The influence of the presence of ammonium ions in growth culture on malic enzyme (ME) isoforms activity and lipogenesis in Cunninghamella bainieri 2A1 was investigated. The fungus was cultivated in a nitrogen-limiting medium for 120 h at 30°C under two conditions. One of the cultures was intermittently fed with ammonium tartrate to maintain the ammonium concentrations above 0.5 g/L. The second culture was performed without any feeding to allow N limitation, thus promoting lipid accumulation. Activity staining of ME isoforms was carried out for both cultures. The culture which was not intermittently fed with ammonium tartrate achieved a maximum lipid content of 35% (g/g biomass) at 48 h. This culture possessed five ME isoforms (A, B, C, D and E) with isoform E showing a parallel correlation to lipid accumulation profile. In contrast, intensity of bands representing isoform D decreased as lipid accumulated. No appreciable differences of all other isoforms were observed. However, the culture which was intermittently fed with ammonium tartrate, accumulated only up to 16% lipid (g/g biomass). All isoforms were present but with a more pronounced activity of isoform D and a lower activity of isoform E was observed. These findings support further evidence that isoform E is the key isoform for lipid synthesis in C. bainieri 2A1.