Bioremediation Efficacy of Extracellular Chromate Reductase from Bacillus amyloliquefaciens (CSB 9) for Detoxification of Hexavalent Chromium

  • B. P. Rath
  • S. Das
  • H. Thatoi


Extracellular Cr(VI) reductase was produced by a chromate resistant bacterium Bacillus amyloliquefaciens with optimized physico-chemical parameters. Subsequently, the purified Cr(VI) reductase showed specific activity of 0.167 U/mg, 6% yield and 30.36-fold increase in purity. Based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the molecular weight (M) of the purified enzyme of interest was estimated to be ~116 KDa. The purified enzyme was further subjected for functional characterization (influence of pH, temperature and storage stability). The Cr(VI) reductase activity of the purified enzyme for temperature and pH optima was at 35 °C and 7.0 respectively on standard analysis conditions. Using potassium dichromate as substrate, the enzyme showed maximum activity (Vmax) of 3.5 U/mL with its corresponding KM value of 27.78 μM. The purified enzyme exhibited higher stability when treated with certain additives. These remarkable qualities found with this enzyme produced by B. amyloliquefaciens could make this an ideal candidate for bioremediation of Cr(VI) under a wide range of environmental conditions.


Bacillus amyloliquefaciens Bioremediation Chromate reductase Electrophoretical homogeneity Enzyme stability 


  1. 1.
    APHA, AWWA, WEF (1995) Standard methods for the examination of water and wastewater, 19th edn. APHA, Washington, DCGoogle Scholar
  2. 2.
    Bagchi D, Stohs SJ, Bernard WO, Bagchi M, Preus HG (2002) Cytotoxicity and oxidation mechanism of different forms of chromium. Toxicology 180:5–22PubMedCrossRefGoogle Scholar
  3. 3.
    Chen JM, Hao OJ (1998) Microbial chromium(IV) reduction. Crit Rev Environ Sci Technol. 28:219–251Google Scholar
  4. 4.
    Cheung KH, Gu JD (2007) Mechanism of hexavalent chromium detoxification by microorganisms and bioremediation application potential: A review. Int Biodeter Biodegr. 59(1):8–15CrossRefGoogle Scholar
  5. 5.
    Das S, Mishra J, Das SK, Pandey S, Rao DS, Chakraborty A, Sudarshan M, Das NN, Thatoi HN (2014) Bacterial chromate reductase, a potential enzyme for bioremediation of hexavalent chromium: a review. Chemosphere 96:112–121Google Scholar
  6. 6.
    Elangovan R, Philip L, Chandraraj K (2010) Hexavalent chromium reduction by free and immobilized cell-free extract of Arthrobacter rhombi-RE. Appl Biochem Biotechnol 160(1):81–97PubMedCrossRefGoogle Scholar
  7. 7.
    Gonzalez CF, Ackerley DF, Park CH, Matin A (2003) A Soluble Flavoprotein Contributes to Chromate Reduction and Tolerance by Pseudomonas putida. Acta Biotechnol 23:233–239CrossRefGoogle Scholar
  8. 8.
    Holt JG, Krieg NR, Sneath PHA, Staley JWS (1994) Bergey’s manual of derminative bacteriology, 9th edn. Williams and Wilkins, BaltimoreGoogle Scholar
  9. 9.
    McLean JS, Beveridge TJ (2001) Chromate reduction by a pseudomonad isolated from a site contaminated with chromated copper arsenate. Appl Environ Microbiol 67:1076–1084PubMedPubMedCentralCrossRefGoogle Scholar
  10. 10.
    Novitsky JA, MacSween MC (1989) Microbiology of a high energy beach sediment: evidence for an active and growing community. Mar Ecol Prog Ser. 52:71–75CrossRefGoogle Scholar
  11. 11.
    Park CH, Keyhan M, Wielinga B, Fendorf S, Matin A (2004) Chromate-Reducing Properties of Soluble Flavoproteins from Pseudomonas putida and Escherichia coli. Appl Environ Microbiol 66(5):1788–1795Google Scholar
  12. 12.
    Puzon GJ, Petersen JN, Roberts AG, Kramer DM, Xun L (2002) A bacterial flavin reductase system reduces chromate to a soluble chromium(III)-NAD(+) complex. Biochem Biophys Res 294(1):76–81PubMedCrossRefGoogle Scholar
  13. 13.
    Rath BP, Das S, Das Mohapatra PK, Thatoi HN (2014) Optimization of extracellular chromate reductase production by Bacillus amyloliquefaciens (CSB 9) isolated from chromite mine environment. Biocatal Agric Biotechnol. 3(3):35–41CrossRefGoogle Scholar
  14. 14.
    Sevgi E, Coral G, Gizir AM, Sagun MK (2009) Investigation of heavy metal resistancein some bacterial strains isolated from industrial soils. Turk J Biol 34:423–431Google Scholar
  15. 15.
    Sultan S, Hasnain S (2005) Isolation of hexavalent chromim-reducing Cr-tolerant facultative anaerobes from tannery effluent. J Gen Appl Microbiol. 47:307–312Google Scholar
  16. 16.
    Suzuki T, Miyata N, Horistu H, Kawai K, Takamizawa K, Tai Y, Okazaki M (1992). NAD(P)H-dependent chromium(VI) reductase of Pseudomonas ambigua G-1: a Cr(V) intermediate is formed during the reduction of Cr(VI) to Cr(III). J Bacteriol. 174(16):5340–5345PubMedPubMedCentralCrossRefGoogle Scholar
  17. 17.
    Vidali M (2001) Bioremediation. An overview*. Pure Appl Chem. 73(7):1163–1172CrossRefGoogle Scholar

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© Springer Nature Singapore Pte Ltd. 2019

Authors and Affiliations

  • B. P. Rath
    • 1
  • S. Das
    • 1
  • H. Thatoi
    • 2
  1. 1.Department of Biotechnology, College of Engineering and TechnologyBiju Patnaik University of TechnologyBhubaneswarIndia
  2. 2.Department of BiotechnologyNorth Orissa UniversityBaripada, MayurbhanjIndia

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