Ladislav Zeman

Deoxynivalenol and its toxicity.

Deoxynivalenol and its toxicity Deoxynivalenol (DON) is one of several mycotoxins produced by certain Fusarium species that frequently infect corn, wheat, oats, barley, rice, and other grains in the field or during storage. The exposure risk to human is directly through foods of plant origin (cereal grains) or indirectly through foods of animal origin (kidney, liver, milk, eggs). It has been detected in buckwheat, popcorn, sorgum, triticale, and other food products including flour, bread, breakfast cereals, noodles, infant foods, pancakes, malt and beer. DON affects animal and human health causing acute temporary nausea, vomiting, diarrhea, abdominal pain, headache, dizziness, and fever. This review briefly summarizes toxicities of this mycotoxin as well as effects on reproduction and their antagonistic and synergic actions.

Utilizing of Square Wave Voltammetry to Detect Flavonoids in the Presence of Human Urine

About biological affecting of flavonoids on animal organisms is known less, thus we selected flavonoids, flavanones and flavones, and their glycosides, which were examined as potential inducers of cytochrome(s) P450 when administrated by gavages into experimental male rats. The study was focused on induction of CYP1A1, the major cytochrome P450 involved in carcinogen activation. The data obtained demonstrate the necessity of taking into account not only ability of flavonoids to bind to Ah receptor (induction factor) but also to concentrate on their distribution and metabolism (including colon microflora) in the body. After that we examined certain flavonoids as potential inducers of cytochrome P450, we wanted to suggest and optimize suitable electrochemical technique for determination of selected flavonoids (quercetin, quercitrin, rutin, chrysin and diosmin) in body liquids. For these purposes, we selected square wave voltannetry using carbon paste electrode. Primarily we aimed on investigation of their basic electrochemical behaviour. After that we have optimized frequency, step potential and supporting electrolyte. Based on the results obtained, we selected the most suitable conditions for determination of the flavonoids as follows: frequency 180 Hz, step potential 1.95 mV/s and phosphate buffer of pH 7 as supporting electrolyte. Detection limits (3 S/N) of the flavonoids were from units to tens of nM except diosmin, where the limit were higher than μM. In addition, we attempted to suggest a sensor for analysis of flavonoids in urine. It clearly follows from the results obtained that flavonoids can be analysed in the presence of animal urine, because urine did not influence much the signals of flavonoids (recoveries of the signals were about 90 %).

Study of Interactions between Metallothionein and Cisplatin by using Differential Pulse Voltammetry Brdickas reaction and Quartz Crystal Microbalance

Treatment strategies for tumour diseases are progressively focusing on personalization of medicine. However, this focus requires methods revealing the early general biological mechanisms, including the formation anti-cancer drugs’ resistance. The low molecular mass protein metallothionein is thought to be the crucial for the formation of resistance in tumour treatment based on the platinum-cytostatics. The interactions between metallothionein (MT) and cisplatin were determined by the adsorptive transfer stripping technique coupled with the differential pulse votlammetry Brdickás reaction. The signals related to the MT-cisplatin complex appeared at −0.9 V. The formation of this complex depended on the time of interaction between cisplatin and MT. The complex formation was consequently confirmed by quartz crystal microbalance analyses. The formation of this complex was detectable even after a 20 s long interaction. Moreover, we detected presence of MT-cisplatin complex in the blood of male rats treated with this drug.

Changes in metallothionein level in rat hepatic tissue after administration of natural mouldy wheat.

