Shulyakovà M.A., Khomyak D.I., Mashchenko O.Y., Shevchuk T.A., Pirog T.P.

National University of Food Technologies, Kiev, Ukraine

BIOCONVERSION OF WASTE PRODUCTS OF BIODIESEL PRODUCTION IN SURFACE-ACTIVE SUBSTANCES OF RHODOCOCCUS ERYTHROPOLIS IMV Ac-5017, ACINETOBACTER CALCOACETICUS IMV B-7241 AND NOCARDIA VACCINII K-8

The wide use of fossil fuels leads to large emissions of greenhouse gases and causes irreversible damage to the environment. The current instability in oil supplies and continuous prices fluctuations are leading to a growing interest to alternative sources of energy. This situation needs to get the solution of economic, environmental and geopolitical issues and is central in the interest of renewable energy sources [1].

Biofuels, including ethanol and biodiesel, are promising substitutes of fossil fuels. Biodiesel is an environmentally friendly type of biofuel, derived from vegetable oils or animal fats and is supposed to replace petroleum. Nowadays etherification of vegetable oils is the most common method of producing biodiesel. Due to the increase in volumes of biodiesel production in the world a problem with utilization of its by-product - glycerol is occurring. For every 100 liters of biodiesel almost 10 liters of technical (crude) glycerol are produced (so-called glycerol fraction that settles after sedimentation) [2]. Glycerol fraction except its main product (60-80% of glycerol) also contains a large number of different impurities that disables its use in many traditional areas of application of glycerol (food processing, pharmaceutical and cosmetic industry, etc.) because of increased alkalinity and high concentrations of methanol. Storage and utilization of glycerol fraction is a serious environmental problem and its treatment is extremely expensive.

Thus, to improve the economic reasonability and profitability of biodiesel production development of new alternative methods of disposal of this waste are required. As possible ways of its disposal incineration, composting, and thermochemical conversion could be considered [1] but he alternative way of glycerol utilization is use of it as a substrate in biotechnological processes for producing of practically valuable products, including surface-active substances (SAS) [2, 3]. Moreover, from the microbiological point of view now the main task is to get the bacteria resistant to inhibitors that are present in crude glycerol, and special attention there should be given to the remains of methanol and sodium or potassium salts, as it is known that they are able to inhibit cell growth [2].

In previous studies we showed the possibility of use of refined glycerol (98%) as a source of carbon and energy for the SAS synthesis by Rhodococcus erythropolis IMV Ac-5017, Acinetobacter calcoaceticus IMV B-7241 and Nocardia vaccinii K-8. But now it was quite necessary to explore the possibility of bioconversion of crude glycerol into biosurfactants by these strains. The average composition of glycerol fraction was simulated by the addition of residual alcohols (methanol or ethanol) and sodium or potassium chlorides to the medium. Thereafter, a substrate was called the "crude" glycerol.

Quantitative content of surfactants in the culture broth was estimated with indicator of SAS (SAS*), so called the "relative concentration of surfactant," as described in work [4].

It was discovered that the addition of KCl or NaCl in concentration of 2,5 % to the medium led to the increase in SAS synthesis (by 4-35 %) by all strains. The presence of these salts, even in concentration of 5-10 %, didn’t cause a significant inhibition on the processes of SAS formation. This may be explained by stimulating effect of cations of metals on the activity of enzymes of anaplerotic reactions and SAS biosynthesis of studied strains. For example in previous works it was shown that during the growth of A. calcoaceticus IMV B-7241 on ethanol Na⁺ acted as the activator of phosphoenolpiruvate (PEP)-carboxylase – in the presence of 100 mM of Na⁺ in the reaction mixture activity of the enzyme increased in 1,2-1,3 fold [5]. The physiological significance of this enzyme during the cultivation of the strain IMV B-7241 on ethanol consists in activation of gluconeogenesis, and, hence, the increase of the synthesis of glycolipid biosurfactants.

Thus, the substitution of potassium nitrate by the equimolar concentration of sodium nitrate in the medium with hexadecane was accompanied by the increase in SAS synthesis of R. erythropolis IMV Ac-5017 in 1,5-2 fold [5]. This effect may be explained by the fact that sodium is the activator of the hexadecane-oxidazing enzyme of strain IMV Ac-5017. It was also shown the feasibility of use of sodium nitrate as nitrogen source during the growth of N. vaccinii K-8 on glycerol [6].

The presence of salts and 0,3 % of methanol or ethanol in the medium with glycerol didn’t inhibit the bacteria growth and even was accompanied by increase in SAS synthesis to 11-77 % compared with cultivation on the medium without salts and alcohols. Under such conditions alcohols may be considered as secondary sources of carbon and was consumed by cells that can be explained by previously established broad substrate specificity of N,N-dimethylnitrosamine (NDMA)-dependent alcohol dehydrogenases of R. erythropolis IMV Ac-5017 and A. calcoaceticus IMV B-7241 [7].

Thus, the proposed method of utilization of crude glycerol allows to improve the profitability of biodiesel production by bioconversion of its by-product into practically valuable microbial surfactants.

References

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