The influence of phosphorus source and the nature of nitrogen substrate on the biomass production and lipid accumulation in oleaginous Mucoromycota fungi

Abstract

Oleaginous filamentous fungi grown under the nitrogen limitation, accumulate high amounts of lipids in the form of triacylglycerides (TAGs) with fatty acid profiles similar to plant and fish oils. In this study, we investigate the effect of six phosphorus source concentrations combined with two types of nitrogen substrate (yeast extract and ammonium sulphate), on the biomass formation, lipid production, and fatty acid profile for nine oleaginous Mucoromycota fungi. The analysis of fatty acid profiles was performed by gas chromatography with flame ionization detector (GC-FID) and the lipid yield was estimated gravimetrically. Yeast extract could be used as both nitrogen and phosphorus source, without additional inorganic phosphorus supplementation. The use of inorganic nitrogen source (ammonium sulphate) requires strain-specific optimization of phosphorus source amount to obtain optimal lipid production regarding quantity and fatty acid profiles. Lipid production was decreased in ammonium sulphate-based media when phosphorus source was limited in all strains except for Rhizopus stolonifer. High phosphorus source concentration inhibited the growth of Mortierella fungi. The biomass (22 g/L) and lipid (14 g/L) yield of Umbelopsis vinacea was the highest among all the tested strains. Key points • The strain specific P requirements of Mucoromycota depend on the nature of N source. • Yeast extract leads to consistent biomass and lipid yield and fatty acids profiles. • Umbelopsis vinacea showed the highest biomass (22 g/L) and lipid (14 g/L) yield. • High P source amounts inhibit the growth of Mortierella fungi. Electronic supplementary material The online version of this article (10.1007/s00253-020-10821-7) contains supplementary material, which is available to authorized users.

Metal and Phosphate Ions Show Remarkable Influence on the Biomass Production and Lipid Accumulation in Oleaginous Mucor circinelloides

“…Only two observations could be considered as significant—increase of biomass under higher Zn ion availability and Ca deficiency. Moreover, under Pi conditions lower than the reference (Pi0.5 and Pi0.25), K and Na ions were compensated with KCl and NaCl salts in order to provide the same Na and K amounts as in the reference Pi1 condition [ 43 , 44 ]. It has been reported that chlorides could have negative effect on the mycelium formation of some fungi [ 74 , 75 ].…”

“…Spores were collected from agar plates using 10 ml of saline solution and a bacteriological loop. The composition of the reference medium, used and modified in our previous studies [ 27 , 43 , 44 , 45 ], was the following: 80 g/L of glucose, 1.5 g/L of (NH 4 ) 2 SO 4 , 7 g/L of KH 2 PO 4 , 2 g/L of Na 2 HPO 4 , 1.5 g/L of MgSO4·7H2O, 0.1 g/L of CaCl2·2H2O, 0.008 g/L of FeCl3·6H2O, 0.001 g/L of ZnSO4·7H2O, 0.0001 g/L of CoSO4·7H2O, 0.0001 g/L of CuSO4·5H2O, 0.0001 g/L of MnSO4·5H2O, where the listed concentrations of the metal ions Ca, Cu, Co, Fe, Mg, Mn, and Zn were assigned as reference concentration and marked as “R” ( Table 1 ). The higher—1000; 100; 10 × R and lower—0.1; 0.01 and 0 × R concentrations of the metal ions were assessed in the study as described in Table 1 .…”

“…Direct transesterification was performed according to Lewis et al [ 55 ], with some modifications [ 44 ]: 2 mL screw-cap polypropylene (PP) tubes were filled with 30 ± 3 mg of freeze-dried biomass, 250 ± 30 mg of acid-washed glass beads, and 500 μL of methanol. Further, the fungal biomass was disrupted in a tissue homogenizer (Bertin Technologies Percellys Evolution, Montigny-le-Bretonneux, France).…”

“…In our previous studies, we have assessed the chemical composition of nine different oleaginous filamentous fungi (including M. circinelloides ) and revealed nutrient-induced coproduction of lipids, chitin/chitosan, and polyphosphates [ 43 , 44 ]. We reported that M. circinelloides has an ability to coproduce lipids, polyphosphate, and chitin/chitosan at different phosphorus concentrations and showed a versatile metabolism with a high adaptability level to different stress conditions.…”

“…Different concentrations of phosphorus source were used in order to study the effect of metal ions on the co-production of lipids, polyphosphates and cell wall polysaccharides, such as chitin/chitosan, triggered by phosphorus availability. Analogous to our previous studies, the study was performed in a high-throughput set-up using a Duetz microtiter plate system (Duetz-MTPs) combined with Fourier transform infrared (FTIR) spectroscopy [ 43 , 44 , 45 , 46 ]. FTIR spectroscopy was applied to obtain a biochemical fingerprint of the microbial cells [ 43 , 47 , 48 , 49 , 50 , 51 , 52 ], while gas chromatography was used to analyze the lipid yield and fatty acid profiles of the extracted lipids.…”