|
|
Production of biologically active substances by yeasts and algae cultivated on different nutrition sources
Kohútová, Darina ; Němcová, Andrea (referee) ; Márová, Ivana (advisor)
Carotenoids are lipophilic pigments and the most widespread antioxidants occurring in nature with great application potencial in the cosmetics, pharmaceuticals and food industry. Lipids are one of the essential organic substances of living organisms as a nutrition source and energy supply. This work deals with the optimalization of nutritional characteristics of cultivation medium for cultivation of carotenogenic yeasts and algae. Two waste substrates were used and different nitrogen sources, NaCl concentrations, and various C/N ratios were applied. The biomass obtained was analysed by chromatography, and the production of carotenoids, ergosterol, coenzyme Q and lipids was evaluated. Of the tested nitrogen sources urea was the most favourable for yeasts, and sodium nitrate for algae. Higher salt concentration had a negative effect on yeast growth but increased degree of fatty acid unsaturation. Tested C/N led different effects: while the C/N ratio 16 most suitable for growth, the increasing C/N ratio caused the increase of the polyunsaturated fatty acids content. Of the used yeast strains, Rhodotorula mucilaginosa and Sporidiobolus salmonicolor exhibited the best production characteristics, of the used algae strains, it was Chlorella.
|
|
|
Use of diffrent nutrition source by selected carotenogenic yeasts
Čačková, Katarína ; Starečková, Terezie (referee) ; Márová, Ivana (advisor)
Carotenoids are the most common naturally occurring terpenoid pigments. Because of their positive biological functions, carotenoids represent a group of important molecules for the pharmaceutical, chemical, food and feed industries. This work was realized as a comparative study focused on use of some waste substrates as nutritional sources for production of carotenoid pigments by red yeast Rhodotorula aurantiaca. Yeast were cultivated in different media with selected waste substrates – apple fiber, whey, wheat etc. To some media hydrolytic enzymes prepared from some fungal strains were added. Most of waste substrates seems to be useful for R. aurantiaca cultivation. As the best nutritional sources for enriched biomass and carotenoid production apple fiber or wheat processed by mixed preparative of extracellular hydrolases from Fusarium solani were found.
|
|
|
Use of Waste Substrates to Production of Enriched Yeast Biomass
Starečková, Terezie ; Demnerová, Kateřina (referee) ; Vávrová, Milada (referee) ; doc.PharmDr.Petr Babula, Ph.D. (referee) ; Márová, Ivana (advisor)
Yeasts are like other organisms constantly exposed to environmental influences. Their survival depends on the skills to adapt to environmental changes, including the ability to use various alternative sources of nutrients. In presented PhD thesis carotenogenic yeast belonging to the genera Rhodotorula, Sporobolomyces and Cystofilobasidium were tested for ability to use of selected waste substrates, and also subjected to several types of exogenous stress effects and mutations in order to increase the production of microbial biomass enriched with specific metabolites. As alternative nutrient sources derived from waste substrates from agricultural and farm production apple peel, pulp, corn germ and more were tested. Yeasts were also exposed to osmotic, oxidative and combined stress (benefits of various concentrations of NaCl and H2O2 to the culture media), followed by metal ions of selenium and chromium in concentrations of 0.01 mM, 0.1 mM and 1 mM. The effect of mutagen methanesulfonic acid ethyl ester was tested too. In all experiments the adaptivity of cells, morphological changes, color pigments produced by the media while some important fungal metabolites production and changes in chromosomal DNA fragmentation were analyzed. In order to evaluate potential changes in the yeast genome after treatment with mutagen and stress factors methods for isolation of intact chromosomal DNA and DNA analysis by pulsed field gel electrophoresis was optimized. The amount of produced metabolites was mainly analyzed by RP-HPLC with UV/VIS and MS detection. The work has been shown that most strains are able to use waste substrates and produced selected target metabolites. Biomass, for example, in R. aurantiaca on apple fiber was about 7 g/l and in C. capitatum cultivated on modified whey reached to 9 g/l. Amount of produced carotenoids by R. aurantiaca cultivated on wheat germ and maize after enzymatic hydrolysis by F. solani was 1.01 mg/g and S. roseus on pasta 4.3 mg/g. The values of ergosterol synthesis in R. aurantiaca are on the apple shells around 4.8 mg/g, in S. roseus on pasta with the enzymatic hydrolysis of P. chrysosporium 8.9 mg/g. The best substrate for biomass production and induction of carotenoids are waste substartes containing a mixture of simple and complex carbohydrates enriched with the addition of nitrogen compounds. Potential cytotoxic effect of stress factors of low concentrations was demonstrated. Red yeast genome was able to distribute by optimized PFGE, the karyotype of tested yeasts contain 11 or more chromosomes with visible differences between yeast species and genera. During exchange internship the ability of recombinant yeast S. cerevisiae to convert xylose to xylitol, which would be achieved by increasing the production of bioethanol as alternative fuel sources was studied. It turned out that both ligninocellulose materials to bioethanol production, as well as various waste substrates for microbial synthesis of carotenoids would reduce costs for industrial production of yeast metabolites, as well as to reduce the negative burden on the environment.
