Agricultural waste materials enhance protease production by Bacillus subtilis B22 in submerged fermentation under blue light-emitting diodes

  • Punniyakotti Elumalai
  • Jeong-Muk Lim
  • Yool-Jin Park
  • Min ChoEmail author
  • Patrick J. Shea
  • Byung-Taek OhEmail author
Research Paper


Bacillus bacteria have major utility in large-scale production of industrial enzymes, among which proteases have particular importance. B. subtilis B22, an aerobic and chemotrophic strain, was isolated from kimchi and identified by 16S rRNA gene sequencing. Extracellular protease production was determined in basic medium, with 1% (w/v) casein as substrate, by submerged fermentation at 37 °C under blue, green, red and white light-emitting diodes (LEDs), white fluorescent light and darkness. Fermentation under blue LEDs maximized protease production (110.79 ± 1.8 U/mL at 24 h). Various agricultural waste products enhanced production and groundnut oil cake yielded the most protease (334 ± 1.8 U/mL at 72 h). Activity and stability of the purified protease were optimum at pH 7–10 and 20–60 °C. Activity increased in the presence of Ca2+, Mg2+ and Mn2+, while Fe2+, Zn2+, Co2+ and Cu2+ moderated activity, and Ni2+ and Hg2+ inhibited activity. Activity was high (98%) in the presence of ethylenediaminetetraacetic acid (EDTA) but inhibited by phenylmethanesulfonyl fluoride (PMSF). The protease was unaffected by nonionic surfactants, tolerated an anionic surfactant and oxidizing agents, and was compatible with multiple organic solvents. These properties suggest utility of protease produced by B. subtilis B22 under blue LEDs for industrial applications.


Agricultural waste products Blue LEDs B. subtilis Protease Submerged fermentation 



This work was supported by Korea Environment Industry & Technology Institute (KEITI) through Public Technology Program based on Environmental Policy, funded by Korea Ministry of Environment (MOE) (2018000200001).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

449_2019_2277_MOESM1_ESM.docx (231 kb)
Supplementary file1 (DOCX 231 kb)


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2020

Authors and Affiliations

  • Punniyakotti Elumalai
    • 1
    • 2
  • Jeong-Muk Lim
    • 2
  • Yool-Jin Park
    • 3
  • Min Cho
    • 2
    Email author
  • Patrick J. Shea
    • 4
  • Byung-Taek Oh
    • 2
    Email author
  1. 1.School of EnvironmentSouth China Normal University, University TownGuangzhouChina
  2. 2.Division of Biotechnology, Advanced Institute of Environment and Bioscience, College of Environmental and Bioresource SciencesChonbuk National UniversityIksanSouth Korea
  3. 3.Department of Ecology Landscape Architecture-Design, College of Environmental and Bioresource SciencesChonbuk National UniversityIksanSouth Korea
  4. 4.School of Natural ResourcesUniversity of Nebraska-LincolnLincolnUSA

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