisting of ammonium formate and acetonitrile in ratio 40:60 (v/v). The isocratic elution and the flow rate of 0.4 ml/min were utilised. Mass spectrometer was chosen as a detector and the quantification was performed in selected reaction monitoring mode. Plasma samples were treated using the...
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Analytical and bioanalytical evaluation of drug candidates from the group of aroylhydrazones
Kresová, Jana ; Kovaříková, Petra (advisor) ; Stariat, Ján (referee)
High-performance liquid chromatography (HPLC) is a progressive analytical method used for qualitative and quantitative drug analysis. This work describes various modifications of the chromatographic method for determining of BSIH (isonicotinic acid [2-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)- benzylidene]-hydrazide) and SIH (salicylaldehyde isonicotinoyl hydrazone) in rabbit plasma. BSIH belongs to prochelators which convert to iron chelator SIH in the presence of hydrogen peroxide. Increased stability in plasma and lower toxicity even during repetitive administration is the advantage of BSIH. Separation of BSIH, SIH and internal standard (o-108) was tested on various stationary phases using different chromatographic conditions. However, none of them provided better results than the initial one. Tha analysis was performed on chromatographic column Zorbax Bonus-RP (150 mm x 3,0 mm, particle size 3,5 µm,) with guard column using identical sorbent at a flow rate of 0,3 ml/min. Phosphate buffer (10 mM monosodium phosphate, pH6, adjusted with sodium hydroxide) : acetonitrile with methanol (60:40) (v/v) with the ratio of 60:40 (v/v)) as the mobile phase. Detector response was registered at 297 nm. Linearity of the method was verified in the concentration range of 10 µM to 100 µM for both BSIH and...
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Development of a bioanalytical method for the determination of novel drug candidates derived from thiosemicarbazone in a biological material
Suprunová, Vlasta ; Stariat, Ján (advisor) ; Kučera, Radim (referee)
Nowadays, high-performance liquid chromatography is widely used analytical method for separation and quantification of drugs in biological material due to its high sensitivity and selectivity. HPLC is suitable for quantitative as well as qualitative analysis. 2-benzoylpyridine-4-ethyl-3-thiosemicarbazone (Bp4eT) is a potential drug from the group of thiosemicarbazones, which are currently intensively studied and developed as anticancer agents. Their specific mechanism of action, based on chelation of iron, might overcome resistance to standard chemotherapy. The aim of this study was to develop and assess essential validation parameters of an HPLC-MS/MS method for determination of Bp4eT and its main phase I metabolites in rat plasma and to utilise the method for the analysis of samples from a pilot pharmacokinetic study in rats. The separation of Bp4eT and its metabolites was achieved on chromatographic column Discovery HSC18 (75 x 4.6 mm, 3 µm) protected by the same type of guard column using the mobile phase consisting of ammonium formate and acetonitrile in ratio 40:60 (v/v). The isocratic elution and the flow rate of 0.4 ml/min were utilised. Mass spectrometer was chosen as a detector and the quantification was performed in selected reaction monitoring mode. Plasma samples were treated using the...
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Analysis of selected drugs by liquid chromatography I.
