Bioprocessing development for anaerobic cultivation of probiotic bacteria bifidobacterium longum for high cell mass

Abstract

Bifidobacteria are used as probiotic mainly in the dairy industry as cell suspensions or as freeze-dried additives. Bifidobacterium longum is important in maintaining general health. Anaerobic growth of B. longum and its organic acids byproduct give some restriction to their growth. Therefore, the goal of this research is to select suitable optimized medium as production media of B. longum and its growth characteristics on this medium. Several media were tested for the potential effect to the cell growth. The best medium gave the cell mass of 2.58 g L-1. In shake flasks, further optimization by using classical and statistical method gave the cell mass of 6.03 g L-1 and 5.95 g L-1, respectively. The medium optimized using classical method was selected and used in the bioreactor. Cultivation in bioreactor was carried out with controlled and un-controlled pH. High cell mass was observed in controlled pH with bioreactor which yielded 11.97 g L-1, compared with uncontrolled pH which gave only 7.5 g L-1. Fed-batch cultivation was carried out by constant supplementation of glucose or complete media. Feeding with complete medium gave the highest cell mass comparing with glucose feeding, the cells obtained were 18.5 g L-1 and 17.7 g L-1, respectively. Study on its growth characteristics revealed that B. longum has good resistance to bile salt concentration with 79 % cell survival at concentration of 0.3 % of bile salts. However, the resistance to bile salt was found to be affected by pH value. Following two-hour treatment, no cells survived at pH 1.0 while about 41.7 % of cells survived at pH 4. Surface adherence of the cells was tested by using hydrophobicity assays. High degree of hydrophobicity was observed during exposure to chloroform which gave the value of 86.1 %. Susceptibility of the cells was also tested with antimicrobial and antibiotics. B. longum was found resistant to all the pathogenic microorganism tested and susceptible to rifampicin, gentamycin, erythromycin, and ampicillin.