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Sequential injection chromatography-testing and comparison of modern chromatographic columns
Molnárová, Petra ; Chocholouš, Petr (advisor) ; Sklenářová, Hana (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Deparment of Analytical chemismy Candidate: Bc. Petra Molnárová Supervisor: PharmDr. Petr Chocholouš, PhD. Title of diploma thesis: Sequential injection chromatogramy - testing and comparison of modern chromatographic columns This diploma thesis deals with testing and comparison of two modern chromatoghraphic columns in Sequential injection chromatography system. They were used for separation of selected vitamins B (thiamine, nikotinamide, pyridoxine, riboflavin and folic acid) and vitamin C. Developed method was based on UV detection at 220, 272 and 290nm. The Ascentis Express HILIC column (3 cm x 4.6 mm, 2.7 µm) was tested by using acetonitrile with ammonium acetate as a mobile phase. The method was not siccessfully developed because of low retention times of vitamins. The separation of six vitamins was not able, because of low selectivity. There was developed a method with use of Ascentis Express RP- Amide column (3 cm x 4.6 mm, 2.7 µm) using gradient elution, which was able to separate all 6 vitamins. Acetonitrile and phosphoric acid (pH 1.8) were components of the mobile phases. There was made calibration, reproducibility and recovery tests, but there was not good results enough to validate the method. For recovery was used...
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Sledování norfloxacinu, ciprofloxacinu a enrofloxacinu pomocí HPLC-FD ve vzorcích odpadních vod a jejich vliv na životní prostředí
Spurná, Marie ; Solich, Petr (advisor) ; Chocholouš, Petr (referee)
8 Abstract Fluoroquinolones (FQs) are used for therapy and prophylaxis of human and animal diseases and they are also used as growth promoters of animals. The usage of antibiotic is in connection with development of antibiotic resistance among bacterial populations. Since swine production industry is considered to be a relevant source of antibiotic resistant bacteria, measures to control the use of antimicrobial agents in animal husbandry have been adopted in recent years (1). The presence of FQs in wastewater from swine farms can introduce antibiotics in surface waters through agricultural runoff. FQs are rather resistant to microbial degradation and these compounds may be persisting within environmental waters because of their sorption properties, favouring the accumulation in sewage sludge, manure and soil (2). To trace the sources and ways of contamination, and evaluate the environmental impact of an outdoor swine production system, the occurrence norfloxacin (NOR), ciprofloxacin (CIP) and enrofloxacin (ENRO) residues were evaluated in wastewater and surface water samples. A LC-FD method based on the application of monolithic column (Chromolith Performance RP-18e (100 x 4.6 mm) successfully developed in previous studies (3,4,5) was applied. FQs were isocratically eluted using a mobile phase consisting...
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Study of temperature factor in gradient and isocratic HPLC separation
Habart, Miroslav ; Nováková, Lucie (advisor) ; Chocholouš, Petr (referee)
5 Abstract This thesis was focused on the determination of the temparature influence upon analysis of phenolic acids and fluoroquinolone antibiotics using high performance liquid chromatography. Analysis was performed on Zorbax SB C18 (50 x 4.6 mm; 1.8 µm) analytical column using ultra - viollet detection (UV) for both phenolic acids and fluoroquinolon antibiotics. The optimization of separation conditions was carried out for determination of phenolic acids and fluoroquinolone antibiotics. The effect of composition and concentration of aqueous part of mobile phase and especially the effect of temperature was investigated. The gradient elution was used for determination of phenolic acids. The mobile phase was formed by 0.1% formic acid and methanol. The isocratic elution was used for determination of fluoroquinolone antibiotics and the mobile phase was also formed by 0.1% formic acid and methanol. Accelerating of analysis and saving time and used solvents by using higher temperature were the main goals of this thesis. This premise came true in the analysis of phenolic acids. The analysis was faster with good peak resolution. The results for fluoroquinolone antibiotics were not better than basic 25žC measurement. Contrariwise, lower than laboratory temperature was more advantageous for their analysis. System...