Mycotoxins are secondary metabolites produced by microfungi that are capable of causing disease and death in humans and other animals. This work was aimed at investigation of influence of mouldy wheat contaminated by pathogenic fungi producing mycotoxins on metallothionein levels in hepatic tissue of rats. The rats were administrating feed mixtures with different contents of vitamins or naturally mouldy wheat for 28 days. It was found that the wheat contained deoxynivalenol (80 ± 5 μg per kg of mouldy wheat), zearalenone (56 ± 3 μg/kg), T2-toxin (20 ± 2 μg/kg) and aflatoxins as a sum of B1, B2, G1 and G2 (3.9 ± 0.2 μg/kg). Rats were fed diets containing 0, 33, 66 and 100% naturally moulded wheat. Control group 0, 33, 66 and 100% contained vitamins according to Nutrient Requirements of Rats (NRC). Other four groups (control group with vitamins, vit33, vit66 and vit100%) were fed on the same levels of mouldy wheat, also vitamins at levels 100% higher than the previous mixtures. We determined weight, feed conversion and performed dissection to observe pathological processes. Changes between control group and experimental groups exposed to influence of mouldy wheat and experimental groups supplemented by higher concentration of vitamins and mouldy wheat were not observed. Livers were sampled and did not demonstrate significant changes in morphology compared to control either. In the following experiments the levels of metallothionein as a marker of oxidative stress was determined. We observed a quite surprising trend in metallothionein levels in animals supplemented with increased concentration of vitamins. Its level enhanced with increasing content of mouldy wheat. It was possible to determine a statistically significant decline (p<0.05) between control group and groups of animals fed with 33, 66 and 100% mouldy wheat. It is likely that some mycotoxins presented in mouldy wheat are able to block the mechanism of metallothionein synthesis.

Whole tract and post ruminal digestibility determined by in situ ruminal, intestinal mobile nylon bag and whole tract nylon capsule methods

Whole tract and post ruminal digestibility of crude protein (CP) and dry matter (DM) in ryegrass, alfalfa hay (high and medium quality) and maize silage were determined by means of a rumen in situ nylon bag, a mobile nylon bag and a nylon capsule method as well as by calculations that combined the methods. Calculation of whole tract digestibility was from results of rumen in situ nylon bag and mobile nylon bag methods and calculation of post ruminal digestibility was from results of the nylon capsule and rumen in situ nylon bag methods. Three dry cows (Red Holstein × Black Friesian) fitted with ruminal and duodenal cannulas were used for testing the nylon bag and mobile bag methods, whereas the nylon capsule method utilised three dairy cows (Red Holstein × Black Friesian). Differences between determined and calculated values for whole tract digestibility of CP and DM ranged from −3.3 to +2.9%, and from −0.1 to −8.5%, respectively. Differences between determined and calculated values of intestinal digestibility of undegraded CP were higher and ranged from −10.6 to +13.8%, and from +0.4 to +17.5% for CP and DM, respectively.

Effect of selenium in organic and inorganic form on liver, kidney, brain and muscle of Wistar rats

AbstractSelenium is a micronutrient, localized in the active sites of enzymes such as glutathione peroxidase and thioredoxin reductase, and participating together with these enzymes in an antioxidant defence system of organisms against free radicals. Administration of selenium is necessary for maintaining oxidative homeostasis. The present experiment is aimed at investigation of selenium impact on basal metabolic processes and selected antioxidants in a Wistar rat model, fed selenium in organic and inorganic forms. Liver, kidney, brain and muscle were sampled during a month-long feeding with four different doses of selenium (0.075 mg or 1.5 mg of inorganic and/or organic selenium per kg of feed). We found a significant reduction in glutathione level in liver tissue regardless of the form of the administered selenium. On the other hand, selenium caused a decreased glutathione reductase level in the liver and metallothionein level in the liver, kidney and muscle.

Complexes of glutathione with heavy metal ions as a new biochemical marker of aquatic environment pollution

Reduced glutathione (GSH) plays a number of key roles in many biochemical pathways. This peptide is highly reactive and forms conjugates with other molecules via its sulfhydryl moiety. The interactions of the common heavy metal pollutant Cd(II) with GSH were determined by using the Brdicka reaction to evaluate whether this technique would be suitable as a biomarker. After GSH interaction with Cd(II) ions, two characteristic changes in the measured voltammogram were observed: Cat2 signal height decreased, and a new signal called P1 was found. The observed signal probably relates to the formation of a GSH-heavy metal ion complex adsorbed on the surface of the working electrode. When the interaction of GSH with cisplatin was studied, the same characteristic changes in the voltammogram were observed, which confirmed our hypothesis. Moreover, changes in the height of P1 and Cat2 signals with increasing time of GSH interaction with Cd(II) ions and/or cisplatin were also investigated. Cat2 peak height decreased proportionally with increasing time of interaction. This decrease can be explained by shielding of free sulfhydryl moiety by heavy metal ions, so it cannot catalyze the evolution of hydrogen from the supporting electrolyte. In addition, we found that, with increasing time of the interaction, the P1 signal was enhanced and shifted to more positive potentials for both Cd(II) ions and cisplatin.

