Abstract
Importance of bacterial alginate is attributed to its employment in multiple fields like pharmaceutical, biomedical and food industries. This research was aiming to discover a new Egyptian isolate of Azotobacter chroococcum and thereafter stimulation of production and extraction of alginate under various stress conditions. Extracted alginate then entered in a process of composition of hybrid inorganic composite based on an apatite and a polysaccharide of microbial origin. Isolates of Azotobacter chroococcum bacteria were isolated from different habitats in Egypt to assess the highest alginate productivity. Mannitol, sucrose, ethanol, tryptone, beef extract, peptone, yeast extract, pH, incubation time and agitation were tested for enhancement of alginate productivity. Biochemical tests and molecular study were done for the isolated Azotobacter and the new isolate was registered in GenBank with accession no MH179061. In vitro bone bioactivity was carried out via soaking the prepared materials in simulated body fluid (SBF) for 7 and 21 days in presence of hydroxyapatite nanoparticles (HA) and polymethyl methacrylate (PMMA) in different ratios. The examination of apatite formation on the prepared materials after soaking in SBF was carried out by fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy with energy dispersive X-ray (SEM-EDX) and (3-(4,5-Dimethylthiazol)-2,5-Diphenyltetrazolium bromide) (MTT) assay was used to assess the cytotoxic effect of sodium alginate/hydroxyapatite analogs against normal splenocyte. The new strain was identified as Azotobacter chroococcum MH179061 which has the ability to maximize alginate production (3.8- 4 g/L) in bacterial medium. Extracted sodium alginate has the ability of formation of a thick apatite layer on the surface of the composite films in special way on this containing SA/PMMA with ratio (1.5:1 and 1:1) compared to other ones. Significant increase in splenocyte proliferation noticed the nontoxic effects of the SA/HA particles, moreover, the cell division was stimulated; cells were able to maintain their healthy and being even in the presence of a different concentration of HA particles more than control. The novel composites based on sodium alginate produced from Azotobacter chroococcum MH179061 has promising bioactivity properties, which can be applied in bone implants and tissue engineering applications after further investigations.