Low production rates are still one limiting factor for the industrial climate-neutral production of biovaluable compounds in cyanobacteria. Next to optimized cultivation conditions, new production strategies are required. Hence, the use of established molecular tools could lead to increased product yields in the cyanobacterial model organism Synechocystis sp. PCC6803. Its main storage compound glycogen was chosen to be increased by the use of these tools. In this study, the three genes glgC, glgA1 and glgA2, which are part of the glycogen synthesis pathway, were combined with the Pcpc560 promoter and the neutral cloning site NSC1. The complete genome integration, protein formation, biomass production and glycogen accumulation were determined to select the most productive transformants. The overexpression of glgA2 did not increase the biomass or glycogen production in short-term trials compared to the other two genes but caused transformants death in long-term trials. The transformants glgA1_11 and glgC_2 showed significantly increased biomass (1.6-fold - 1.7-fold) and glycogen production (3.5-fold - 4-fold) compared to the wild type after 96 h making them a promising energy source for further applications. Those could include for example a two-stage production process, with first energy production (glycogen) and second increased product formation (e.g. ethanol).
- MeSH
- glykogen MeSH
- Synechocystis * genetika MeSH
- Publikační typ
- časopisecké články MeSH
Microalgae accumulate bioavailable selenium-containing amino acids (Se-AAs), and these are useful as a food supplement. While this accumulation has been studied in phototrophic algal cultures, little data exists for heterotrophic cultures. We have determined the Se-AAs content, selenium/sulfur (Se/S) substitution rates, and overall Se accumulation balance in photo- and heterotrophic Chlorella cultures. Laboratory trials revealed that heterotrophic cultures tolerate Se doses ∼8-fold higher compared to phototrophic cultures, resulting in a ∼2-3-fold higher Se-AAs content. In large-scale experiments, both cultivation regimes provided comparable Se-AAs content. Outdoor phototrophic cultures accumulated up to 400 μg g-1 of total Se-AAs and exhibited a high level of Se/S substitution (5-10%) with 30-60% organic/total Se embedded in the biomass. A slightly higher content of Se-AAs and ratio of Se/S substitution was obtained for a heterotrophic culture in pilot-scale fermentors. The data presented here shows that heterotrophic Chlorella cultures provide an alternative for Se-enriched biomass production and provides information on Se-AAs content and speciation in different cultivation regimes.
- MeSH
- aminokyseliny analýza metabolismus MeSH
- biomasa MeSH
- Chlorella klasifikace růst a vývoj metabolismus účinky záření MeSH
- fototrofní procesy MeSH
- heterotrofní procesy MeSH
- mikrořasy chemie růst a vývoj metabolismus účinky záření MeSH
- selen analýza metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
We have worked out a rapid 1-day test based on photosynthesis measurements to estimate suitable growth temperature of microalgae cultures. To verify the proposed procedure, several microalgae-Chlorella, Nostoc, Synechocystis, Scenedesmus, and Cylindrospermum-were cultured under controlled laboratory conditions (irradiance, temperature, mixing, CO2, and nutrient supply) to find the optima of photosynthetic activity using the range between 15 and 35 °C. These activities were recorded at each temperature step after 2 h of acclimation which should be sufficient as oxygen production and the PQ cycle are regulated by fast processes. Photosynthetic activity was measured using three techniques-oxygen production/respiration, saturating pulse analysis of fluorescence quenching, and fast fluorescence induction kinetics-to estimate the temperature optima which should correspond to high growth rate. We measured all variables that might have been directly related to growth-photosynthetic oxygen evolution, maximum photochemical yield of PSII, Fv/Fm, relative electron transport rate rETRmax, and the transients Vj and Vi determined by fast fluorescence induction curves. When the temperature optima for photosynthetic activity were verified in growth tests, we found good correlation. For most of tested microalgae strains, temperature around 30 °C was found to be the most suitable at this setting. We concluded that the developed test can be used as a rapid 1-day pre-screening to estimate a suitable growth temperature of microalgae strains before they are cultured in a pilot scale.