Influence of garlic extract on antioxidant status of chicken.

In 2006 the European Union banned the feeding of antibiotic growth promoters because of possible risk of drug resistance in human pathogens bacteria. This is the reason for the study of various phytogenic additives and their extracts as a natural source of biologically important compounds. Antimicrobial substances are a commonly included in chicken feed rations. They are used mainly as prevention against various diseases, and also to stimulate growth. The beneficial effects of garlic on animal organism resulting from their antimicrobial, antioxidative and antihypertensive properities. Studies focused on growth, conversion and meat quality of different types of animals indicate its positive effects. In our experiment we studied the influence of garlic extract in a dose of 0, 10 g and 15 g per 1 kg of chicken feed mixture. We focused on weight gains and antioxidant status of an organism. The experiment took 39 days. 54 seven-day-old chickens were included in the experiment. The chickens were weighed once a week, when aged 11, 17, 24, 31 and 38 days, at the same time of the day. The chickens had ad libitum access to feed ration and water. The chickens were taken blood sample at the end of the experiment when 39 days old. Their antioxidant status were measured using ABTS, FRAP and DPPH methods. Our results show that owing to higher concentration of garlic extract in feed ration the antioxidant status of observed chickens was increased. DPPH method showed an increase in antioxidant status of both experimental groups by 38% (a group with a dose of 10 g/kg of mixture) and by 46% (a group with a dose of 15 g/kg of mixture) compared to the control group. When using FRAP method, antioxidant status of both G10 and G15 groups increased by 24%, resp. 16%. No evidential differences in antioxidant activity between the experimental groups and control group were found using ABTS method. The supplement of garlic extract into a feed ration did not have any influence on weight gains of chickens.

Effect of chamomile supplements to feeding doses on antimicrobial parameters in poultry.

Due to a ban of use of antibiotic growth promoters in the poultry industry it is necessary to look for alternative solutions. The use of some herbs showing antimicrobial effects can be one of such alternatives. In this experiment, effects of three different concentrations of chamomile (Matricaria chamomilla) extract, (0.3%; 0.6% and 1.2%) in feeding doses on the microbial population in the gastrointestinal tract of growing broiler chickens were studied. The main attention was paid to the population of Clostridium perfringens and to numbers of coliform microbes. Clostridia were cultivated under anaerobic conditions at 46 °C on the Tryptone Sulfite Neomycin (TSN) agar for a period of 24 hours. Coliform microbes were grown on the violet red bile lactose (VRBL) agar at 37 °C for a period of 24 hours. The experiment lasted 39 days and involved 80 chicks that were slaughtered in the course of their growth period at the age of 18, 25, 32 and 39 days; there were 5 chicks in each group. The obtained results indicated that increasing doses of chamomile in the feeding ration decreased numbers of coliform microbes in the digestive tract of chicks and also reduced the population of C. perfingens.

Study of deoxynivalenol effect on metallothionein and glutathione levels, antioxidant capacity, and glutathione-S-transferase and liver enzymes activity in rats

Deoxynivalenol (DON, vomitoxin) is one of several human mycotoxins acquired through residues in foods of plant origin (cereals) and through foods of animal origin (kidneys, liver, milk, eggs). The aim of this study was to investigate the impact of deoxynivalenol-contaminated mouldy grain on the health status of rats and their ability to resist oxidative stress. The rats were fed a diet supplemented with DON along with organic and inorganic Zn(II), and vitamins for 28 days. The study focuses particularly on protective mechanisms and levels of reduced glutathione and metallothioneins (MT) against the adverse effects of xenobiotics. The highest concentrations of MT were detected in the tissues of kidneys ((6.70 ± 0.05) μg g−1) and liver ((6.00 ± 0.05) μg g−1), both of which are involved in detoxifying cationic xenobiotics. Lower concentrations were also detected in heart, brain, testes, and muscle tissues (approximately 3.0 μg g−1). Antioxidant capacity, glutathione-S-transferase, and liver enzymes activity were also studied. In conclusion, MT may play an important role in the detoxification of mycotoxins. Even though its role is not fully understood, MT are crucial for the redistribution of ions important for transcription factors and interaction with reactive oxygen species that form mycotoxins.