Imobilização de células microbianas em esferas de alginato de cálcio e avaliação da viabilidade celular e estabilidade bioquímica em diferentes condições de armazenamento

Abstract

The preservation of microbial cultures aims to maintain the viability of the crop, without any morphological, physiological or genetic changes occur, providing cell stability. In this study the cellular immobilization technique was studied as a new proposal for the maintenance and preservation of microbial cultures. Escherichia coli and Enterobacter aerogenes cells were entrapped in calcium alginate beads, which were submitted to different storage conditions for 240 days. Moist spheres were maintained under refrigeration (5 ºC) conditions and deep freezing (-60 ºC) and lyophilized beads were kept under freezing (-18 ºC). The viability and biochemical stability of the immobilized microbial cells during the storage period were evaluated by the Petrifilm method and the automated microorganism identification system (VITEK 2), respectively. In addition, the morphology of the beads was evaluated by optical microscopy (OM) and scanning electron microscopy (SEM). The immobilization method employed achieved efficiency over 99% in spheres of uniform size with a mean diameter of 3.52 mm. The results of OM e SEM revealed spherical shape with a thick characteristic and small irregularities on the surface for the wet spheres. For the lyophilized spheres, the microscopy techniques revealed the spherical characteristic with wrinkled surface and well-defined grooves, being possible to observe cells bonded between the pores. E. aerogenes cells entrapped in calcium alginate beads kept moist under refrigeration and lyophilized under freezing showed viability and kept biochemical stability during 240 days of storage. The maintenance of the wet spheres under ultra-freezing allowed the viability of E. aerogenes cells for 150 days. The viability of E. coli cells was maintained during the entire period under all storage conditions studied. However, biochemical changes resulting in enzymatic inhibition of β-galactosidase and β-glucuronidase were observed throughout the 240 days of storage, and the storage of lyophilized beads under freezing preserved the characteristics of the cells for a greater period (150 days). The results obtained in this study suggest that the immobilization in lyophilized beads stores under freezing reach the best results of preservation of E. coli and E. aerogenes cells. These results indicate cellular immobilization in calcium alginate beads as a promising technique for the maintenance of E. coli and E. aerogenes cultures.