|
|
Biotechnological production of polyhydroxyalkanoates using wastes of coffee production
Vašíčková, Monika ; Benešová, Pavla (referee) ; Obruča, Stanislav (advisor)
This bachelor thesis is focused on study of production of polyhydroxyalkanoates (PHA) by bacteria Burkholderia cepacia and Burkholderia sacchari. Production of PHA has been studied using spent coffee grounds hydrolysates which have been prepared by different aproaches. In the thesis is compared standard method of coffee ground hydrolysis by mineral acid followed by enzymatic hydrolysis. Hydrolysates which have been produced were analyzed in terms of concentration of saccharides and efficiency of hydrolysis. The best producer of PHA (based on results) was bacterium Burkholderia cepacia, in the medium which has been prepared by hydrolysis of spent coffee grounds extracted by 60% solution of ethanol. Biomass yield of this cultivation was 3,553 g/l with 32,472% PHA content. This PHA contained 6,09% 3-hydroxyvalerate. In the other experiment, we verified alternative hydrolysis of spent coffee grounds – by commercially available enzymes. We used cellulase, hemicellulase and the enzymatic cocktail which has been produced by mould. Hemicellulase was the most effective hydrolytic enzyme and its application resulted in production of the highest amount of biomass – 5,708 g/l. In this cultivation, only homopolymer PHB has been which is probably caused by the fact, that during the enzymatic hydrolysis levulinic acid or any other potential precursor of 3HV is formed.
|
|
|
Influence of selected parameters on biotechnological production of polyhydroxyalkanoates
Eremka, Libor ; Pekař, Miloslav (referee) ; Obruča, Stanislav (advisor)
The aim of this work is to study microbial production of polyhydroxyalkanoates (PHA). Theoretical part is focused on production of PHA using microoganisms and transgenic plants. Bacterial strain Cupriavidus necator H16 was used for laboratory production of PHA. Various waste oils were used as sole carbon and energy source. Salt of propionic adic and 1-propanol were used as intermediate for 3-hydroxyvalerate monomer (3HV) unit. Incorporation of 3HV to polymer can improve material features of PHA. The major part of experimental work was focused to study influence of aeration (concentration of dissolved oxygen) to bacterial growth, selected metabolic pathways and formation of PHA. Furthermore, influence of aeration to monomer composition of polymer was evaluated. According to experimental conclusion of this work it was approved dependance between aeration and monomer composition of PHA. Moreover, it was approved that higher concentration of oxygen supports bacterial growth and influences PHA content in cells. In addition, NADPH is one of the substrates influencing flux of acetyl-CoA throughout the metabolism; higher intracellular concentration of NADPH inhibits TCA cycle and enhances accumulation of PHA in cells. For this reason, specific enzymatic aktivity of several selected intracelular enzymes were measured, including those enzymes which can generate NADPH.
|
|
|
METHODS OF POLY-3-HYDROXYBUTYRATE EXTRACTION AND THEIR OPTIMIZATION
Pospíšilová, Aneta ; Plachá,, Daniela (referee) ; Kowalczuk, Marek (referee) ; Přikryl, Radek (advisor)
Poly-3-hydroxybutyrát (PHB) je biolodegradovatelný a biokompatibilní termoplastický polymer se širokou škálou potenciálních aplikací. Výroba PHB typicky zahrnuje fermentaci obnovitelných surovin bakteriemi. Získávání PHB z bakterií je však často náročné a neefektivní. V této práci jsou zkoumány a porovnány různé metody pro získání PHB, včetně digesce, mechanického rozrušení a extrakce. Jsou diskutovány různé optimalizační strategie, které lze použít ke zlepšení účinnosti izolace PHB. Experimentální a výsledková část popisuje snahy o zlepšení metod izolace, zejména kyselé a mýdlové digesce, které byly vybrány jako potenciálně ekonomické, bezpečné a šetrné k životnímu prostředí. Společně s našimi průmyslovými partnery jsme tyto technologie úspěšně otestovali v čtvrtprovozním měřítku a plánuje se další vývoj. Řešily se také možné aplikace a komercializace PHB. Bylo zjištěno, že PHB může nahradit některé mikroplasty v kosmetice a že toto úsilí má potenciál, protože přítomnost mikroplastů v běžných výrobcích je stále vysoká a na úrovni Evropské unie je plánován jejich zákaz. Dále byla studována tepelná stabilita PHB v závislosti na metodě získávání a byly vyvinuty dvě stabilizační metody, které umožnily použít PHB získaný digescí za použití mýdla v materiálových aplikacích.
