Optimization of Medium Composition for Cultivating Clostridium butyricum with Response Surface Methodology

ABSTRACT: Probiotic bacteria, such as some strains of Clostridium butyricum , have been frequently used as the active ingredient in functional foods. However, slow growth of the probiotic bacteria has been one of the big concerns for their potential commercial application. In the present study, a fractional factorial design was applied to investigate the main factors, namely, the concentrations of the glucose, tryptone, yeast extract, beef extract, cys-teine, (NH₄) ₂SO₄, NaCl, K₂HPO₄, and the medium initial pH that affected the growth of 1 probiotic strain, C. butyricum ZJUCB, currently preserved in our laboratory. Central composite experimental design and response surface methodology were adopted to derive a statistical model for optimizing the composition of the medium. The experimental results showed that the optimum medium for incubating the C. butyricum ZJUCB was composed of 2.44% (w/v) glucose, 2.08% yeast extract (w/v), 1% tryptone (w/v), 0.1% (NH₄)₂SO₄ (w/v), 0.1% NaHCO₃ (w/v), 0.02% MnSO₄. H₂O (w/v), 0.02% MgSO₄. 7H₂O (w/v), 0.002% CaCl₂ (w/v), and 2% agar (w/v) (if necessary) at pH 8.55. After incubation for 24 h in the optimum medium, the populations of the viable organisms could reach 10⁹ colony-forming units (CFU)/mL, which was 100 times higher than that incubated in the initial medium.