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Effect of prenylated flavonoids on biotransformation enzymes in vitro
Pataráková, Paula ; Szotáková, Barbora (advisor) ; Boušová, Iva (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Paula Pataráková Supervisor: Prof. Ing. Barbora Szotáková, Ph.D. Title of diploma thesis: Effect of prenylated flavonoids on biotransformation enzymes in vitro Prenylated flavonoids xanthohumol (XH), isoxanthohumol (IXH), 6-prenylnaringenin (6PN) and 8-prenylnaringenin (8PN) are flavanones and chalcones occurring in hops and their characteristic feature is prenyl chain on A ring. As these substances, present in beer and dietary supplements, are foreign compounds for organism, they can affect drug- metabolizing enzymes, including carbonyl reducing enzymes. The aim of our study was to find out, whether XH, IXH, 6PN, and 8PN have any impact on cytosolic carbonyl reductase 1 (CBR1) and aldo-keto reductase 1C subfamily (AKR1C). Viability test revealed that prenylated flavonoids in lower concentrations do not affect or even increase the viability of primary rat hepatocytes, but higher concentrations are toxic. AKR1C activity was increased in 8PN treated hepatocytes, the activity of CBR1 was significantly increased in IXH and 8PN treated samples. An increase in expression of AKR1C3 in IXH and 6PN treated samples, and decrease in XH and 8PN samples was observed. Expression of CBR1 was at the detection...
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Modification of selected biotransformation enzymes during ageing (in rat)
Vyskočilová, Erika ; Boušová, Iva (advisor) ; Svobodová, Hana (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Erika Vyskočilová Supervisor: PharmDr. Iva Boušová, Ph.D. Title of diploma thesis: MODIFICATION OF SELECTED BIOTRANSFORMATION ENZYMES DURING AGEING (IN RAT) Ageing is an inevitable biological process characterized by progressive functional decline of all organ systems with simultaneous increase in oxidative damage as a result of reactive oxygen species (ROS) accumulation. Increased activity of antioxidant enzymes, such as peroxidase and catalase, is related to increased production of ROS during ageing. Activity of biotransformation enzymes such as cytochrome P450 (CYP) and glutathion-S-transferase (GST) changes during ageing too. In general, activity of enzymes is lower in very young and very old individuals than in the period of adulthood. The aim of the present work was to study age-related changes in activities and expression of selected xenobiotic-metabolizing enzymes (GST, CYP 1A, CYP 2B, CYP 3A4) and antioxidant enzymes (catalase, peroxidase, superoxiddismutase) in rat liver. Male Wistar rats were sacrificed by decapitation under ether anesthesia at the age of 6 weeks and 21 months (6 individuals in each group). Subsequently, liver specimens were processed to obtain the subcellular...
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Formation of carbonyl compounds by non-enzymatic protein glycation
Krautová, Lenka ; Boušová, Iva (advisor) ; Dršata, Jaroslav (referee)
Author: Lenka Krautová Publishing place: Charles University in Prague, Faculty of Pharmacy in Hradec Králové Title: Formation of carbonyl compounds by non-enzymatic protein glycation Supervisor: PharmDr. Iva Boušová, Ph.D. This diploma thesis was aimed at the optimization of carbonyl groups determination in biological material. Carbonyl groups are formed by effect of protein oxidation or glycation. As a model protein, I used bovine serum albumin for spectrophotometric and aspartate aminotransferase for electrophoretic method of assessment, which were glycated by several glycating agents (glucose; glucose + copper; fructose; fructose + copper; methylglyoxal). I optimized different parameters of spectrophotometric determination: solvent, concentration of reagent, amount of protein, and duration of incubation. I managed to find suitable conditions of this determination, which I have used in subsequent experiments with different glycating agents. At electrophoretic determination, I performed SDS-PAGE with following silver staining or immunoblotting with chemiluminescent detection. The results of spectrophotometric determination showed that methylglyoxal has the highest glycating ability, followed by fructose with addition of cupric ions, fructose itself, glucose with addition of cupric ions, and the...
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Changes of lipid spectrum during body mass reduction in patients with diabetes mellitus
Šmídová, Barbora ; Dršata, Jaroslav (advisor) ; Boušová, Iva (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Barbora Šmídová Supervisor: Prof. MUDr. Jaroslav Dršata, CSc. Supervisor - specialist: RNDr. Mgr. Alena Tichá, PhD. Title of diploma thesis: CHANGES OF LIPID SPECTRUM DURING BODY MASS REDUCTION IN PATIENTS WITH DIABETES MELLITUS The thesis deals with the determination of lipid parameters (plasma total fatty acids, plasma total cholesterol, lipoprotein cholesterol, plasma triglycerides, plasma cholesterol precursors (lathosterol and squalene) and markers of cholesterol absorption (β-sitosterol and campesterol)) in obese patiens with diabetes mellitus type 1 and type 2 who were tested with a seven-day fasting, followed by low-calorie diabetic diet. It is assumed that weight loss should improve insulin resistence. The aim of this work is to evaluate the lipid parameters during the body weight reduction in obese patients suffering with diabetes mellitus. Lipids were determined before and after a seven-day fasting and after one month from the beginning of fasting (in patiens with diabetes mellitus type 1 also after one year). Gas chromatography was used for the determination of fatty acids and non-cholesterol sterols and squalene. Cholesterol and triglycerides were determined by a routine...