|
|
|
Production of selected metabolites by carotenogenic yeasts cultivated on food wastes
Turková, Lucie ; Hlaváček, Viliam (referee) ; Márová, Ivana (advisor)
The production of selected metabolites, especially carotenoids, ergosterol and coenzyme Q was observed in four red yeast strains (Cystofilobasidium capitatum, Rhodotorula glutinis, Sporobolomyces shibatanus a Sporobolomyces roseus) cultivated on viticulture waste substrates such as skins and stem. In these substrates the content of carbohydrates, fat, tannins and polyphenols was analysed. Cultivations were done also on chemically and enzymatically hydrolysed substrates. For enzyme hydrolysis the extracellular enzyme cocktails of the fungi Phanerochaete chrysosporium and Penicillium lilacinum were used. The highest production of metabolites was observed in S. roseus cultivated on stem hydrolysed by enzymes from P. chrysosporium. The yield of -carotene reached 616,1 g/g dry mass, carotenoids 753,6 g/g dry mass, ergosterol 415,5 g/g dry mass and coenzym Q 77,9 g/g dry mass. S. roseus reached very high yields also in the other series of cultivation. The lowest production of metabolites was observed in S. shibatanus. The yield coefficients of biomass and -carotene were calculated based on consumption of reducing sugars.
|
|
|
Using of waste substrates for the lipid production by Metschnikowia yeasts
Cagáňová, Linda ; Márová, Ivana (referee) ; Němcová, Andrea (advisor)
This thesis was focused on study of biotechnological utilization of waste substrates to produce lipids by yeast of the genus Metschnikowia. Waste materials and their subsequent transformation into high value-added products such as microbial lipids are currently considered as an alternative source for biofuel production. Therefore, the experimental part was aimed at investigating the influence of a carbon source to the controlled overproduction of lipids by yeast Metschnikowia. Total of 12 yeast strains of the genus Metschnikowia were selected. Yeast strains M. pulcherrima , M. pulcherrima 147, M. pulcherrima 149, M. andauensis 129 a M. fructicola 15 were purchased from Culture Collection of Yeasts (CCY, Bratislava, Slovakia). The growth characteristics of this yeast strains were also studied. It may serve to better understanding of the physiology of the yeast strains and also to help in further analysis of the produced metabolites. The other strains M. chrysoperlae 1158, M. pulcherrima 1232, M. fructicola 1235, M. andauensis 1241, M. sinensis 1244, M. zizyphicola 1247 a M. shanxiensis 1250 were purchased from CBS (Centraalbureau voor Schimmelcultures, Utrecht, the Netherlands).Yeast strains were cultivated on crude animal fat, glycerol and cheese whey under conditions of different C/N ratios. Because of higher lipid yields, cultivation was carried out at 14°C for 14 days. The accumulated lipid content was determined by gas chromatography and Raman spectroscopy. The glycerol-containing medium was evaluated as the most suitable for microbial lipids production. The total amount of lipids present in cells of M. pulcherrima 1232 was 36,31%. At the same time, quantitative screening of lipase enzymatic activity in Metschnikowia yeast was performed using spectrophotometric method with p-NPP. Controlled production of lipolytic enzymes has been monitored by using two types of media: crude animal fat and crude animal fat with addition of emulsifier (Tween 80). The conclusion of the work was supplemented by analysis of the karyotype of yeasts of the genus Metschnikowia using the technique of pulsed gel electrophoresis.
