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The employment of wastes from food production
Hurčíková, Andrea ; Babák, Libor (referee) ; Omelková, Jiřina (advisor)
The waste from agricultural and food industry are accessible in large quantity anywhere in the whole world nowadays. Most of these wastes include cellulose (30 - 40 %), hemicellulose (20 - 40 %) and lignine (10 - 20 %). Therefore these waste materials have wide use as the substrates for the microbial growth and the production of the enzymes. The microorganisms are able to use organic compounds from the wastes as the source of energy for the growth and carbon for synthesis of cellular biomass [24]. Wheat and rice straw are possible to use as the substrates for cultivation of the microorganisms and following production of the enzymes. In this thesis the utilization of the wastes from food industry for the production of the enzymes by the microorganisms was studied. We observed utilization of wheat straw as source of energy for growth of tested microorganisms and investigated their ability for the production of oxidoreductase (laccase). The optimalization of growth conditions of Aureobasidium pullulans was proceeded. Further the activity of laccase was studied. Milled wheat straw was used as the substrate. The cultivation was done in the thermoregulator at the temperature of 27°C. The activity of laccase was not found in this thesis. Petri dishes were contaminated by three unknown microoganisms during optimalization of growth of Aureobasidium pullulans. One of them produced laccase in cultivation with straw.
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Texturní a barevné vlastnosti sýrů ošetřených extracelulárními enzymy
Jašková, Jana
This diploma thesis deals with textural and color properties of cheeses treated with an extracellular enzyme. The cheese samples were matured for three weeks. The penetration and compression method was used to determine the textural properties of the cheeses. The color of the cheeses was measured on a CM 3500d spectrophotometer. The lowest strength determined by the penetration method had the sample with laccase after production (0.55 ± 0.07 N) and the highest strength had the control sample, which was aged for one week (1.31 ± 0.14 N). The lowest strength determined by the compression method had the sample with laccase, which had matured for two weeks (3.11 ± 1.07 N) and the highest strength had the control sample of cheese matured for three weeks (12.28 ± 2.36 N). The values of L*(D65) varied for all samples in the range from 72.89 ± 0.31 to 87.76 ± 0.01, the values of a*(D65) from -1.95 ± 0.01 to 0.66 ± 0.05 and b*(D65) values were measured between 6.73 ± 0.35 and 20.93 ± 0.03 Laccase enzyme treatment had no significant effect on the firmness and color of the cheese samples.
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Vliv receptury a délky zrání na obsah polyaminů ve zrajících sýrech
Kolková, Andrea
The theoretical part of my thesis was focused on the characteristics of polyamines, their synthesis, degradation and effect in the organism. The practical part was involved on determining the content of polyamines (putrescine, spermine and spermidine) in ripening cheese without addition and with the addition of laccase. The high-performance liquid chromatography method was used for the analysis. Quantitatively, the most significant polyamine in all tested cheese variants was spermine (laccase in cheese grains 16.6 mg/kg > control 15.5 mg/kg > surface laccase 14.8 mg/kg; p < 0.05). Spermidine content was in the range (surface laccase 8.8 mg/kg > laccase in cheese grains 2.9 mg/kg > control 2.4 mg/kg; p < 0.05). Putrescine was detected only in low precipitation (laccase in cheese grains 2.1 mg/kg > control 1.3 mg/kg > laccase in surface 0.7 mg/kg; p < 0.05). During storage (X; weeks), the total PA content (Y; mg/kg) in the control cheese sample without the addition of laccase increased according to the equations: y = 256.93 + 207.38x (R² = 0.8955; p = 0.0021 ), the amount of PA in the sample of cheese with laccase in the cheese grains also increased linearly according to equation: y = 196.05 + 228.22x (R² = 0.8226; p = 0.0024). The total PA content in the cheese sample with laccase on the surface increased up to 4 weeks and then decreased according to the equation: y = 629.1 + 621.45x - 59.539x2 (R² = 0.8984; p = 0.0163). The effect of laccase on the degradation of amines described in the literature has not been proven. However, this is a pilot experiment that investigates the effect of laccase on the content of polyamines in ripening cheeses. For a better understanding of this problematics, will be needed more extensive research.