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Dispersive liquid-phase microextraction of propofol using Dual valve SIA systém with fluorimetric detection
Rojková Kovačíková, Zuzana ; Boonjob, Warunya (advisor) ; Chocholouš, Petr (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Zuzana Rojková-Kovačíková Supervisor: Warunya Boonjob, Ph.D Consultant: Doc. PharmDr. Hana Sklenářová, Ph.D. Title of diploma thesis: Dispersive liquid-phase microextraction of propofol using Dual valve SIA system with fluorimetric detection This thesis deals with determination of propofol using automated dispersive liquid - liquid microextraction (DLLME) in SIA system with fluorimetric detection. Amyl acetate was selected as the extraction solvent; acetonitrile was used as the disperser solvent. Measurement parameters in SIA system were optimized. The calibration curve was linear in the range of propofol concentration 8 - 64 µg/ml with the correlation coefficient 0.9945. The limit of detection (LOD = 0.83 µg/ml) and the limit of quantification (LOQ = 7.25 µg/ml) were calculated. Repeatability for concentrations 16 and 64 µg/ml was proved; the values of the relative standard deviations (RSDs) were 1.54% and 1.52%. The analysis time was 152 s in case of one sample injection.
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Identification of phenolic compounds in plant extracts using UHPLC-MS/MS
Kmeťová, Ivana ; Nováková, Lucie (advisor) ; Chocholouš, Petr (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Analytical chemistry Candidate: Ivana Kmeťová Supervisor: Doc. PharmDr. Lucie Nováková, Ph.D. Title of Diploma thesis: Identification of phenolic compounds in plant extracts using UHPLC-MS/MS The aim of this Diploma thesis was identification of phenolic compounds in dry lyophilized water extract of leaves of wild strawberry (Fragaria vesca L.) using ultra-high performance liquid chromatography coupled with electrospray tandem mass spectrometry (UHPLC-ESI-MS/MS), advantageous in identification of complex plant samples. Phenolic compounds represent a group of very complex naturally occurring molecules in plants, possessing a wide variety of biological and physiological functions and acting as chemotaxonomic marker substances. This work was focused on flavonoids and related substances. For separation, UHPLC was used instead of more common HPLC due to its much higher separation efficiency that is crucial for successful separation of complex samples. Moreover, UHPLC enabled reduction of analysis time. Determination of structures of phenolics was based on evaluation and comparison of MS and MS/MS spectra in both negative and positive ESI ion modes. Full scan spectra, product ion spectra, neutral loss scans and reconstructed...
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HPLC determination of selected incestisides in cosmetic
Kameníčková, Daniela ; Šatínský, Dalibor (advisor) ; Chocholouš, Petr (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Daniela Kameníčková Supervisor: Doc. RNDr. Dalibor Šatínský, Ph.D. Title of Diploma Thesis: HPLC determination of selected incestisides in cosmetic Active ingredients fenoxycarb and permethrin were determined in cosmetic anti- parasitic product Arpalit® Neo shampoo against parasites with bamboo extract. Analysis was performed by HPLC using RP-Amide column 100 x 3 mm with a particle size of 2.7 micron and detection by UV at a wavelength of 225 nm. Separation proceeded by isocratic elution with mobile phase of acetonitrile - water (55:45) at a flow rate 1.0 ml/min. Temperature during measurement was 60 řC. For evaluation method was used an internal standard method, as the internal standard was used dye sudan II. For separation was needed time interval of 6 min. The retention time of fenoxycarb was 1.01, sudan II 2.87 and permethrin 4.42 min. Resolution of chromatographic peaks fenoxycarb and sudan II was 10.612 and sudan II and permethrin 6.524. Keywords: fenoxycarb, permethrin, HPLC.