|
|
|
Screening of extremozyme production of selected extremophilic PHA producers
Dyagilev, Dmitry ; Obruča, Stanislav (referee) ; Pernicová, Iva (advisor)
This bachelor thesis deals with the screening of the production of extracellular hydrolytic enzymes in thermophilic microorganisms of the genera Aneurinibacillus, Brevibacillus, Chelatococcus, Pseudomonas, Schlegelella, Tepidimonas and Caldimonas. The ability of selected enzymes, namely proteases, lipases, amylases, xylanases, cellulases and pectinases, was tested in the investigated microorganisms. Such testing made it possible to assess in which microorganisms the production of specific enzymes can be observed. Based on the results of the screening, it was found that Schlegelella aquatica LMG 23380, Tepidimonas fonticaldi LMG 26746 and the investigated microorganisms of the genus Chelatococcus did not show the ability to produce any of the tested enzymes extracellularly. In natural isolates of Brevibacillus borstelensis LK 99 and Aneurinibacillus thermoaerophilus LK 102, only the ability to produce lipolytic enzymes was detected. The isolate Brevibacillus borstelensis Bz acts as a universal producer of all selected extremozymes. Enzyme activity was determined for selected producers. The bacterium Brevibacillus borstelensis Bz proved the ability to produce all six selected hydrolytic enzymes and has the highest activity of lipases, xylanases, cellulases and pectinases from the tested microorganisms. The highest proteolytic activity was measured in Thermomonas hydrothermalis DSM 14834 when cultured on skimmed milk powder.
|
|
|
Preparation and application particles from copolymer of P(3HB-co-4HB)
Ostanina, Kseniia ; Sedláček, Petr (referee) ; Obruča, Stanislav (advisor)
This bachelor thesis deals with preparation of particles from poly(3-hydroxybutyráte-co-4-hydroxybutyráte). Copolymer was dissolved in acetone and then the solution had been dosed into the anti-solvents. Acetone and anti-solvent are perfectly miscible. Theoretical part focuses on the properties of the copolymer, its production and potential use in medical applications. In the experimental part the copolymer was dripped into selected anti-solvents by syringe and pipette above and below the surface of anti-solvent. The following anti-solvent were chosen: water, methanol, dimethylsulfoxid (DMSO), 50 % acetone-water, 75% acetone-water, 50 % methanol-water, 75 % methanol-water mixtures. There were three copolymers used with different compositions of 4HB units (63 %, 87 % a 33 %). Copolymer composition was analyzed by gas chromatography. Copolymers were prepared by bacterial strain Aneurinibacillus sp. H1. Due to the results particles of size approximately 100-500 nm can be prepared by methods described in this work. The produced particles can be successfully used in medical applications.
|
|
|
Pathways Identification In The Gram-Positive Bacterium Aneurinibacillus Species H1
Musilová, Jana
Here, we present the first insight into the Gram-positive, promising polyhydroxyalkanoatesproducer Aneurinibacillus species H1. Both static and dynamic properties are described inthis paper with the aim on identification of pathways occurring in the organism. The genome consistsof a circular, 3,663,644 bp long chromosome and contains 4,654 protein-coding sequences and129 RNAs in total. The GC content is 44.8%. Functional properties identification showed that CDSdivide into 26 categories with the most prevalent Amino Acids and Derivatives group. Pathwaysinference revealed 201 pathways. The most represented group is metabolic pathways, which is furtherdivided into 12 groups.
|
|
|
Extraction of Poly(3-hydroxybutyrate) from Biomas
Jašek, Vojtěch ; Ing. Jiří Švrček, Ph.D (referee) ; Figalla, Silvestr (advisor)
Polyhydroxyalkanoates are a group of biopolymers which occur in cells of microorganisms. These biopolymers can be considered as alternatives to commonly used petrochemical materials such as polypropylene and polyethylene due to their physical and chemical properties. This bachelor thesis focuses on isolation of particular kind of PHA from biomass and contains both experimental and theoretical part. It consists of information about the origin and usage of PHA, challenges and problems of their biotechnological production and mainly various ways of downstream. Experimental part includes comparison of PHA extraction from biomass done by two ester solvents. Studied parameters were molecular weight of the polymer, solvent extraction ability and their resistance against hydrolysis. The main goal of this thesis was to verify the best process conditions of isolation done by nonhalogenated bio-solvent.
