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Study of retention and separation of peptides in HPLC
Boudová, Hana ; Kalíková, Květa (advisor) ; Kubíčková, Anna (referee)
This diploma thesis deals with the study of retention and separation of structurally different peptides and digest products of cytochrome c using reverse phase and mixed modes of liquid chromatography. As the model peptides, five unmodified dipeptides, nine dipeptides containing a protective benzoyl group on the N-terminal amino acid and eight therapeutic peptides containing five to ten amino acids were selected. The chromatographic behaviour of the model peptides was tested on three different columns: XBridge C18, XSelect CSH C18 and Atlantis Premier BEH C18 AX under different conditions. The influence of the pH of the aqueous part of the mobile phase, i.e. 2,1; 3,0; 4,7 and 6,8, and the amount of acetonitrile in the mobile phase on retention and separation of model peptides was tested. The effect of the concentration of ammonium acetate buffer, pH = 6,8 in the mobile phase on the retention and peak symmetry in the concentration range of 10-50 mmol L-1 was assessed. The chromatographic conditions for the baseline separation of 20 different model peptides were optimized. The conditions were as follows: XSelect CSH C18 column, mobile phase composed of acetonitrile/10mM ammonium acetate, pH = 6,8 (v/v) under gradient elution. The suitable chromatographic conditions for the analysis and identification...
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Multidimensional characterization of polyelectrolytes and interpolyelectrolyte complexes in aqueous solutions
Murmiliuk, Anastasiia ; Štěpánek, Miroslav (advisor) ; Šachl, Radek (referee) ; Hoffmann, Ingo (referee)
Multidimensional characterization of polyelectrolytes and interpolyelectrolyte complexes in aqueous solutions Abstract: This PhD thesis is dedicated to the study of polyelectrolytes and their self-assembly in aqueous solutions. The morphology and ionization state of individual polymer chains were investigated as well as their co-assembly with oppositely charged species. First, the charge regulation of short and long weak polyelectrolytes in aqueous solution was investigated to deeper understand their pH-responsive properties. The changes of ionization degree and charge of oligopeptides composed of 5 amino acids with acid side-chains and 5 with basic side chains were followed upon varying the pH in order to reveal the effect of interactions between the like-charged and oppositely charged groups. It was shown that intra-molecular electrostatic interactions and conformational flexibility caused the suppression of the total charge and enhancement of ionization of the peptides. To get an insight into the distribution of local H+ concentration in the proximity of a polyelectrolyte chain, a modified polyelectrolyte structure was used with a fluorescent pH- indicator, covalently attached at the end of the chain. Ionization of the pH-responsive fluorophore revealed its effective pK which was compared with the local...
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Multidimensional characterization of polyelectrolytes and interpolyelectrolyte complexes in aqueous solutions
Murmiliuk, Anastasiia ; Štěpánek, Miroslav (advisor) ; Šachl, Radek (referee) ; Hoffmann, Ingo (referee)
Multidimensional characterization of polyelectrolytes and interpolyelectrolyte complexes in aqueous solutions Abstract: This PhD thesis is dedicated to the study of polyelectrolytes and their self-assembly in aqueous solutions. The morphology and ionization state of individual polymer chains were investigated as well as their co-assembly with oppositely charged species. First, the charge regulation of short and long weak polyelectrolytes in aqueous solution was investigated to deeper understand their pH-responsive properties. The changes of ionization degree and charge of oligopeptides composed of 5 amino acids with acid side-chains and 5 with basic side chains were followed upon varying the pH in order to reveal the effect of interactions between the like-charged and oppositely charged groups. It was shown that intra-molecular electrostatic interactions and conformational flexibility caused the suppression of the total charge and enhancement of ionization of the peptides. To get an insight into the distribution of local H+ concentration in the proximity of a polyelectrolyte chain, a modified polyelectrolyte structure was used with a fluorescent pH- indicator, covalently attached at the end of the chain. Ionization of the pH-responsive fluorophore revealed its effective pK which was compared with the local...
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Design and characterization of advanced polymer-coated upconversion nanoparticles
Kostiv, Uliana ; Horák, Daniel (advisor) ; Kaman, Ondřej (referee) ; Brožek, Jiří (referee)
Lanthanide-based upconversion nanoparticles are able to absorb low-energy near-infrared (NIR) photons and emit visible or ultraviolet light. This unique optical property enables luminescent detection without autofluorescence from biological samples that is crucial for bioimaging and diagnostics. Moreover, NIR irradiation allows to deliver light deep into the tissue, which can be used for drug or gene delivery and NIR-induced photodynamic therapy (PDT) of tumors. Despite huge effort during the last 10 years devoted to the upconversion nanoparticle synthesis and their surface engineering, preparation of the particles suitable for biomedical applications still remains a big challenge. The current work is focused on synthesis of the upconversion nanoparticles with controllable morphology, size, distribution, crystallinity, and high upconversion efficiency, as well as on desirable surface modification by biocompatible inorganic or organic polymers. Neat and functionalized homogeneous silica shell was introduced on the NaYF4:Yb3+ /Er3+ and NaGdF4:Yb3+ /Er3+ nanoparticles to enhance their biocompatibility and colloidal stability in water. To render the nanoparticles with targeting moieties, their surface was decorated with cell-adhesive RGD or cell-penetrating TAT peptides. The RGD- and TAT- conjugated...
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Radiolabelled antibodies - the perspective for diagnosis and therapy
Mejtská, Jana ; Bárta, Pavel (advisor) ; Ramos Mandíková, Jana (referee)
1 CHARLES UNIVERSITY FAKULTY OF PHARMACY IN HRADEC KRÁLOVÉ DEPARTMENT OF BIOPHYSICS AND PHYSICAL CHEMISTRY DISSERTATION THESIS RADIOLABELED ANTIBODIES - THE PERSPECTIVE FOR DIAGNOSIS AND THERAPY Supervizor: Mgr. PAVEL BÁRTA, Ph.D. HRADEC KRÁLOVÉ, 2017 Bc. JANA MEJTSKÁ 2 ABSTRACT EN Different types of tissues have a characteristic cell morphology. Each cell has typical molecules on its surface, which may be either of physiological or pathological type. The presence of these surface structures can be interesting for possible modulation of specific cell populations from neighboring cells. Utilization of this property is then essential particularly in a case of tumor cells. Targeting on tumor specific cell structures involves the use of receptor specific peptides or monoclonal antibodies. The discovery of the preparation of monoclonal antibodies has opened a new chapter in the treatment and diagnosis not only tumor diseases. The advantage of monoclonal antibodies is their specificity and also high affinity to the type of the target cell structures. This study is focused on the summary of monoclonal antibodies which are currently being applied on the treatment or diagnosis of a particular cancer. Furthermore, this work also includes antibodies which are under development for intended medical applications with...
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