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Study of the effect of microbial degradation of lignin on its molecular weight
Fialová, Tereza ; Hudečková, Helena (referee) ; Kovalčík, Adriána (advisor)
This diploma thesis deals with the microbial degradation of lignin using wood-degrading fungi Fomitopsis pinicola and Lenzites betulina. In the first phase of the experimental part, media screening was carried out, the aim of which was to select the best media for fungal growth and production of ligninolytic enzymes. Based on the determination of activity, in the next step, the conditions for their production were optimized in selected media. As part of these experiments, the optimal concentration of lignin and glucose was selected for subsequent cultivations. Commercially available lignin was used as substrate. Based on the results of measured enzyme activities, Lenzites betulina was selected as the better producer. The most produced enzyme of this fungus was laccase, followed by manganese-dependent peroxidase and the least lignin peroxidase. Further experiments were performed only with L. betulina. In the next part of the experiment, lignin was characterized, and its properties compared before and after microbial degradation. Its molecular weight was determined by SEC-MALS and the change in structure by FTIR. The molecular weight of lignin decreased after cultivation and a change in its structure was also noted. The results show that the microbial degradation of lignin was successful.
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Comparison of the Covalent Laccase Immobilization at Amino- and Carboxylfunctionalized Mesoporous Silica, Glassy Carbon, and Graphite Powders using Different Coupling Agents for Optimal Biosensor Preparation
Tvorynska, Sofiia ; Barek, J. ; Josypčuk, Bohdan
In order to find the most suitable immobilization protocol, a comparison of three strategies\nbased on the application of –NH2 and –COOH functionalized supports with the different\nactivation agents (glutaraldehyde and carbodiimide) have been conducted for the covalent\nenzyme (laccase) attachment. Two kinds of the supports, namely mesoporous silica (SBA−15,\nMCM−41) and carbonaceous (glassy carbon, graphite) powders, have been used. It was found\nthat a biosensor consisted of tubular detector of silver solid amalgam as a working electrode\nand the enzymatic mini-reactor with laccase covalently attached to glutaraldehyde activated\n–NH2 functionalized MCM−41 shows the best results regarding sensitivity and stability for\ndopamine detection.
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Study of the interaction between fungus Pleurotus ostreatus and bacterial cultures on the abiotic surfaces - morphological, biochemical and proteomic analysis
Kozická, Barbora ; Petráčková, Denisa (advisor) ; Konopásek, Ivo (referee)
Ligninolytic fungi are well known for their ability to degrade a wide range of xenobiotics contaminating the environment, including synthetic industrial dyes. In this work Pleurotus ostreatus was used for decolorization of a synthetic textile dye Remazol Brilliant Blue R (RBBR). To set up a model fungal "fixed-bed" bioreactor the fungus was immobilized on a polyurethane foam and artificially contaminated with a model bacterium Rhodococcus erythropolis. The development of bacterial contamination can be expected during a real application of fungal bio filters in wastewater treatment. The main aim of the work was to study interspecies interactions in the model bioreactors during the dye decolorization. Ligninolytic enzyme activities were followed in the bioreactor cultures as markers of fungal biodegradation ability. In contrast to the controls, no bacterial growth was observed in the P. ostreatus bioreactor culture liquid. The results showed that fungal laccase, pH of the culture liquid, and glucose consumption by the fungus had no effect on the bacterial growth. However, 4*105 - 1,3*106 CFU/ml of R. erythropolis was detected to be associated with the fungal solid support. The presence of these bacteria had no effect on the decolorization performance of the bioreactors. Dye decolorization efficiency...
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Laccase activity profiling in Trametes versicolor cultures degrading endocrine-disrupting compound Delor 103
Plačková, Martina
In this work endocrine disrupting potential of Delor 103, a commercial mixture of PCB congeners, was studied along with its effect on production of laccase by the ligninolytic fungus Trametes versicolor. Using a gene-reporter yeast assay for evaluation of hormonal activity Delor 103 showed an androgenic activity with an EC50 value of 2.29. 10-2 mg/l. Chlorbenzoic acids, Delor 103 potential metabolites resulting from microbial degradation, displayed on the other hand an estrogenic activity, indicating possible changes in hormonal activity of Delor 103 during its microbial degradation. The addition of Delor 103 to mineral medium T. versicolor cultures resulted in an up to 257times higher laccase activities detected in fungal cultures. Delor 103 induced enzymes showed different pI values from those of control cultures. In a complex malt-extract glucose medium (MEG) the stimulation effect of Delor 103 was kept down. Further, the production of laccase and synthesis of different pI forms depended strongly on the growth phase of fungal cultures. Exponencially growing cultures of T. versicolor were able to produce up to 7 different pI forms of laccase in responce to Delor 103 whereas stationary cultures produced only 4 enzyme forms with higher pI values. Stimulation of laccase activities in T. versicolor,...