- MeSH
- Chlorella růst a vývoj metabolismus účinky záření MeSH
- fotosyntéza MeSH
- kinetika MeSH
- kultivační techniky metody MeSH
- kyslík metabolismus MeSH
- mikrořasy růst a vývoj metabolismus účinky záření MeSH
- Scenedesmus růst a vývoj metabolismus účinky záření MeSH
- sinice růst a vývoj metabolismus účinky záření MeSH
- světlo MeSH
- teplota MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
In this work, the key moments of the development of the so-called thin-layer cascades (TLC) for microalgae production are described. Development started at the end of the 1950s when the first generation of TLCs was set-up in former Czechoslovakia. Since, similar units for microalgae culturing, which are relatively simple, low-cost and highly productive, have been installed in a number of other countries worldwide. The TLCs are characterized by microalgae growth at a low depth (< 50 mm) and fast flow (0.4-0.5 m/s) of culture compared to mixed ponds or raceways. It guarantees a high ratio of exposed surface to total culture volume (> 100 1/m) and rapid light/dark cycling frequencies of cells which result in high biomass productivity (> 30 g/m2/day) and operating at high biomass density, > 10 g/L of dry mass (DW). In TLCs, microalgae culture is grown in the system of inclined platforms that combine the advantages of open systems-direct sun irradiance, easy heat derivation, simple cleaning and maintenance, and efficient degassing-with positive features of closed systems-operation at high biomass densities achieving high volumetric productivity. Among significant advantages of thin layer cascades compared to raceway ponds are the operation at much higher cell densities, very high daylight productivities, and the possibility to store the culture in retention tanks at night, or in unfavourable weather conditions. Concerning the limitations of TLCs, one has to consider contaminations by other microalgae that limit cultivation to robust, fast-growing strains, or those cultured in selective environments.
The worldwide growing demand for energy permanently increases the pressure on industrial and scientific community to introduce new alternative biofuels on the global energy market. Besides the leading role of biodiesel and biogas, bioethanol receives more and more attention as first- and second-generation biofuel in the sustainable energy industry. Lately, microalgae (green algae and cyanobacteria) biomass has also remarkable potential as a feedstock for the third-generation biofuel production due to their high lipid and carbohydrate content. The third-generation bioethanol production technology can be divided into three major processing ways: (i) fermentation of pre-treated microalgae biomass, (ii) dark fermentation of reserved carbohydrates and (iii) direct "photo-fermentation" from carbon dioxide to bioethanol using light energy. All three technologies provide possible solutions, but from a practical point of view, traditional fermentation technology from microalgae biomass receives currently the most attention. This study mainly focusses on the latest advances in traditional fermentation processes including the steps of enhanced carbohydrate accumulation, biomass pre-treatment, starch and glycogen downstream processing and various fermentation approaches.
In this work we present a simple and cost-effective approach for the determination of selenium species in algae and yeast biomass, based on a combination of thin-layer chromatography (TLC) with diode laser thermal vaporization inductively coupled plasma mass spectrometry (DLTV ICP MS). Extraction of freeze-dried biomass was performed in 4M methanesulphonic acid and the selenium species were vaporized from cellulose TLC plates employing a continuous-wave infrared diode laser with power up to 4 W using a simple laboratory-built apparatus. Selenomethionine and selenocysteine were quantified with limits of detection 3 μg L-1 in a Se-enriched microalgae Chlorella vulgaris and yeast certified reference material SELM-1. Results delivered by TLC-DLTV ICP MS were consistent with those obtained by a routine coupling of high-performance liquid chromatography (HPLC) to ICP MS. In addition, the TLC approach is capable of analyzing extract containing even undiluted crude hydrolysates that could damage HPLC columns.
- MeSH
- chemické techniky analytické ekonomika metody MeSH
- Chlorella vulgaris chemie MeSH
- chromatografie na tenké vrstvě * MeSH
- hmotnostní spektrometrie * přístrojové vybavení MeSH
- lasery polovodičové MeSH
- Saccharomyces cerevisiae chemie MeSH
- selenocystein analýza MeSH
- selenomethionin analýza MeSH
- spektrální analýza MeSH
- volatilizace MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- Publikační typ
- časopisecké články MeSH
Quantification of selenated amino-acids currently relies on methods employing inductively coupled plasma mass spectrometry (ICP-MS). Although very accurate, these methods do not allow the simultaneous determination of standard amino-acids, hampering the comparison of the content of selenated versus non-selenated species such as methionine (Met) and selenomethionine (SeMet). This paper reports two approaches for the simultaneous quantification of Met and SeMet. In the first approach, standard enzymatic hydrolysis employing Protease XIV was applied for the preparation of samples. The second approach utilized methanesulfonic acid (MA) for the hydrolysis of samples, either in a reflux system or in a microwave oven, followed by derivatization with diethyl ethoxymethylenemalonate. The prepared samples were then analyzed by multiple reaction monitoring high performance liquid chromatography tandem mass spectrometry (MRM-HPLC-MS/MS). Both approaches provided platforms for the accurate determination of selenium/sulfur substitution rate in Met. Moreover the second approach also provided accurate simultaneous quantification of Met and SeMet with a low limit of detection, low limit of quantification and wide linearity range, comparable to the commonly used gas chromatography mass spectrometry (GC-MS) method or ICP-MS. The novel method was validated using certified reference material in conjunction with the GC-MS reference method.
- MeSH
- Chlorella vulgaris metabolismus MeSH
- limita detekce MeSH
- lineární modely MeSH
- malonáty MeSH
- methansulfonáty MeSH
- methionin analýza metabolismus MeSH
- pronasa MeSH
- reprodukovatelnost výsledků MeSH
- selenomethionin analýza metabolismus MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- Publikační typ
- časopisecké články MeSH
