Oleaginous yeasts are considered as a potential lipid source for food, feed and biofuel production. In order to make the yeast-based lipid production environmentally and economically sustainable, there is a need for screening studies in order to find the best yeast lipid producers on different substrates, and to optimize cultivation conditions. Since the target parameter of such screening studies are lipid amounts and profiles, an analytical technique that is able to perform lipid analyses rapidly, reproducible and with high precision is highly desirable. The main objective of this study was to establish the non-invasive high-throughput Fourier transform infrared (FTIR) spectroscopy analysis for the prediction of lipid content and profile in oleaginous yeasts. High-throughput FTIR spectroscopy allowed characterizing the total biochemical profile of oleaginous yeasts and enabled us to identify strains and substrate(s) providing the highest total lipid content. Some of the yeast strains grown under nitrogen-limiting conditions with glucose/xylose/mixture of glucose and xylose as carbon sources were accumulating lipids with a high proportion of free fatty acids. FTIR spectra were used to predict gravimetric and gas chromatography data by establishing multivariate calibration models. Coefficients of determination (R2) for calibration models were obtained in a range between 0.62 and 0.92 for predicting lipid content. When using an independent test set, R2 values between 0.53 and 0.79 were achieved for predicting fatty acid profile. The best spectral region(s) for the prediction of total lipid content was 3100–2800 cm−1 combined with 1800–700 cm−1, and for prediction of summed saturated (SAT), monounsaturated (MUFA) and polyunsaturated (PUFA) fatty acids: 3100–2800 cm−1, 3100–2800 cm−1 combined with 1700–1715 cm−1 and 3100–2800 cm−1 combined with 1800–1715 cm−1, respectively. The highest lipid accumulation was observed for strains Rhodotorula babjevae DBVPG 8058 on glucose and mixture of glucose and xylose and Lipomyces starkeyi CBS 2512 on xylose. Applying FTIR spectroscopy combined with multivariate data analysis allows performing rapid, non-invasive, reproducible and precise quantitative predictions of total lipid content and lipid profile. It allows also detecting different lipid fractions as triacylglycerols (TAGs) and free fatty acids and evaluating the total biochemical profile of cells. Several yeast strains with high lipid accumulation were identified.

Oleaginous fungi can accumulate lipids by utilizing a wide range of waste substrates. They are an important source for the industrial production of omega-6 polyunsaturated fatty acids (gamma-linolenic and arachidonic acid) and have been suggested as an alternative route for biodiesel production. Initial research steps for various applications include the screening of fungi in order to find efficient fungal producers with desired fatty acid composition. Traditional cultivation methods (shake flask) and lipid analysis (extraction-gas chromatography) are not applicable for large-scale screening due to their low throughput and time-consuming analysis. Here we present a microcultivation system combined with high-throughput Fourier transform infrared (FTIR) spectroscopy for efficient screening of oleaginous fungi. The microcultivation system enables highly reproducible fungal fermentations throughout 12 days of cultivation. Reproducibility was validated by FTIR and HPLC data. Analysis of FTIR spectral ester carbonyl peaks of fungal biomass offered a reliable high-throughput at-line method to monitor lipid accumulation. Partial least square regression between gas chromatography fatty acid data and corresponding FTIR spectral data was used to set up calibration models for the prediction of saturated fatty acids, monounsaturated fatty acids, polyunsaturated fatty acids, unsaturation index, total lipid content and main individual fatty acids. High coefficients of determination (R2 = 0.86–0.96) and satisfactory residual predictive deviation of cross-validation (RPDCV = 2.6–5.1) values demonstrated the goodness of these models. We have demonstrated in this study, that the presented microcultivation system combined with rapid, high-throughput FTIR spectroscopy is a suitable screening platform for oleaginous fungi. Sample preparation for FTIR measurements can be automated to further increase throughput of the system.

The relative abundance of specific fatty acids in milk can be important for consumer health and manufacturing properties of dairy products. Understanding of genes controlling milk fat synthesis may contribute to the development of dairy products with high quality and nutritional value. This study aims to identify key genes and genetic variants affecting de novo synthesis of the short- and medium-chained fatty acids C4:0 to C14:0. A genome-wide association study using 609,361 SNP markers and 1,811 animals was performed to detect genomic regions affecting fatty acid levels. These regions were further refined using sequencing data to impute millions of additional genetic variants. Results suggest associations of PAEP with the content of C4:0, AACS with the content of fatty acids C4:0-C6:0, NCOA6 or ACSS2 with the longer chain fatty acids C6:0-C14:0, and FASN mainly associated with content of C14:0. None of the top-ranking markers caused amino acid shifts but were mostly situated in putatively regulating regions and suggested a regulatory role of the QTLs. Sequencing mRNA from bovine milk confirmed the expression of all candidate genes which, combined with knowledge of their roles in fat biosynthesis, supports their potential role in de novo synthesis of bovine milk fatty acids.

The biomass of Mucor circinelloides, a dimorphic oleaginous filamentous fungus, has a significant nutritional value and can be used for single cell oil production. Metal ions are micronutrients supporting fungal growth and metabolic activity of cellular processes. We investigated the effect of 140 different substrates, with varying amounts of metal and phosphate ions concentration, on the growth, cell chemistry, lipid accumulation, and lipid profile of M. circinelloides. A high-throughput set-up consisting of a Duetz microcultivation system coupled to Fourier transform infrared spectroscopy was utilized. Lipids were extracted by a modified Lewis method and analyzed using gas chromatography. It was observed that Mg and Zn ions were essential for the growth and metabolic activity of M. circinelloides. An increase in Fe ion concentration inhibited fungal growth, while higher concentrations of Cu, Co, and Zn ions enhanced the growth and lipid accumulation. Lack of Ca and Cu ions, as well as higher amounts of Zn and Mn ions, enhanced lipid accumulation in M. circinelloides. Generally, the fatty acid profile of M. circinelloides lipids was quite consistent, irrespective of media composition. Increasing the amount of Ca ions enhanced polyphosphates accumulation, while lack of it showed fall in polyphosphate.

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.