|
|
|
Bioproduction, Characterization and Application of Selected Biopolymers
Benešová, Pavla ; Pekař, Miloslav (referee) ; Čertík, Milan (referee) ; Márová, Ivana (advisor)
Proposed doctoral thesis deals with microbial production of biopolymers (polysaccharides and polyesters) using renewableble inexpensive resources as carbon and nitrogen source. At first, production of extracellular metabolites, mainly pullulan and poly-L-malic acid, by polymorphic microorganism Aureobasisium pullulans was studied. Further part of thesis was focused on production of bacterial biopolyesters polyhydroxyalkanoates (PHA). PHA are produced in the form of intracellular granules by various bacteria species. During the study of PHA production, sythesis of poly(3-hydroxybutyrate) and technologically significant copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate) was investigated as well with regard to waste substrate utilization in biorafinery concept. Spent coffee ground seemed to be applicable waste in biorefinery proces. One of the coffee wastes used for PHA production was spent coffee ground hydrolysate which was converted to PHA by Burkholderia cepacia. Moreover, copolymer of 3HB and 3HV was accumulated when SCGH was used as a sole carbon source, no precursor had to be added. Another coffee waste, which was utillized by Cupriavidus necator H16, is represented by oil extracted from spent coffee grounds. Hydrolysis proces of waste substrates had to be optimized, to reach the highest possible saccharide concentration. Due to the hydrolysis process several toxic compounds can be formed, hence, influence of hydrolysis procesings on polyphenols content in hydrolysates was monitored. Waste substrates were also tested as potentional complex nitrogen source for PHA production. Hydrolysates of cheese whey and chicken feather were used as inexpensive complex nitrogen source for PHA production by C. necator, when waste frying oil was used as a carbon substrate. The finnal part of the thesis is dealing with possibilities how the PHA can be processed into atractive aplication forms. Liposomes with partial content of PHB had been prepared, moreover, for improving of antioxidant activity and antimicrobial effect the waste coffee oil was added. Micro-fibrous PHB materials were prepared for the purposes of water filtration. Higher filtration effectivity was observed when composite PHB fibrous material containing active charcoal or metal oxides was used. Important part of thesis is represented by study of degradation processes of prepared PHA-based aplication forms. Various microorganisms were tested on PHA depolymerases production. Biodegradability of PHA materials was tested in of composting conditions by standard composting test IS/ISO 20200. Changes in mechanical properties of polyurethan, where polyol was partialy replaced by PHB, were tested depending on the exposure of composting condition.
|
|
|
Production of carotenoids by red yeasts grown on some waste substrates
Kubáčková, Martina ; Čarnecká, Martina (referee) ; Márová, Ivana (advisor)
Carotenoids belong to the most widespread and abundant classes of natural pigments with utilised in food industry, pharmacy and cosmetics. Presented work was realized as a comparative study of some substrates for cultivation of red yeast Rhodotorula glutinis, which produces carotenoid pigments. Yeasts were cultivated in different media with several waste substrates (for example potato fiber, apple waste, cereals, grains, etc.) as nutrition sources. To some substrates hydrolytic enzymes isolated from the fungi genus Fusarium were added. Majority of waste substrates were acceptable for cultivation. However, the best conditions for production of carotenoid enriched biomass (6,4 g/l of biomass enriched 3,2 mg /g of beta-carotene) exhibited media with hydrolytic fungal enzymes.
|
|
|
Microbial production of carotenoid pigments using waste substrates
Němcová, Andrea ; Ing.Martina Čarnecká, Ph.D. (referee) ; Márová, Ivana (advisor)
Carotenoids are naturally occurring pigments produced by bacteria, yeasts, filamentous fungi and plants. They exhibit significant biological effects and are widely used in the food industry, pharmacy and cosmetics. The aim of this diploma thesis proposed as a comparative study was regulation of carotenoid and ergosterol production in red yeasts using several waste substrates as whey, corn germs, wheat, apple fiber and pasta. To selected production media extracellular hydrolytic enzymes degrading polysaccharide were added. These enzymes were obtained from the cultivation media of four fungal strains. In this study three carotenogenic yeast strains were used: Rhodotorula glutinis, Sporobolomyces roseus and Cystofilobasidium capitatum. All strains were cultivated simultaneously and changes in biomass and carotenoid production in different production media were monitored and compared. As the best waste substrate apple fiber was utilized, particularly in Rhodotorula glutinis, which exhibited mainly biomass production increase. In Sporobolomyces roseus increased production of biomass and carotenoids have been reported in media with hydrolyzed fiber and pasta as well. Beta-carotene production in this strain reached 4776,38 mg/g of dry weight. The strain Cystofilobasidium capitatum exhibited in waste media a decerase of biomass production accompanied with increased production of carotenoids, especially in wheat mush and pasta medium hydrolyzed by enzyme preparative from Aureobasidium pullulan. It can be concluded that hydrolyzed waste substrates are very hopeful as cheap nutrient sources for yeast strains producing carotenoids and ergosterol. Nevertheless, further study of substrate processing for individual strains is needed.
