Lignocellulose is an abundant raw material and renewable carbon source for the production of single cell oils which can replace plant-derived oils in food, feed, fuels, and oleochemicals. Mucor circinelloides produces both fatty acids and amino polysaccharides, such as chitin and chitosan. This study evaluates hydrolysates of Norway spruce (Picea abies) as a carbon source for their simultaneous production. Cultivation in spruce hydrolysate media yielded 15.8 g/L of biomass, with fatty acids comprising ~ 50% of the cell dry weight and amino polysaccharides up to 8.5%. The fatty acid methyl ester (FAME) content and fatty acid profile were comparable to glucose fermentation. Optimal harvesting times ranged from 72 to 120 h, depending on desired yields. These findings demonstrate that Norway spruce hydrolysates are a viable and sustainable substrate for microbial lipid and polysaccharide production, supporting their potential use in biotechnology and industrial applications.

• Klíčová slova
• Mucor circinelloides, Amino polysaccharides, Fatty acids, Fermentation, Lignocellulose,
• MeSH
• biomasa MeSH
• fermentace MeSH
• hydrolýza MeSH
• mastné kyseliny * biosyntéza   MeSH
• Mucor * metabolismus   růst a vývoj   MeSH
• polysacharidy * biosyntéza   MeSH
• smrk * metabolismus   chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• mastné kyseliny * MeSH
• polysacharidy * MeSH

Nitrogen is one of the most important nutrient sources for the growth of microalgae. We studied the effects of nitrogen starvation on the growth responses, biochemical composition, and fatty acid profile of Dunaliella tertiolecta, Phaeodactylum tricornutum, and Nannochloropsis oculata. The lack of nitrogen caused changes in carbohydrate, protein, lipid, and fatty acid composition in all examined microalgae. The carbohydrate content increased 59% in D. tertiolecta, while the lipid level increased 139% in P. tricornutum under nitrogen stress conditions compared to the control groups. Nitrogen starvation increased the oligosaccharide and polysaccharide contents of D. tertiolecta 4.1-fold and 3.6-fold, respectively. Furthermore, triacylglycerol (TAG) levels in N. oculata and P. tricornutum increased 2.3-fold and 7.4-fold, respectively. The dramatic increase in the amount of TAG is important for the use of these microalgae as raw materials in biodiesel. Nitrogen starvation increased the amounts of oligosaccharides and polysaccharides of D. tertiolecta, while increased eicosapentaenoic acid (EPA) in N. oculata and docosahexaenoic acid (DHA) content in P. tricornutum. The amount of polyunsaturated fatty acids (PUFAs), EPA, DHA, oligosaccharides, and polysaccharides in microalgal species can be increased without using the too costly nitrogen source in the culture conditions, which can reduce the most costly of living feeding.

• Klíčová slova
• Biochemical composition, Fatty acid profile, Microalgae, Nitrogen starvation,
• MeSH
• dusík * metabolismus   MeSH
• mastné kyseliny * metabolismus   analýza   MeSH
• mikrořasy * metabolismus   růst a vývoj   MeSH
• triglyceridy metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• dusík * MeSH
• mastné kyseliny * MeSH
• triglyceridy MeSH

BACKGROUND: Monitoring and control of both growth media and microbial biomass is extremely important for the development of economical bioprocesses. Unfortunately, process monitoring is still dependent on a limited number of standard parameters (pH, temperature, gasses etc.), while the critical process parameters, such as biomass, product and substrate concentrations, are rarely assessable in-line. Bioprocess optimization and monitoring will greatly benefit from advanced spectroscopy-based sensors that enable real-time monitoring and control. Here, Fourier transform (FT) Raman spectroscopy measurement via flow cell in a recirculatory loop, in combination with predictive data modeling, was assessed as a fast, low-cost, and highly sensitive process analytical technology (PAT) system for online monitoring of critical process parameters. To show the general applicability of the method, submerged fermentation was monitored using two different oleaginous and carotenogenic microorganisms grown on two different carbon substrates: glucose fermentation by yeast Rhodotorula toruloides and glycerol fermentation by marine thraustochytrid Schizochytrium sp. Additionally, the online FT-Raman spectroscopy approach was compared with two at-line spectroscopic methods, namely FT-Raman and FT-infrared spectroscopies in high throughput screening (HTS) setups. RESULTS: The system can provide real-time concentration data on carbon substrate (glucose and glycerol) utilization, and production of biomass, carotenoid pigments, and lipids (triglycerides and free fatty acids). Robust multivariate regression models were developed and showed high level of correlation between the online FT-Raman spectral data and reference measurements, with coefficients of determination (R2) in the 0.94-0.99 and 0.89-0.99 range for all concentration parameters of Rhodotorula and Schizochytrium fermentation, respectively. The online FT-Raman spectroscopy approach was superior to the at-line methods since the obtained information was more comprehensive, timely and provided more precise concentration profiles. CONCLUSIONS: The FT-Raman spectroscopy system with a flow measurement cell in a recirculatory loop, in combination with prediction models, can simultaneously provide real-time concentration data on carbon substrate utilization, and production of biomass, carotenoid pigments, and lipids. This data enables monitoring of dynamic behaviour of oleaginous and carotenogenic microorganisms, and thus can provide critical process parameters for process optimization and control. Overall, this study demonstrated the feasibility of using FT-Raman spectroscopy for online monitoring of fermentation processes.