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Study of the interaction between fungus Pleurotus ostreatus and bacterial cultures on the abiotic surfaces - morphological, biochemical and proteomic analysis
Kozická, Barbora ; Petráčková, Denisa (advisor) ; Konopásek, Ivo (referee)
Ligninolytic fungi are well known for their ability to degrade a wide range of xenobiotics contaminating the environment, including synthetic industrial dyes. In this work Pleurotus ostreatus was used for decolorization of a synthetic textile dye Remazol Brilliant Blue R (RBBR). To set up a model fungal "fixed-bed" bioreactor the fungus was immobilized on a polyurethane foam and artificially contaminated with a model bacterium Rhodococcus erythropolis. The development of bacterial contamination can be expected during a real application of fungal bio filters in wastewater treatment. The main aim of the work was to study interspecies interactions in the model bioreactors during the dye decolorization. Ligninolytic enzyme activities were followed in the bioreactor cultures as markers of fungal biodegradation ability. In contrast to the controls, no bacterial growth was observed in the P. ostreatus bioreactor culture liquid. The results showed that fungal laccase, pH of the culture liquid, and glucose consumption by the fungus had no effect on the bacterial growth. However, 4*105 - 1,3*106 CFU/ml of R. erythropolis was detected to be associated with the fungal solid support. The presence of these bacteria had no effect on the decolorization performance of the bioreactors. Dye decolorization efficiency...
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Laccase activity profiling in Trametes versicolor cultures degrading endocrine-disrupting compound Delor 103
Plačková, Martina
In this work endocrine disrupting potential of Delor 103, a commercial mixture of PCB congeners, was studied along with its effect on production of laccase by the ligninolytic fungus Trametes versicolor. Using a gene-reporter yeast assay for evaluation of hormonal activity Delor 103 showed an androgenic activity with an EC50 value of 2.29. 10-2 mg/l. Chlorbenzoic acids, Delor 103 potential metabolites resulting from microbial degradation, displayed on the other hand an estrogenic activity, indicating possible changes in hormonal activity of Delor 103 during its microbial degradation. The addition of Delor 103 to mineral medium T. versicolor cultures resulted in an up to 257times higher laccase activities detected in fungal cultures. Delor 103 induced enzymes showed different pI values from those of control cultures. In a complex malt-extract glucose medium (MEG) the stimulation effect of Delor 103 was kept down. Further, the production of laccase and synthesis of different pI forms depended strongly on the growth phase of fungal cultures. Exponencially growing cultures of T. versicolor were able to produce up to 7 different pI forms of laccase in responce to Delor 103 whereas stationary cultures produced only 4 enzyme forms with higher pI values. Stimulation of laccase activities in T. versicolor,...
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Biodegradation of 17alfa-ethinylestradiol by enzymes of ligninolytic fungi
Přenosilová, Lenka ; Stiborová, Marie (advisor) ; Černá, Věra (referee)
This work is aimed at the study of the effect of 17α-ethinylestradiol (EE2) on the production and characteristics of ligninolytic enzymes (laccase, Mn-dependent peroxidase and lignin peroxidase) in I. lacteus, T. versicolor, P. chrysosporium and P. ostreatus cultures grown on two types of liquid media. Enzyme activity production in fungal cultures was affected by the composition of culture medium. In the case of P. chrysosporium, the addition of EE2 to the complex- medium cultures led to a MnP activity stimulation and simultaneously LiP production was partially repressed in these cultures. In the mineral MM medium, no effect of EE2 on enzyme production by P. chrysosporium was observed. In EE2 treated MM cultures of P. ostreatus lower MnP activities were found when compared to biotic controls. In the case of T. versicolor cultures, the addition of EE2 to the complex medium caused laccase and LiP stimulation in the cultures. In the MM medium, however, only laccase production was affected by EE2. I. lacteus MnP production was partially repressed by EE2 in MM medium. In contrast to that, significantly higher MnP activities were detected in complex- medium I. lacteus cultures after the treatment with EE2. Further EE2 degradation by the fungal cultures was studied. The highest degradation effeciency was...
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