Crypthecodinium cohnii is a marine heterotrophic dinoflagellate that can accumulate high amounts of omega-3 polyunsaturated fatty acids (PUFAs), and thus has the potential to replace conventional PUFAs production with eco-friendlier technology. So far, C. cohnii cultivation has been mainly carried out with the use of yeast extract (YE) as a nitrogen source. In the present study, alternative carbon and nitrogen sources were studied: the extraction ethanol (EE), remaining after lipid extraction, as a carbon source, and dinoflagellate extract (DE) from recycled algae biomass C. cohnii as a source of carbon, nitrogen, and vitamins. In mediums with glucose and DE, the highest specific biomass growth rate reached a maximum of 1.012 h−1, while the biomass yield from substrate reached 0.601 g·g−1. EE as the carbon source, in comparison to pure ethanol, showed good results in terms of stimulating the biomass growth rate (an 18.5% increase in specific biomass growth rate was observed). DE supplement to the EE-based mediums promoted both the biomass growth (the specific growth rate reached 0.701 h−1) and yield from the substrate (0.234 g·g−1). The FTIR spectroscopy data showed that mediums supplemented with EE or DE promoted the accumulation of PUFAs/docosahexaenoic acid (DHA), when compared to mediums containing glucose and commercial YE.

The Kluyveromyces marxianus yeast recently has gained considerable attention due to its applicability in high-value-added product manufacturing. In order to intensify the biosynthesis rate of a target product, reaching high biomass concentrations in the reaction medium is mandatory. Fed-batch processes are an attractive and efficient way how to achieve high cell densities. However, depending on the physiology of the particular microbial strain, an optimal media composition should be used to avoid by-product synthesis and, subsequently, a decrease in overall process effi-ciency. Thus, the aim of the present study was to optimise the synthetic growth medium and feeding solution compositions (in terms of carbon, nitrogen, phosphorous, magnesium, and calcium concentrations) for high cell density K. marxianus fed‑batch cultivations. Additionally, the biomass yields from the vitamin mixture and other macro/microelements were identified. A model predictive control algorithm was successfully applied for a fed-batch cultivation control. Biomass growth and substrate consumption kinetics were compared with the mathematical model predictions. Finally, 2‑phenylethanol biosynthesis was induced and its productivity was estimated. The determined optimal macronutrient ratio for K. marxianus biomass growth was identified as C:N:P = 1:0.07:0.011. The maximal attained yeast biomass concentration was close to 70 g·L-1 and the 2-PE biosynthesis rate was 0.372 g·L−1·h−1, with a yield of 74% from 2-phenylalanine.

Methanotrophs display the ability to consume methane as a carbon source and produce a wide-range of high-value products, e.g. ectoine/hydroxyectoine, poly-b-hydroxybutyrate (PHB), single cell protein, extracellular polysaccharides and lipids. Usually methanotrophs show low specific substrate consumption rates, which restricts their application at pilot and industrial scale. Thus, in order to reduce the time and costs of the cultivation process, it is vital to accelerate the growth of applied organisms. Usually, methanotrophic bacteria cultivations are carried out using fully synthetic mineral mediums (nitrate mineral salts medium (NMS)) without the addition of any growth factors. Potentially, higher biomass growth and substrate uptake rates can be achieved by supplementing the growth medium with vitamins, amino acids etc. or by using more bioavailable substrates.The aim of our research was to study the influence of growth factors such as vitamins, and different nitrogen sources (yeast extract, yeast nitrogen base with/without amino acids and tryptone) on the growth of Methylomonas methanica, Methylomicrobium alcaliphilum and Methylosinus trishosporium.Experiments for studying the influence of growth factors were carried out in shake flasks by varying the medium compositions and analyzing the effects of said variations on the kinetics of the cultivation, e.g. specific biomass growth rate and biomass yield from substrate.Subsequent tests of the developed nutrient medium, which promotes higher biomass growth rates, were carried out in laboratory 5 L bioreactor Methylosinus trishosporium cultivations to study the main process parameters.From the statistical analysis of experimental data it was observed, that supplementation of the growth medium with yeast extract or tryptone, seems to promote the growth rate of methanotrophs, when methanol is used as the main substrate. Furthermore, specific growth rates observed during cultivations in mediums containing vitamins (including cobolamin) also seem to positively affect the biomass growth rate. Based on the results of lab-scale bioreactor cultivations, using the identified medium composition it was possible to achieve a maximal biomass specific growth rate of 0.15 L⸱h‑1 and productivity of 0.16 g⸱L-1⸱h-1.

Soy legHemoglobin is one of the most important and key ingredients in plant based meat substitutes that can imitate the colour and flavour of the meat. In order to improve the high yield production of legHemoglobin protein and its main component heme in the yeast Pichia pastoris, glycerol and methanol cultivation conditions were studied. Additionally, in silico metabolic modelling analysis of growth coupled enzyme quantity, suggests metabolic gene up/down regulation strategies for heme production. First, cultivations and metabolic modelling analysis of P. pastoris were performed on glycerol and methanol in different growth media. Glycerol cultivation uptake and production rates can be increased by 50 % according to metabolic modelling results, but methanol cultivation is near the theoretical maximum. Growth coupled metabolic optimisation results revealed the best feasible upregulation (33 reactions) (1.47 % of total reactions) and 67 downregulation/deletion (2.98 % of total) reaction suggestions. Finally, we describe reaction regulation suggestions with the highest potential to increase heme production yields.