• Klíčová slova
• Carotenoids, Infrared spectroscopy, Lipids, Partial least squares (PLS) regression, Process analytical technology, Raman spectroscopy, Real-time monitoring, Rhodotorula, Schizochytrium,
• MeSH
• biomasa MeSH
• fermentace MeSH
• glukosa metabolismus   MeSH
• glycerol MeSH
• karotenoidy metabolismus   MeSH
• Ramanova spektroskopie * metody   MeSH
• triglyceridy MeSH
• uhlík * metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• glukosa MeSH
• glycerol MeSH
• karotenoidy MeSH
• triglyceridy MeSH
• uhlík * MeSH

Oleaginous filamentous fungi can accumulate large amount of cellular lipids and biopolymers and pigments and potentially serve as a major source of biochemicals for food, feed, chemical, pharmaceutical, and transport industries. We assessed suitability of Fourier transform (FT) Raman spectroscopy for screening and process monitoring of filamentous fungi in biotechnology. Six Mucoromycota strains were cultivated in microbioreactors under six growth conditions (three phosphate concentrations in the presence and absence of calcium). FT-Raman and FT-infrared (FTIR) spectroscopic data was assessed in respect to reference analyses of lipids, phosphorus, and carotenoids by using principal component analysis (PCA), multiblock or consensus PCA, partial least square regression (PLSR), and analysis of spectral variation due to different design factors by an ANOVA model. All main chemical biomass constituents were detected by FT-Raman spectroscopy, including lipids, proteins, cell wall carbohydrates, and polyphosphates, and carotenoids. FT-Raman spectra clearly show the effect of growth conditions on fungal biomass. PLSR models with high coefficients of determination (0.83-0.94) and low error (approximately 8%) for quantitative determination of total lipids, phosphates, and carotenoids were established. FT-Raman spectroscopy showed great potential for chemical analysis of biomass of oleaginous filamentous fungi. The study demonstrates that FT-Raman and FTIR spectroscopies provide complementary information on main fungal biomass constituents.

• Klíčová slova
• biodiesel, biopolymers, carotenoids, chitin, chitosan, fatty acids, fermentation, fungi, oleaginous microorganisms, pigments,
• MeSH
• analýza hlavních komponent MeSH
• biologické pigmenty analýza   MeSH
• biomasa MeSH
• biotechnologie MeSH
• chromatografie plynová MeSH
• fosfor analýza   metabolismus   MeSH
• Fourierova analýza MeSH
• houby chemie   růst a vývoj   MeSH
• karotenoidy analýza   MeSH
• lipidy analýza   MeSH
• magnetická rezonanční spektroskopie MeSH
• Ramanova spektroskopie metody   MeSH
• spektrofotometrie ultrafialová MeSH
• spektroskopie infračervená s Fourierovou transformací MeSH
• vápník metabolismus   MeSH
• vysokoúčinná kapalinová chromatografie MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• biologické pigmenty MeSH
• fosfor MeSH
• karotenoidy MeSH
• lipidy MeSH
• vápník MeSH