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OCT activity and glycation damage in rat liver mitochondria
Merhautová, Lucie ; Dršata, Jaroslav (advisor) ; Boušová, Iva (referee)
OCT ACTIVITY AND GLYCATION DAMAGE IN RAT LIVER MITOCHONDRIA Lucie Merhutová Diploma thesis. Charles University in Prague, Faculty of Pharmacy in Hradec Králové. 2007 This work was created following investigations carried out in LBBCV (Laboratoire de Biologie et Biochimie Cellulaire du Vieillissement) at University Paris Diderot - Paris 7. The team of this laboratory studies the mechanisms of posttranslational non-enzymatic modifications of proteins, involved in the pathophysiology of aging. Glycation is one of the mechanisms responsible for the modification of intracellular macromolecules leading to the loss of their structure and function. In previous investigations, ornithine carbamoyltransferase (OCT), the second enzyme of urea cycle, has been identified as one of the markedly glycated proteins in liver mitochondria of senescent rats. The goal of this work was to investigate in vitro and ex vivo glycation of OCT and the effect of glycation on its function. Purified OCT and mitochondrial matrix extracts were incubated with methylglyoxal, one of the most reactive physiological glycating agents, and submitted to the enzymatic assay and immunochemical analysis. We demonstrated that methylglyoxal modifies OCT both in vitro and ex vivo and causes rapid and extensive decrease in its enzymatic activity....
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In vitro effects of 3-hydroxytyrosol on renal hypoxia and inflammation
Kamasová, Terézia ; Boušová, Iva (advisor) ; Macháček, Miloslav (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Terézia Kamasová Supervisor: Asst. Prof. PharmDr. Iva Boušová, Ph.D. Prof. Alice Santos-Silva Maria João Valente, Ph.D. Title of diploma thesis: In vitro effects of 3-hydroxytyrosol on renal hypoxia and inflammation Chronic kidney disease (CKD) results from a group of heterogeneous disorders affecting the kidneys. The renal hypoxia and hypoxia-derived oxidative stress, renal fibrosis, and inflammation are highly prevailing conditions appearing in the diseased kidney, contributing to the progression of CKD. Phytochemicals are an essential part of contemporary therapeutic strategies for the treatment of various diseases. 3-Hydroxytyrosol (HT), a phenolic compound extracted from olives and olive-derived products (e.g. olive oil), is believed to carry a potent antioxidant, anti-inflammatory, antithrombotic, bactericidal and bacteriostatic activity. The aim of this work was to determine the preventive effect of HT in hypoxic renal cells and evaluate the effect of HT on hypoxia-related inflammation, fibrosis, and oxidative stress, in order to summarize the value of this phenolic compound as a promising novel remedy in the treatment of CKD. A cell line of human renal proximal tubular cells (HK-2) was...
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Modulatory effects of apigenin and luteolin on cytochrome P4501A expression and activity in intestinal cells
Šalovská, Barbora ; Skálová, Lenka (advisor) ; Boušová, Iva (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Title, Name, Surname of candidate: Barbora Šalovská Title, Name, Surname of tutor: Doc. RNDr. Lenka Skálová, Ph.D. Title of diploma thesis: Modulation effects of apigenin and luteolin on activity and expression of cytochrome P4501A2 in intestinal cells Flavonoids are a large group of polyphenolic compounds that occur naturally in plant kingdom. They have been shown to posses a variety of biological activities including anti-inflammatory, anti-oxidant and anti-mutagenic effects and play a prominent role in cancer prevention. The aim of this work was research of the effect of flavones apigenin and luteolin on an activity and expression of subfamily 1A of cytochromes P450 in human colon cancer cell line HCT-8. The CYP1A isozymes enzymatic activity was measured by EROD/MROD activity assay; the quantity of CYP1A's protein was set by Western blotting. We found out that apigenin and luteolin significantly decreased CYP1A activity in HCT-8 cells. During synchronous application of model inductor CYP1A β-naftoflavon (β-NF) with apigenin or luteolin (at the concentrations of 5µM and higher), the induction effect of β-NF on CYP1A was reduced. Apigenin and luteolin significantly decreased the CYP1A activity...
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Influence of hydroxycinnamic acids on methylglyoxal-induced protein glycation
Vejvalková, Hana ; Boušová, Iva (advisor) ; Trnková, Lucie (referee)
Author: Hana Vejvalková Publishing place: Charles University in Prague, Faculty of Pharmacy in Hradec Králové Title: Influence of hydroxycinnamic acids on methylglyoxal-induced protein glycation Supervisor: PharmDr. Iva Boušová, Ph.D. The diploma thesis was aimed at potencial antiglycation effect of antioxidants of natural origin (hydroxycinnamic acids) on the modification of bovine serum albumine and asparate aminotransferase by methylglyoxal as a glycating agent. I monitored the effect of hydroxycinnamic acids on generation of glycation products using fluorescent, absorption, and electrophoretic methods. The influence of hydroxycinnamic acids on the glycation of model proteins was ambiguous. The only derivative with proven antiglycation effect on production of non-specific glycation products and pentosidin measured by fluorescence and spectrophotometry was p-coumaric acid in 100 μM concentration. On the other hand, chlorogenic acid 100 μM displayed strong proglycation effects in all methods used for detection of glycation products. The effect of all tested substances was concentration dependent. Results show that the more substituents given acid contained in its molecule, the stronger proglycation effect this compound exerted.
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