Cultivation of carotenogenic yeasts on waste substrates usinf solid state fermentation

„Semi-solid fermentation“ je čoraz viac využívanou technikou k produkcii významných metabolitov, či obohatenej biomasy za nízkych finančných nákladov, nízkej spotreby kultivačnej vody a zároveň s malou záťažou na životné prostredie. Problémom tejto techniky ale môže byť voľba vhodného mikroorganizmu, ktorý je schopný rásť a produkovať pri nízkej aktivite vody. Predložená práca je poňatá ako pilotná štúdia troch kvasinkových kmeňov Rhodotorula glutinis, Cystofilobasidium capitatum a Sporobomomyces roseus kultivovaných technikou „semi-solid state fermentation“ (semi-SSF). Kvasinky boli kultivované v sériách produkčných médií s postupným znižovaním obsahu kultivačnej vody. Ako uhlíkatý zdroj sacharidov boli použité cestoviny a hydrolyzované cestoviny, v kontrolných médiách glukóza. Všetky študované kmene boli schopné rásť a produkovať sledované lipidické metabolity i pri nízkych obsahoch vody. Produkcie karotenoidov a sterolových látok boli vyššie práve v médiách s nízkou aktivitou vody. Bunky sa tak pravdepodobne chránili pred väčším osmotickým tlakom. Ako najlepší producent karotenoidov i biomasy sa ukázal kmeň Cystofilobasidium capitatum, kultivovaný na médiu s hydrolyzovanými cestovinami a obsahom vody 40%. Semi-SSF sa javí ako dobrá technika pre selekciu kmeňov s nadprodukčnými vlastnosťami. Výberom vhodného produkčného média a koncentrácie vody sa dá optimalizovať stimulácia produkcie sledovaných metabolitov v kvasinkových bunkách.Semi-solid fermentation is an eco - friendly technique more and more used for production of significant metabolites or enriched biomass at low entrance cost and low comsumption of water. The problem of this technique might be the right choice of microorganism able to grow and produce at low water activity. This work is a pilot study of three red yeast strains – Rhodotorula glutinis, Cystofilobasidium capitatum and Sporobomomyces roseus cultivated by semi-solid state fermentation (semi-SSF). Yeasts were cultivated in series of production media with gradual reduction of cultivation water content. Pasta and hydrolyzed pasta were used as source of sacharides, glucose served as the carbon source in control media. All studied strains we able to grow and produce observed lipidic metabolites also at low water contents. Production of carotenoids and sterols was higher in semi-solid media. Cystofilobasidium capitatum strain was identified as the best producer of carotenoids and biomass. This strain was cultivated on hydrolyzed pasta media with 40% water content. Semi-SSF seems to be an adequate technique for selection of strains having over-productive properties. Observed metabolites production in yeast cells can be optimized by choosing the appropriate production media and water activity.

Determination of protective properties of Bardejovske Kuple spa curative waters by rotational viscometry and ABTS assay

Mineral waters from Bardejovske Kupele spa are natural, strongly mineralized, with healing effects. They are classified as hydrocarbonic - containing chloride and sodium - carbonic, cold, hypotonic, with a relatively high boric acid content. Potential anti-oxidative effects of curative waters from Bardejovske Kupele were investigated against the hyaluronan (HA) degradation. High-molar-mass HA was exposed to the action of ascorbate and cupric ions, which initiate free-radical chain degradation. Time-dependent changes of dynamic viscosity (h) of the HA solutions were monitored by rotational viscometry. The radical scavenging capacity of curative waters was determined by the ABTS assay. Despite a significantly high content of transition metal ions, especially iron, remarkable protective effects of the two curative spa waters were found, namely Alzbeta and Klara. Even though “Alzbeta´s“ iron content was 3.5-fold higher than “Klara´s“, “Alzbeta“ was shown to have better protective properties against the HA degradation compared to “Klara“. Bolus addition of ferric ions to the reaction system instead of the natural iron-containing curative water caused a significant HA degradation. The ABTS decolorization assay revealed that the curative spa waters were proven as poorly effective scavengers of the ABTS·+ cation radical

New multiresponsive chromic soft materials: dynamic interconversion of short 2,7-dicyanomethylenecarbazole-based biradicaloid and the corresponding cyclophane tetramer

This work reports on a quinodimethane-type molecule, 2,7-dicyanomethylene-9-(2-ethylhexyl)carbazole (1), one of the shortest π-conjugated biradicaloids reported to be stable in solution under ambient conditions. This carbazole-based quinoidal precursor is able to form a macrocyclic σ-bonded tetramer (2). The resolved single-crystal X-ray structure of tetramer 2 shows that four molecules of 1 are linked together through four long (CN)2C−C(CN)2 bonds (1.631 Å) resulting from coupling of the unpaired electrons in biradicaloid 1. Dynamic interconversion between monomer 1 and cyclophane tetramer 2 is achieved by reversible cleavage and recovery of the four (CN)2C−C(CN)2 bonds upon soft external stimuli (light absorption, temperature and pressure), which is accompanied by significant color changes. These novel photo-, thermo-, and mechanochromic properties expand the versatility of π-conjugated biradicaloid compounds as novel functional materials that, in combination with spin chemistry and dynamic covalent chemistry, can be relevant in molecular machines, sensors, and switches

