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Enhanced amylase production by Bacillus subtilis using a dual exponential feeding strategy

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Abstract

A recombinant Bacillus subtilis strain (ATCC 31784) haboring the plasmid pC194 with a thermostable α-amylase gene was cultured in a 22-l B. Braun Biostat C fermenter. Traditional batch operations suffer from low cell mass and protein productions because a high initial glucose concentration causes substrate inhibition and also product inhibition due to acetate accumulation. An exponential fed-batch strategy to prevent these inhibitions was developed in this work. The host strain is auxotrophic for phenylalanine, tyrosine and tryptophan. Due to low solubilities of tyrosine and tryptophan in the feed stream, tyrosine and tryptophan were dissolved separately in ammonia water to form a second feed stream. By dual feeding both streams at different exponential feed rates, a high cell density of 17.6 g/l and a final α-amylase activity of 41.4 U/ml and the overall biomass yield of 0.39 g cell/g glucose were achieved.

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Abbreviations

F :

Volumetric feed rate of feed stream 1 containing glucose (l/h)

F 1 :

Volumetric feed rate for feed stream 1 (l/h)

F 2 :

Volumetric feed rate for feed stream 2 (l/h)

S :

Glucose concentration in the broth (g/l)

S 0 :

Glucose concentration in the feed (g/l)

S N0 :

Concentration of the nutrient other than glucose (e.g., ammonium sulfate) in the feed (g/l)

S N :

Initial concentration of the nutrient other than glucose (e.g., ammonium sulfate) in the fermenter for a batch run (g/l)

t :

Time (h)

V 0 :

The broth volume when feed starts (l)

V :

The broth volume at time t (l)

X 0 :

The cell mass concentration when feed starts (g/l)

X :

The cell mass concentration in the broth at time t (g/l)

Y X/S :

Biomass yield on glucose (g/g)

\(Y_{X/S_N } \) :

Biomass yield on nutrient (g/g)

μ:

Specific growth rate (h−1)

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Authors and Affiliations

  1. Department of Chemical Engineering, Ohio University, Athens, OH, 45701, USA

    Hanjing Huang, Darin Ridgway & Tingyue Gu

  2. Department of Chemical Engineering, University of Waterloo, Waterloo, ON, Canada, N2L 3G1

    Murray Moo-Young

Authors
  1. Hanjing Huang
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  2. Darin Ridgway
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  3. Tingyue Gu
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  4. Murray Moo-Young
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Correspondence to Tingyue Gu.

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Huang, H., Ridgway, D., Gu, T. et al. Enhanced amylase production by Bacillus subtilis using a dual exponential feeding strategy. Bioprocess Biosyst Eng 27, 63–69 (2004). https://doi.org/10.1007/s00449-004-0391-z

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  • DOI: https://doi.org/10.1007/s00449-004-0391-z

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