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Analysis of historical pharmaceutical preparation containing chicory from the 18th century
Lener, Tomáš ; Nesměrák, Karel (advisor) ; Kubíčková, Anna (referee)
A historical pharmaceutical preparation - chicory extract - from the 18th century whose authenticity was investigated was qualitatively analyzed using HPLC with UV/VIS and MS2 detection. A current dry chicory root was used as a source of selected chicory chemotaxonomic markers and an indicator of the success of extraction with different solvents. The authenticity of the historical extract was proved by a water solubility test. A VIS spectrometric determination of inulin was used and the inuline content was determined to be 52.4% in the current dry chicory root and 7.45% in the historical extract. The detection wavelength 254 and 320 nm was selected from measured UV/VIS spectra of selected standards and the literature. The authenticity of the historical chicory extract was proven by tandem mass spectrometry, and, moreover, arteincultone - a new chicory chemotaxonomic marker - was identified. Key words: HPLC-MS, HPLC-UV/VIS, UV/VIS Spectrometry, Chicory, Historical extract, Chicory root, Arteincultone
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Analysis and isolation of intermediates involved in the biosynthetic pathway of alkylproline derivatives
Cudlman, Lukáš ; Bosáková, Zuzana (advisor) ; Michalíková, Klára (referee)
(EN) This work aims at preparation, analysis and isolation of intermediates of biosynthetic pathways of 4-alkyl-L-proline derivatives for their structural elucidation. Compounds with incorporated 4-alkyl-L-proline derivatives include clinically used lincosamide antibiotic, lincomycin A, antitumor pyrrolobenzodiazepines and bacterial hormone hormaomycin. Detailed knowledge of biosynthetic pathways of these biological active substances can be used to prepare new, more efficient derivatives. The first part of this work focuses on yellow-coloured dicarboxylic intermediates 1 and 2 of the biosynthetic pathway of 4-propyl-L-proline - the precursor of lincomycin A. In the presence of the methylation agent, S-adenosyl-L-methionine, and LmbW C- -methyltransferase, 1 was partially converted into intermediate 2. Using ultra-high performance liquid chromatography, both intermediates were identified from absorption and mass spectrometry spectra. A semi-preparative chromatographic method for isolation of both intermediates was developed. Surprisingly, a significantly lower stability of 2 compared to intermediate 1 was observed in an in vitro enzymatic reaction mixture. The second part of the work focuses on 4-ethylidene-L-proline - the precursor of tomaymycin belonging to pyrrolobenzodiazepines. After...
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Analysis of the degradation products of the active substances in historical pharmaceutical relicts from 18th and 20th century.
Čambal, Peter ; Nesměrák, Karel (advisor) ; Kubíčková, Anna (referee)
Historical pharmaceutical preparations analyzed in this thesis were a senna extract more than 200 years old, an ointment "Naso-Merfen" 75 years old, and an ointment "Sulfathiazol" 42 years old. The active substances in the analyzed samples were sennosides A and B (senna extract), ephedrine and menthol (the ointment "Naso-Merfen"), and sulfathiazole (the ointment "Sulfathiazol"). The senna extract was analyzed by RP-HPLC and HPLC-MS. Separation conditions were optimized, especially for separation of the sennoside A and B enantiomers. The active substances were not detected in the sample. One degradation product and substances characteristic for senna were identified. Their presence in the historical and contemporary sample was compared. Detailed ESI− -MSn fragmentation mechanisms of sennoside A and B have been proposed. The sample of the ointment "Naso-Merfen" was analyzed by HILIC-UV, HPLC-MS, GC-MS, and AAS. Separation conditions were optimized. The active substances were quantified. Degradation products of the active substances were not detected in the sample. The sample of the ointment "Sulfathiazol" was analyzed by RP-HPLC and HPLC-MS. Separation conditions were optimized. The active substance was quantified. Degradation products of the active substance were not detected. The authenticity of...
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Ergothioneine and mycothiol in the biosynthesis of lincosamides
Seidlová, Bára ; Kameník, Zdeněk (advisor) ; Kopecký, Jan (referee)
Specialized microbial metabolites are described as low-molecular-weight bioactive compounds, which are dispensable for the growth, evolution, or reproduction of its producer. This group of substances includes the lincosamides, which are produced mainly by the bacteria of the Streptomyces genera. Apart from other precursors, two low-molecular-weight thiols, ergothioneine and mycothiol, are essential participants of the lincosamide biosynthesis. Mycothiol (MSH) serves in this pathway as a source of sulphur, on the other hand, ergothioneine (ESH) constitutes a conjugate with the aminosugar moiety of lincosamide structure. The conjugate is condensed with an activated amino acid, which is catalyzed by an unusual enzyme to form a core of the lincosamide molecule. The objective of this diploma thesis is to isolate the conjugate of ESH and aminooctose, which serves as a substrate of the LmbD biosynthetic protein. Another aim is to study the links between the thiol metabolism and the biosynthesis of three lincosamides, lincomycin, celesticetin, and intervencin, which are produced by different bacterial strains. Bacterial strains were cultivated under laboratory conditions and methods of liquid chromatography with UV and MS detection were used for the analysis. The parameters of the methods were developed...
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Analysis and isolation of intermediates involved in the biosynthetic pathway of alkylproline derivatives
Cudlman, Lukáš ; Bosáková, Zuzana (advisor) ; Michalíková, Klára (referee)
(EN) This work aims at preparation, analysis and isolation of intermediates of biosynthetic pathways of 4-alkyl-L-proline derivatives for their structural elucidation. Compounds with incorporated 4-alkyl-L-proline derivatives include clinically used lincosamide antibiotic, lincomycin A, antitumor pyrrolobenzodiazepines and bacterial hormone hormaomycin. Detailed knowledge of biosynthetic pathways of these biological active substances can be used to prepare new, more efficient derivatives. The first part of this work focuses on yellow-coloured dicarboxylic intermediates 1 and 2 of the biosynthetic pathway of 4-propyl-L-proline - the precursor of lincomycin A. In the presence of the methylation agent, S-adenosyl-L-methionine, and LmbW C- -methyltransferase, 1 was partially converted into intermediate 2. Using ultra-high performance liquid chromatography, both intermediates were identified from absorption and mass spectrometry spectra. A semi-preparative chromatographic method for isolation of both intermediates was developed. Surprisingly, a significantly lower stability of 2 compared to intermediate 1 was observed in an in vitro enzymatic reaction mixture. The second part of the work focuses on 4-ethylidene-L-proline - the precursor of tomaymycin belonging to pyrrolobenzodiazepines. After...
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