Study of Metabolic Adaptation of Red Yeasts to Waste Animal Fat Substrate

Carotenogenic yeasts are non-conventional oleaginous microorganisms capable to utilize various waste substrates. In this work 4 red yeast strains (Rhodotorula, Cystofilobasidium and Sporobolomyces sp.) were cultivated in media containing crude, emulsified and enzymatically hydrolysed animal waste fat, compared with glucose and glycerol as single C-sources. Cell morphology (cryo-SEM, TEM), production of biomass, lipase, biosurfactants, lipids (GC/FID) carotenoids, ubiquinone, ergosterol (HPLC/PDA) in yeast cells was studied depending on medium composition, C-source and C/N ratio. All studied strains are able to utilize solid and processed fat. Biomass production at C/N=13 was higher on emulsified/hydrolysed fat than on glucose/glycerol. Production of lipids and lipidic metabolites was enhanced for several times on fat; the highest yields of carotenoids (24.8 mg/l) and lipids (54.5%/CDW) were found in S.pararoseus. Simultaneous induction of lipase and biosurfactants was observed on crude fat substrate. Increased C/N ratio (13-100) led to higher biomass production in fat media. Production of total lipids increased in all strains to C/N 50. Oppositely, production of carotenoids, ubiquinone and ergosterol dramatically decreased with increased C/N in all strains. Compounds accumulated in stressed red yeasts are having great application potential and can result from valorization of animal waste fat in the biorefinery concept.Karotenogenní kvasinky jsou schopny utilizovat řadu různých substrátů včetně odpadů. V této práci je popsána kultivace vybraných kmenů karotenogenních kvasinek rodů Rhodotorula, Cystofilobasidium, and Sporobolomyces sp. v médiu obsahujícícm surový odpadní živočišný tuk a upravený substrát emulsifikací a enzymovou hydrolýzou. Výsledky byly srovnány s kultivací na glukóze a glycerolu jako jednoduchých zdrojích uhlíku

Revealing the Potential of Lipid and beta-Glucans Coproduction in Basidiomycetes Yeast

Beta (beta)-glucans are polysaccharides composed of D-glucose monomers. Nowadays, beta-glucans are gaining attention due to their attractive immunomodulatory biological activities, which can be utilized in pharmaceutical or food supplementation industries. Some carotenogenicBasidiomycetesyeasts, previously explored for lipid and carotenoid coproduction, could potentially coproduce a significant amount of beta-glucans. In the present study, we screened elevenBasidiomycetesfor the coproduction of lipids and beta-glucans. We examined the effect of four different C/N ratios and eight different osmolarity conditions on the coproduction of lipids and beta-glucans. A high-throughput screening approach employing microcultivation in microtiter plates, Fourier Transform Infrared (FTIR) spectroscopy and reference analysis was utilized in the study. Yeast strainsC. infirmominiatumCCY 17-18-4 andR. kratochvilovaeCCY 20-2-26 were identified as the best coproducers of lipids and beta-glucans. In addition,C. infirmominiatumCCY 17-18-4,R. kratochvilovaeCCY 20-2-26 andP. rhodozymaCCY 77-1-1 were identified as the best alternative producers of beta-glucans. Increased C/N ratio led to increased biomass, lipid and beta-glucans production for several yeast strains. Increased osmolarity had a negative effect on biomass and lipid production while the beta-glucan production was positively affected

New Water-Soluble Copper(II) Complexes with Morpholine-Thiosemicarbazone Hybrids: Insights into the Anticancer and Antibacterial Mode of Action

Six morpholine-(iso)thiosemicarbazone hybrids HL1-HL6 and their Cu(II) complexes with good-to-moderate solubility and stability in water were synthesized and characterized. Cu(II) complexes [Cu(L1-6)Cl] (1-6) formed weak dimeric associates in the solid state, which did not remain intact in solution as evidenced by ESI-MS. The lead proligands and Cu(II) complexes displayed higher antiproliferative activity in cancer cells than triapine. In addition, complexes 2-5 were found to specifically inhibit the growth of Gram-positive bacteria Staphylococcus aureus with MIC50 values at 2-5 μg/mL. Insights into the processes controlling intracellular accumulation and mechanism of action were investigated for 2 and 5, including the role of ribonucleotide reductase (RNR) inhibition, endoplasmic reticulum stress induction, and regulation of other cancer signaling pathways. Their ability to moderately inhibit R2 RNR protein in the presence of dithiothreitol is likely related to Fe chelating properties of the proligands liberated upon reduction

New Water-Soluble Copper(II) Complexes with Morpholine-Thiosemicarbazone Hybrids: Insights into the Anticancer and Antibacterial Mode of Action

Six morpholine-(iso)­thiosemicarbazone hybrids HL1–HL6 and their Cu­(II) complexes with good-to-moderate solubility and stability in water were synthesized and characterized. Cu­(II) complexes [Cu­(L1–6)­Cl] (1–6) formed weak dimeric associates in the solid state, which did not remain intact in solution as evidenced by ESI-MS. The lead proligands and Cu­(II) complexes displayed higher antiproliferative activity in cancer cells than triapine. In addition, complexes 2–5 were found to specifically inhibit the growth of Gram-positive bacteria Staphylococcus aureus with MIC50 values at 2–5 μg/mL. Insights into the processes controlling intracellular accumulation and mechanism of action were investigated for 2 and 5, including the role of ribonucleotide reductase (RNR) inhibition, endoplasmic reticulum stress induction, and regulation of other cancer signaling pathways. Their ability to moderately inhibit R2 RNR protein in the presence of dithiothreitol is likely related to Fe chelating properties of the proligands liberated upon reduction