Microbiological Production of Isocitric Acid from Biodiesel Waste and Its Effect on Spatial Memory.

Within this work, the microbial synthesis of (2R,3S)-isocitric acid (ICA), a metabolite of the nonconventional yeast Yarrowia lipolytica, from biodiesel waste, has been studied. The selected strain Y. lipolytica VKM Y-2373 synthesized ICA with citric acid (CA) as a byproduct. This process can be regulated by changing cultivation conditions. The maximal production of ICA with the minimal formation of the byproduct was provided by the use of a concentration of (NH4)2SO4 (6 g/L); the addition of biodiesel waste to cultivation medium in 20-60 g/L portions; maintaining the pH of the cultivation medium at 6, and degree of aeration between 25% and 60% of saturation. Itaconic acid at a concentration of 15 mM favorably influenced the production of ICA by the selected strain. The optimization of cultivation conditions allowed us to increase the concentration of ICA in the culture liquid from 58.32 to 90.2 g/L, the product yield (Y) by 40%, and the ICA/CA ratio from 1.1:1 to 3:1. Research on laboratory animals indicated that ICA counteracted the negative effect of ammonium molybdate (10-5 М) and lead diacetate (10-7 М) on the learning and spatial memory of rats, including those exposed to emotional stress.

Biosynthesis of isocitric acid in repeated-batch culture and testing of its stress-protective activity.

Biosynthesis of Ds(+)-threo-isocitric acid from ethanol in the Yarrowia lipolytica batch and repeated-batch cultures was studied. Repeated-batch cultivation was found to provide for a good biosynthetic efficiency of the producer for as long as 748 h, probably due to maintenance of high activities of enzymes involved in the biosynthesis of isocitric acid. Under optimal repeated-batch cultivation conditions, the producer accumulated 109.6 g/L Ds(+)-threo-isocitric acid with a production rate of 1.346 g/L h. The monopotassium salt of isocitric acid isolated from the culture liquid and purified to 99.9% was found to remove neurointoxication, to restore memory, and to improve the learning of laboratory rats intoxicated with lead and molybdenum salts. Taking into account the fact that the neurotoxic effect of heavy metals is mainly determined by oxidative stress, the aforementioned favorable action of isocitric acid on the intoxicated rats can be explained by its antioxidant activity among other pharmacological effects.