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Application of the SIA system for aluminium determination
Fialová, Barbora ; Sklenářová, Hana (advisor) ; Chocholouš, Petr (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of analytical chemistry Candidate: Barbora Fialová Supervisor: PharmDr. Hana Sklenářová, PhD. Title of Diploma Thesis: Application of the SIA system for aluminium determination This diploma work deals with determination of aluminium ions in samples of surface water in a sequential injection system. For determination of aluminium ions method described in ČSN 83 0520 part 22 B was used. Determination was based on complex formation between aluminium ions and aluminon. The incurred complex is colored and color intensity is directly proportional to aluminium ions concentration in the sample. In this work spectrophotometric detection was used. Primarily, an optimization of reaction conditions in the test tube was performed. The calibration graph was linear in the range 0.02 - 1 mg/l Al with correlation coefficient R = 0.9932. Also detection limit (LOD = 0.0119 mg/l) and quantification limit (LOQ = 0.0390 mg/l) were assumed. Consequently reaction was converted into the SIA system and optimization of reaction conditions in the SIA system was performed. Calibration curve was linear in concentration range from 0.02 to 0.6 mg/l with R = 0.9971. Deformation of calibration curve in the field of higher aluminium concentrations is due...
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A study of HILIC mechanism on selected UHPLC stationary phases
Ježková, Kateřina ; Nováková, Lucie (advisor) ; Chocholouš, Petr (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Candidate: Kateřina Ježková Supervisor: Doc. PharmDr. Lucie Nováková, Ph.D. Title of Diploma thesis: A study of HILIC mechanism on selected UHPLC stationary phases This diploma thesis deals with a study of HILIC mechanism of Ascentis HILIC Express (2.1 x 100 mm, 2.7 µm, Sigma-Aldrich), ACQUITY UPLC BEH HILIC (2.1 x 100 mm, 1.7 µm, Waters) and ACQUITY UPLC BEH AMIDE (2.1 x 100 mm, 1.7 µm, Waters) stationary phases. Totally 15 analytes had been selected for testing of the characteristics of the stationary phases. The analytes were divided into groups of acidic, basic and neutral substances. The measurement was performed on UHPLC instrument by Waters with detection by a PDA detector at the wave-length of 254 nm. Isocratic elution was used for the measurement. The flow was set to 0.4 ml/min, injection volume was 1-2 µl. Mobile phase was composed of ultrapure water or buffer and acetonitrile. For an easier evaluation, tables and charts were constructed from measured data. Lin-lin, log-lin and log-log plots were constucted. The stationary phases were evaluated in terms of selectivity and retention of the individual groups of analytes. The influence of a percentage of water and acetonitrile in mobile...
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Verification of practical application of parallel SIA determination of inorganic anions in real samples of surface water
Grebeníček, Lukáš ; Sklenářová, Hana (advisor) ; Chocholouš, Petr (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of: Analytical chemistry Candidate: Bc. Lukáš Grebeníček Supervisor: PharmDr. Hana Sklenářová, Ph.D Title of Diploma Thesis: Verification of practical application of parallel SIA determination of inorganic anions in real samples of surface water This diploma thesis is divided into two parts. In the first one, the experimental determination deals with verification of practical application of chosen anions (SiO3 2- , Cl- , NO2 - , NO3 - ) by sequential injection (SIA) assay followed by valid ISO norms for flow injection (FIA) assays. The second part compares determination of nitrites and nitrates by the SIA assay and comparative assay from MERCK company. Determination of dissolved silicates was accomplished in accordance with the Czech technical norm ČSN EN ISO 16264. This technical standard for FIA assay is based on reaction of dissolved silicates with reagent solution. Reagent is composed of acidic solution of molybdenite, which reacts with silicates and phosphates generating molybdic-silicate acid and molybdic-phosphate acid. Molybdic-phosphate acid is decomposed by oxalic acid solution and molybdic-silicate acid is reduced by stannic chloride to molybdic blue - the final product of reaction - assayed...
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