|
|
|
Production of Selected Microbial Metabolites and Energy Using Different Waste Materials
Petrik, Siniša ; Rychtera, Mojmír (referee) ; Němec, Miroslav (referee) ; Pekař, Miloslav (referee) ; Márová, Ivana (advisor)
Pro zpracování a nakládání s odpadními substráty lze použít řadu postupů a možností. Stále se rozšiřující spektrum metod a technologií umožňuje další využití materiálů a energie ve formě obnovitelných zdrojů. Jedním z řešení pro zpětné získávání některých odpadních materiálů je využití tzv. bílé (průmyslové) biotechnologie, která zahrnuje praktickou aplikaci metabolických aktivit celé řady různých mikroorganizmů včetně jejich specifických biologických drah k produkci látek s vysokou přidanou hodnotou. V předložené práci screeningového typu bylo pro zhodnocení odpadních surovin využito několik druhů mikroorganizmů kultivovaných za různých specifických podmínek včetně kultivace na odpadních materiálech získaných zejména ze zemědělství a potravinářství. Cílem bylo získání vybraných typů průmyslově cenných metabolitů, případně energie. Předložená studie byla zaměřena na srovnání růstu a produkčních vlastností několika kmenů karotenogenních kvasinek rodu Rhodotorula, Sporobolomyces a Cystofilobasidium, kultivovaných v médiích s obsahem glycerolu (technický a odpadní glycerol), dále v médiích obsahujících pšeničnou slámu, hydrolyzovanou slámu zpracovanou v hydrotermálním procesu při vysoké teplotě a zbytky po filtraci hydrolyzátu. Dalším testovaným odpadním substrátem byla syrovátka. Všechny testované kvasinky byly schopny využít glycerol jako jediný zdroj uhlíku. Produkce biomasy při kultivaci na technickém glycerolu se více či méně přibližovala kontrole (cca 7 - 10 gl-1), zatímco při kultivaci na odpadním glycerolu byla produkce vyšší (10.9 - 14.5 gl-1). Produkce karotenoidů a ergosterolu byla vyšší v glukózovém médiu než v médiu s obsahem glycerolu. Všechny testované kvasinky byly rovněž schopny produkovat neutrální lipidy, a to v rozmezí 11 - 15 %, s výjimkou C. capitatum, kde produkce dosahovala více než 22 % obsahu neutrálních lipidů. Pšeničná sláma a produkty z ní připravené se ukázaly být využitelnými substráty s vysokým potenciálem pro produkci biomasy i metabolitů, a to zejména u kmene S. roseus. Syrovátka, jako odpadní produkt mlékarenství, byla účinně využita jako substrát pro kokultivaci karotenogenních kvasinek a bakterií mléčného kvašení. Kokultivační proces může vyvolat nadprodukci pigmentů a ergosterolu, přičemž získaná biomasa díky obohacení o bakterie L. casei dosahovala vyšší kvality. Za účelem energetického využití mikrobiálního metabolismu formou mikrobiálních palivových článků, tzv. „Microbial Fuel Cell“ byla aplikována směsná kultura bakterií získaných z čistírny odpadních vod. Tyto mikroorganizmy hrají významnou roli při výrobě elektrické energie a současně také při čištění odpadních vod. Elektřina je generována přímo z organických látek přítomných v kultivačním médiu a lze ji použít pro provoz čistírny samotné a případně i pro další aplikace.
|
|
|
Production, characterization and application of polymers of malic and glutamic acids
Čangelová, Katarína ; Kučera, Dan (referee) ; Obruča, Stanislav (advisor)
Subject of this bachelor thesis was study of biopolymers production using microorganisms. Polymer of malic acid was produced by Aureobasidium pullulans, a yeast-like fungus. Biosynthesis of polymer of glutamic acid was carried out by Bacillus licheniformis bacterium. Particular polymers, their microbial producers and possible application in different areas are characterised in the theoretical part of the thesis. Through experiments, the effect of various conditions on cell growth and production of these polymers was studied. Cultivation conditions were optimised in terms of polymers production. Subsequently, waste substrates - whey and poultry feathers were used. Microbially-produced polymer of malic acid was hydrolysed with 2 M sulfuric acid. Concentration of malic acid was analysed by IEC with a conductivity detector. Concentration of polymer of glutamic acid was determined by a spectrophotometric method using CTAB.
|
guest ::
