O on lipid and GLA production by Cunninghamella sp. 2A1 were studied in submerged culture fermentation. Cultivation was performed in 250 ml conical flask (100ml working volume), at 30°C and 250 rpm agitation for 120h. Mg 2+ , Fe 2+ and Zn 2+ have significant effect on lipid accumulation as inclusion of each ions resulted in a 64, 43 and 33% increase in lipid content compared to cultures which was cultivated with the omission of each ions, respectively. All three metal ions also affected GLA production by Cunninghamella sp. 2A1 with Zn 2+ having the most profound effect with an increase of up to 74% of GLA yield (g GLA/g lipidless biomass) achieved. As omission of any single metal ions did not affect biomass concentration achieved, we concluded that metal ions components in production media may have affected lipid and GLA production by affecting pathways involved in the biosynthesis of lipid and GLA. This study reports the critical concentration of each metal ion for optimal lipid and GLA production by Cunninghamella sp. 2A1.

Strategic feeding of ammonium and metal ions (Mg2+, Mn2+, Fe3+, Cu2+, Ca2+, Co2+, and Zn2+) for enhanced GLA-rich lipid accumulation in C. bainieri 2A1 was established. When cultivated in nitrogen-limited medium, the fungus produced up to 30% lipid (g/g biomass) with 12.9% (g/g lipid) GLA. However, the accumulation of lipid stopped at 48 hours of cultivation although glucose was abundant. This event occurred in parallel to the diminishing activity of malic enzyme (ME), fatty acid synthase (FAS), and ATP citrate lyase (ACL) as well as the depletion of metal ions in the medium. Reinstatement of the enzymes activities was achieved by feeding of ammonium tartrate, but no increment in the lipid content was observed. However, increment in lipid content from 32% to 50% (g/g biomass) with 13.2% GLA was achieved when simultaneous feeding of ammonium, glucose, and metal ions was carried out. This showed that the cessation of lipid accumulation was caused by diminishing activities of the enzymes as well as depletion of the metal ions in the medium. Therefore, strategic feeding of ammonium and metal ions successfully reinstated enzymes activities and enhanced GLA-rich lipid accumulation in C. bainieri 2A1.