Turňová, Ivana ; Pilařová, Pavla (advisor) ; Stariat, Ján (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Pharmaceutical Analysis Candidate: Ivana Turňová Supervisor: PharmDr. Pavla Pilařová, Ph.D. Title of Bachelor Thesis: Analysis of selected drugs by liquid chromatography I. High-temperature liquid chromatography (HTLC) is one of the possibilities of development and further progress in the field of liquid chromatography that is ranked among the most used separation methods also in the pharmaceutical analysis. Also, HTLC belongs in the so-called "green chemistry", which highlights performing the chemical practice in the most environment- friendly way. This thesis presents the HTLC issues in the form of literature search focused on the characterization of the fundamental features of HTLC and essential effect of the elevated temperature on the chromatographic processes given. It describes the most important advantages but also the limits of the HTLC method, mainly in its possible use in routine analyses. Finally, the thesis gives the overview of the selected results of several experimental studies, which have dealt with the use of HTLC in the analysis of some concrete drugs. Key words high-temperature liquid chromatography, drug analysis, green chemistry
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Development of a bioanalytical method for the determination of novel drug candidates derived from thiosemicarbazone in a biological material
Suprunová, Vlasta ; Stariat, Ján (advisor) ; Kučera, Radim (referee)
Nowadays, high-performance liquid chromatography is widely used analytical method for separation and quantification of drugs in biological material due to its high sensitivity and selectivity. HPLC is suitable for quantitative as well as qualitative analysis. 2-benzoylpyridine-4-ethyl-3-thiosemicarbazone (Bp4eT) is a potential drug from the group of thiosemicarbazones, which are currently intensively studied and developed as anticancer agents. Their specific mechanism of action, based on chelation of iron, might overcome resistance to standard chemotherapy. The aim of this study was to develop and assess essential validation parameters of an HPLC-MS/MS method for determination of Bp4eT and its main phase I metabolites in rat plasma and to utilise the method for the analysis of samples from a pilot pharmacokinetic study in rats. The separation of Bp4eT and its metabolites was achieved on chromatographic column Discovery HSC18 (75 x 4.6 mm, 3 µm) protected by the same type of guard column using the mobile phase consisting of ammonium formate and acetonitrile in ratio 40:60 (v/v). The isocratic elution and the flow rate of 0.4 ml/min were utilised. Mass spectrometer was chosen as a detector and the quantification was performed in selected reaction monitoring mode. Plasma samples were treated using the...
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Metabolomic analysis of urine samples using ultra performance liquid chromatography and high resolution mass spectrometry
Mrvová, Andrea ; Mokrý, Milan (advisor) ; Stariat, Ján (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Chemistry and Drug Control Candidate: Andrea Mrvová Consultants: Prof. Seppo Auriola; Department of Pharmaceutical Chemistry, Faculty of Health Sciences, Kuopio, University of Eastern Finland RNDr. Milan Mokrý, CSc. Department of Pharmaceutical Chemistry and Drug Control, Faculty of Pharmacy in Hradec Králové, Charles University in Prague Title of Thesis: Metabolomic analysis of urine samples using ultra performance liquid chromatography and high resolution mass spectrometry Metabolomics is inquiring into the analysis of small-molecule metabolites within biologic system. This branch promises great progress not only as a diagnostic tool for disease. As suitable objects for the analysis are bio-fluids (e.g. urine, serum, saliva) and as common analytical techniques that are employed for the metabolomic analysis are NMR, GC/MS and LC/MS. Metabolomic analysis provides a great number of data and it is a problematic task to go through it. There is an effort to design system of effective identification. The aim of this work is to find optimal conditions for metabolomic analysis and to test reliability of computer-assisted workflow for the detection and identification of compounds. This should be as the first...
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Analytical Evaluation of Drugs and Drug Candidates from the Group of Iron Chelating Agents
Stariat, Ján ; Klimeš, Jiří (advisor) ; Šatínský, Dalibor (referee) ; Šišpera, Luděk (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Department of Pharmaceutical Chemistry and Drug Control Candidate Mgr. Ján Stariat Supervisor Prof. RNDr. Jiří Klimeš, CSc. Title of Doctoral Thesis Analytical evaluation of drugs and drug candidates from the group of iron chelating agents High performance liquid chromatography (HPLC) ranks among the most important analytical techniques in the field of pharmaceutical analysis thanks to its robustness and the ability to separate and qualitatively and quantitatively analyse the compounds presented in various samples. The ongoing progress in the development of novel stationary phases and detection techniques allows rapid analysis and to get comprehensive characterization of the analytes presented in the complex samples even at very low concentrations. Cancer still remains a leading cause of death, thus the need for novel, efficient treatment strategies is of crucial importance. The mechanism based on iron chelation (Fe) inside the tumour cells represents one of the most promising strategy which could enhance the prognosis of patients suffering from cancer resistant to standard chemotherapy. Thiosemicarbazone iron chelators are currently under intensive development as novel anticancer drugs. Their strong antiproliferative...
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