Burmistrova T.V., Kamanina O.A., Rybochkin P.V., Ponamoreva O.N.

Tula State University, Russia

Methylotrophic yeast Ogataea polymorpha BKM Y-2559 immobilized in polymer structures

In recent years, particular attention is directed to exploring possibilities sol-gel chemistry to obtain the immobilized microorganism in the silicate matrix. Silicon-based materials have the advantage over other polymers used for the immobilization of cells [1]. Encapsulated bacteria are of considerable interest for the development of effective biocatalysts, including the creation of biosensors [2].

The disadvantages of sol-gel method for the immobilization of microorganisms include excretion of lower alcohols during matrix formation, which are toxic for microorganisms. Because of this, we must introduce an additional step of removing alcohol from the reaction mixture. Low molecular weight alcohols are natural substrates for methylotrophic yeast, because of presence of an active oxidation system, which gives them significant biotechnological potential. Therefore we proposed to use these microorganisms to produce biohybrid materials using sol-gel process without an intermediate step of removing the alcohol.

Previous approaches for obtaining encapsulated in an organosilicate shell methylotrophic yeasts have been developed for potential methanol wastewater treatment and creation of microbial sensors. Hybrid materials were prepared on the basis of different ratios of methyltriethoxysilane (MTES), tetraethoxysilane (TEOS), and methylotrophic yeast cells.

Using scanning electron microscopy (SEM) formed structures were studied. Biosilicate architecture template derived from MTES and TEOS in a ratio of 85%: 15% is of greatest interest (Figure 1c). Each cell is formed around the capsule, wherein the encapsulated cells form a solid hybrid architecture biomaterial. Changing of the MTES content leads to inefficient encapsulation of yeast.

Figure 1. SEM image Ogataea polymorpha BKM Y-2559, encapsulated in a bimodal sol-gel matrices of different compositions: a – 10% on a hydrophobic additive; . b – 50 vol%; c – 85 vol%; d – 90 vol%).

An important feature of the consumer biocatalyst is its resistance to the harmful effects of various environmental factors. Since methylotrophic yeasts are encapsulated in sol-gel matrix, it is important to evaluate the effect of stressors on the activity of the biomaterial.

a biosensor system based on the oxygen electrode which surface is in direct contact with the encapsulated cells was used to study the degree of stress factors influence on the respiratory activity of the encapsulated methylotrophic yeast cells.

Wastewaters often contain heavy metal ions, which reduce the results of the determination of methanol. To identify the protective function of the capsules to the effects of heavy metal ions a comparative analysis of the methanol oxidize ability of the encapsulated yeast and microorganisms without capsule in the presence of heavy metal ions was carried out. The biosensor response values are almost independent of the presence of metal ions, except for Cu (II) and Cd (II) due to their high toxicity on all living organisms. Thus, the presence of heavy metal ions in the solution does not affect the respiratory activity of the encapsulated yeast, indicating that the capsules demonstrate their protective function.

Thus, the encapsulation of living cells in the methylotrophic yeast organosilicate sol-gel matrix provides the protection from the damaging effects of heavy metals and acidity.

Bibliography:

1. Sakai-Kato K., Ishikura K. Integration of biomolecules into analytical systems by means of silica sol-gel technology // National Institute of Health Science. ‒ 2005. ‒ V. 6. ‒ P. 70-75

2. Jia J., Tang M., Chen X., Qi L., Dong S. Co-immobilized microbial biosensor for BOD estimation based on sol–gel derived composite material // Biosensors and Bioelectronics. ‒ 2003. ‒ V. 18, I. 8. ‒ P. 1023-1029

The work on state orders in the field of scientific activities of the Russian Federation Ministry of Education (Task N 2014/227).