The co-cultivation of red yeasts and microalgae works with the idea of the natural transport of gases. The microalgae produce oxygen, which stimulates yeast growth, while CO2 produced by yeast is beneficial for algae growth. Both microorganisms can then produce lipids. The present pilot study aimed to evaluate the ability of selected microalgae and carotenogenic yeast strains to grow and metabolize in co-culture. The effect of media composition on growth and metabolic activity of red yeast strains was assessed simultaneously with microalgae mixotrophy. Cultivation was transferred from small-scale co-cultivation in Erlenmeyer flasks to aerated bottles with different inoculation ratios and, finally, to a 3L bioreactor. Among red yeasts, the strain R. kratochvilovae CCY 20-2-26 was selected because of the highest biomass production on BBM medium. Glycerol is a more suitable carbon source in the BBM medium and urea was proposed as a compromise. From the tested microalgae, Desmodesmus sp. were found as the most suitable for co-cultivations with R. kratochvilovae. In all co-cultures, linear biomass growth was found (144 h), and the yield was in the range of 8.78-11.12 g/L of dry biomass. Lipids increased to a final value of 29.62-31.61%. The FA profile was quite stable with the UFA portion at about 80%. Around 1.98-2.49 mg/g CDW of carotenoids with torularhodine as the major pigment were produced, ubiquinone production reached 5.41-6.09 mg/g, and ergosterol yield was 6.69 mg/g. Chlorophyll production was very low at 2.11 mg/g. Pilot experiments have confirmed that carotenogenic yeasts and microalgae are capable of symbiotic co-existence with a positive impact om biomass growth and lipid metabolites yields.

• Klíčová slova
• Desmodesmus sp, Rhodotorula kratochvilovae, carotenogenic yeasts, carotenoids, co-cultivation, lipids, microalgae,
• Publikační typ
• časopisecké články MeSH

Beta (β)-glucans are polysaccharides composed of D-glucose monomers. Nowadays, β-glucans are gaining attention due to their attractive immunomodulatory biological activities, which can be utilized in pharmaceutical or food supplementation industries. Some carotenogenic Basidiomycetes yeasts, previously explored for lipid and carotenoid coproduction, could potentially coproduce a significant amount of β-glucans. In the present study, we screened eleven Basidiomycetes for the coproduction of lipids and β-glucans. We examined the effect of four different C/N ratios and eight different osmolarity conditions on the coproduction of lipids and β-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 strains C. infirmominiatum CCY 17-18-4 and R. kratochvilovae CCY 20-2-26 were identified as the best coproducers of lipids and β-glucans. In addition, C. infirmominiatum CCY 17-18-4, R. kratochvilovae CCY 20-2-26 and P. rhodozyma CCY 77-1-1 were identified as the best alternative producers of β-glucans. Increased C/N ratio led to increased biomass, lipid and β-glucans production for several yeast strains. Increased osmolarity had a negative effect on biomass and lipid production while the β-glucan production was positively affected.

• Klíčová slova
• carbon:nitrogen ratio, high-throughput screening, lipids, osmotic stress, red yeast, β-glucans,
• Publikační typ
• časopisecké články MeSH

Carotenogenic yeasts are non-conventional oleaginous microorganisms capable of utilizing various waste substrates. In this work, four red yeast strains (Rhodotorula, Cystofilobasidium, and Sporobolomyces sp.) were cultivated in media containing crude, emulsified, and enzymatically hydrolyzed animal waste fat, compared with glucose and glycerol, as single C-sources. Cell morphology (cryo-SEM (cryo-scanning electron microscopy), TEM (transmission electron microscopy)), production of biomass, lipase, biosurfactants, lipids (gas chromatography/flame ionization detection, GC/FID) carotenoids, ubiquinone, and ergosterol (high performance liquid chromatography, HPLC/PDA) in yeast cells was studied depending on the medium composition, the C source, and the carbon/nitrogen (C/N) ratio. All studied strains are able to utilize solid and processed fat. Biomass production at C/N = 13 was higher on emulsified/hydrolyzed fat than on glucose/glycerol. The 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 (cell dry weight)) were found in S. pararoseus. Simultaneous induction of lipase and biosurfactants was observed on crude fat substrate. An increased C/N ratio (13-100) led to higher biomass production in fat media. The production of total lipids increased in all strains to C/N = 50. Oppositely, the production of carotenoids, ubiquinone, and ergosterol dramatically decreased with increased C/N in all strains. Compounds accumulated in stressed red yeasts have a great application potential and can be produced efficiently during the valorization of animal waste fat under the biorefinery concept.

• Klíčová slova
• biosurfactants, carotenogenic yeasts, carotenoids, ergosterol, lipase, lipids, ubiquinone,
• Publikační typ
• časopisecké články MeSH