|
|
|
Dynamic Model for Production of Polyhydroxyalkanoates by Thermophilic Bacterium S. thermodepolymerans
Křápková, Monika ; Šafránek, David (referee) ; Sedlář, Karel (advisor)
Tato diplomová práce se zabývá rekonstrukcí dynamického modelu produkce polyhydroxyalkanoátů (PHA) termofilní bakterií Schlegelella thermodepolymerans. První kapitola poskytuje čtenářům krátký úvod do systémové biologie a matematické teorie grafů. Na ni navazuje druhá kapitola zabývající se různými přístupy v dynamickém modelování, včetně běžně používaných nástrojů pro dynamickou analýzu komplexních systémů. Třetí kapitola pak sleduje další pojmy a možnosti týkající se analýzy modelu. Následující kapitola se zaměřuje na metabolomiku a často používané laboratorní techniky a pátá kapitola je pak věnována polyhydroxyalkanoátům, zejména jejich chemické struktuře a vlastnostem. V kapitole šesté je navržen obecný booleovský model pro produkci PHA termofilními bakteriemi. Kapitola sedmá se poté zaměřuje na zdokonalení modelu se zaměřením na S. thermodepolymerans. Výsledný dynamický model je podroben analýze a výsledky jsou diskutovány.
|
|
|
PREPARATION AND CHARACTERIZATION OF BIOPOLYMER-BASED NANOPARTICLES AND NANOFIBERS AND THEIR APPLICATION IN FOOD AND COSMETICS
Kundrát, Vojtěch ; Vilčáková, Jarmila (referee) ; Kráčmar, Stanislav (referee) ; Márová, Ivana (advisor)
The presented dissertation thesis deals with the polymer polyhydroxybutyrate and other biopolymers as a basic building block for the construction of micro- and nanoscopic structures and materials used in food and cosmetics. In the theoretical part, current literary review is prepared to introduce the basics of this application field. The practical part of the work is composed of three blocks developed during the doctoral study. In the first and most important part are summarized comments to the academic and patent outputs, where among the academic ones it is possible to find two peer-reviewed articles dealing with the electrostatic and wet spinning of PHB and properties of prepared materials. The patent outputs consist of several accepted and applied projects, which summarize results on both PHB spinning methods, but also on general approaches enabling the processing of PHB into forms enabling many applications in food and cosmetics. Second part was focused on the patented composition of the UV protection cream based on the prepared nanoscopic and micro- morphologies of PHB. The third block summarizes results focused predominantly on the electrostatic spinning of PHB and other biopolymers. Finally, a short chapter containing a brief description of projects that were in a way related to the dissertation topic, but rather practical development work in Central Tanzania and West Africa, which draw on knowledge and contacts gained during studies at FCH BUT Brno.
|
|
|
Screening of biotechnological potential of selected members of the genus Geobacillus and other related genuses
Kouřilová, Xenie ; Brázda, Václav (referee) ; Obruča, Stanislav (advisor)
This diploma thesis deals with selected thermophilic representatives of genera Geobacillus, Saccharococcus and Bacillus, taking screening of its biotechnological potential into account. Bacteria from the first two genera came from Czech and German collection of microorganisms, while bacteria of genus Bacillus were natural isolates. Researched strains were examined from a viewpoint of carbon source utilization and furthermore, production of biosurfactants, extracellular hydrolytic enzymes (protease, amylase, lipase, cellulase, xylanase), organic acids, antimicrobial agents and microbial plastics – polyhydroxyalkanoates was also tested. Bacteria S. thermophilus, G. uzenensis and G. zalihae evinced a substantial ability of biosurfactant production. Strains G. jurassicus, G. uzenensis, G. gargensis and G. lituanicus were capable of intensive production of all tested, technologically significant enzymes. Highest antimicrobial effects were reached with bacteria G. stearothermophilus and G. thermocatenulatus. Largest production of acetic acid was achieved with G. jurassicus and lactic acid with G. thermodenitrificans. Ability to produce polyhydroxyalkanoates was proved at genotype level by some cultures only, however at fenotype level, response was negative. On the contrary, bacteria genus Bacillus were able to produce polyhydroxyalkanoates, although in small amounts under given circumstances. With remaining researched metabolites, production ability was considerably lower, compared to genera Geobacillus and Saccharococcus.
|
guest ::
