Applied Microbiology and Biotechnology

, Volume 76, Issue 4, pp 783–794 | Cite as

Optimization of riboflavin production by recombinant Bacillus subtilis RH44 using statistical designs

  • Qiu-Li Wu
  • Tao Chen
  • Yu Gan
  • Xun Chen
  • Xue-Ming Zhao
Biotechnological Products and Process Engineering


A sequential optimization strategy, based on statistical experimental designs, was used to enhance the production of riboflavin by recombinant Bacillus subtilis RH44. In the first instance, the medium components were optimized in shake flask cultures. After preliminary experiments of nitrogen source selection, the two-level Plackett–Burman (PB) design was implemented to screen medium components that significantly influence riboflavin production. Among the 15 variables tested, glucose, NaNO3, K2HPO4, ZnSO4, and MnCl2 were identified as the most significant factors (confidence levels above 95%) for riboflavin production. The optimal values of these five variables were determined by response surface methodology (RSM) based on the central composite design (CCD). The validity of the model developed was verified, and the optimum medium led to a maximum riboflavin concentration of 6.65 g/l, which was 44.3 and 76.4% higher than the improved medium and the basal medium, respectively. A glucose-limited fed-batch culture profile in a 5-l fermentor was consequently designed according to the above optimum medium in shake flasks. A final riboflavin concentration of 16.36 g/l was obtained in 48 h, which further verified the practicability of this optimum strategy.


Bacillus subtilis Riboflavin production Plackett–Burman design Central composite design Response surface methodology 



This research was supported by the National Natural Science Foundation of China (NSFC-20536040), the National Project of Key Fundamental Research (2003Cb716003 and 2007CB707802), and the Development Project of Science and Technology of Tianjin (05YFGZGX04500). The authors sincerely thank Mr. A.O. Babatunde of University College Dublin, Ireland for his help in English editing.


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Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Qiu-Li Wu
    • 1
  • Tao Chen
    • 1
  • Yu Gan
    • 1
  • Xun Chen
    • 1
  • Xue-Ming Zhao
    • 1
  1. 1.Department of Biochemical Engineering, School of Chemical Engineering and TechnologyTianjin UniversityTianjinPeople’s Republic of China

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