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Application of Acid and Cold Stresses to Enhance the Production of Clavulanic Acid by Streptomyces clavuligerus

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Abstract

Clavulanic acid (CA) is frequently prescribed for treatment of bacterial infections. Despite the large number of studies concerning CA production, there is still a need to search for more effective and productive processes because it is mainly produced by biochemical route and is chemically unstable. This paper evaluates the influence of acid and cold stresses on CA production by Streptomyces clavuligerus in bench scale stirred tank bioreactor. Four batch cultures were conducted at constant pH (6.8 or 6.3) and temperature (30, 25, or 20 °C) and five batch cultures were performed with application of acid stress (pH reduction from 6.8 to 6.3), cold stress (reduction from 30 to 20 °C), or both. The highest maximum CA concentration (684.4 mg L−1) was obtained in the culture conducted at constant temperature of 20 °C. However, the culture under acid stress, in which the pH was reduced from 6.8 to 6.3 at a rate of 0.1 pH unit every 6 h, provided the most promising result, exhibiting a global yield coefficient of CA relative to cell formation (YCA/X) of 851.1 mgCA gX−1. High YCA/X values indicate that a small number of cells are able to produce a large amount of antibiotic with formation of smaller amounts of side byproducts. This could be especially attractive for decreasing the complexity and cost of the downstream processing, enhancing CA production.

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References

  1. Bush, K., Jacoby, G. A., & Medeiros, A. A. (1995). A functional classification scheme for beta-lactamases and its correlation with molecular-structure. Antimicrobial Agents and Chemotherapy, 39(6), 1211–1233.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Brown, A. G., Butterworth, D., Cole, M., Hanscomb, G., Hood, J. D., & Reading, C. (1976). Naturally-occurring beta-lactamase inhibitors with antibacterial activity. Journal of Antibiotics, 29(6), 668–669.

    Article  CAS  PubMed  Google Scholar 

  3. Baggaley, K. H., Brown, A. G., & Schofield, C. J. (1997). Chemistry and biosynthesis of clavulanic acid and other clavams. Natural Product Reports, 14(4), 309–333.

    Article  CAS  PubMed  Google Scholar 

  4. Teodoro, J. C., Baptista-Neto, A., Araujo, M. L. G. C., Hokka, C. O., & Badino, A. C. (2010). Influence of glycerol and ornithine feeding on clavulanic acid production by Streptomyces clavuligerus. Brazilian Journal of Chemical Engineering, 27(4), 499–506.

    Article  CAS  Google Scholar 

  5. Costa, C. L. L., & Badino, A. C. (2015). Overproduction of clavulanic acid by extractive fermentation. Electronic Journal of Biotechnology, 18(3), 154–160.

    Article  CAS  Google Scholar 

  6. Mayer, A. F., & Deckwer, W. D. (1996). Simultaneous production and decomposition of clavulanic acid during Streptomyces clavuligerus cultivations. Applied Microbiology and Biotechnology, 45(1-2), 41–46.

    Article  CAS  PubMed  Google Scholar 

  7. Ortiz, S. C. A., Hokka, C. O., & Badino, A. C. (2007). Utilization of soybean derivatives on clavulanic acid production by Streptomyces clavuligerus. Enzyme and Microbial Technology, 40(5), 1071–1077.

    Article  CAS  Google Scholar 

  8. Saudagar, P. S., & Singhal, R. S. (2007). Optimization of nutritional requirements and feeding strategies for clavulanic acid production by Streptomyces clavuligerus. Bioresource Technology, 98(10), 2010–2017.

    Article  CAS  PubMed  Google Scholar 

  9. Saudagar, P. S., & Singhal, R. S. (2007). A statistical approach using L-25 orthogonal array method to study fermentative production of clavulanic acid by Streptomyces clavuligerus MTCC 1142. Applied Biochemistry and Biotechnology, 136(3), 345–359.

    Article  CAS  PubMed  Google Scholar 

  10. Bellão, C., Antonio, T., Araujo, M. L. G. d. C., & Badino, A. (2013). Production of clavulanic acid and cephamycin C by Streptomyces clavuligerus under different fed-batch conditions. Brazilian Journal of Chemical Engineering, 30(2), 257–266.

    Article  Google Scholar 

  11. Cerri, M., & Badino, A. (2012). Shear conditions in clavulanic acid production by Streptomyces clavuligerus in stirred tank and airlift bioreactors. Bioprocess and Biosystems Engineering, 35(6), 977–984.

    Article  CAS  PubMed  Google Scholar 

  12. Rosa, J. C., Neto, A. B., Hokka, C. O., & Badino, A. C. (2005). Influence of dissolved oxygen and shear conditions on clavulanic acid production by Streptomyces clavuligerus. Bioprocess and Biosystems Engineering, 27(2), 99–104.

    Article  CAS  PubMed  Google Scholar 

  13. Li, R. F., & Townsend, C. A. (2006). Rational strain improvement for enhanced clavulanic acid production by genetic engineering of the glycolytic pathway in Streptomyces clavuligerus. Metabolic Engineering, 8(3), 240–252.

    Article  CAS  PubMed  Google Scholar 

  14. Kim, S. J., Kim, J. O., Shin, C. H., Park, H. W., & Kim, C. W. (2009). An approach to strain improvement and enhanced production of clavulanic acid in Streptomyces clavuligerus. Bioscience Biotechnology and Biochemistry, 73(1), 160–164.

    Article  CAS  Google Scholar 

  15. Kizildogan, A. K., Jaccard, G. V., Mutlu, A., Sertdemir, I., & Ozcengiz, G. (2017). Genetic engineering of an industrial strain of Streptomyces clavuligerus for further enhancement of clavulanic acid production. Turkish Journal of Biology, 41, 342–353.

    Article  CAS  Google Scholar 

  16. Hamedi, J., Imanparast, F., Tirandaz, H., Laamerad, B., & Sadrai, S. (2012). Improvement of clavulanic acid production by Streptomyces clavuligerus with peanut derivatives. Annals of Microbiology, 62(3), 1227–1234.

    Article  CAS  Google Scholar 

  17. Rodrigues, K. C. S., Souza, A. T., Pedrolli, D. B., Badino, A. C., & Cerri, M. O. (2018). Screening of medium contituents for clavulanic acid production by Streptomyces clavuligerus. Brazilian Journal of Microbiology., 49(4), 832–839.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Servant, P., & Mazodier, P. (1995). Characterization of streptomyces-albus 18-kilodalton heat shock-responsive protein. Journal of Bacteriology, 177(11), 2998–3003.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Nie, W., Wei, G., Du, G., Li, Y., & Chen, J. (2005). Enhanced intracellular glutathione synthesis and excretion capability of Candida utilis by using a low pH-stress strategy. Letters in Applied Microbiology, 40(5), 378–384.

    Article  CAS  PubMed  Google Scholar 

  20. Liang, G., Du, G., & Chen, J. (2008). Enhanced glutathione production by using low-pH stress coupled with cysteine addition in the treatment of high cell density culture of Candida utilis. Letters in Applied Microbiology, 46(5), 507–512.

    Article  CAS  PubMed  Google Scholar 

  21. Vohradsky, J., Li, X. M., Dale, G., Folcher, M., Nguyen, L., Viollier, P. H., & Thompson, C. J. (2000). Developmental control of stress stimulons in Streptomyces coelicolor revealed by statistical analyses of global gene expression patterns. Journal of Bacteriology, 182(17), 4979–4986.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Mikulík, K. & Palecková, P. (2007) Activity of ribosomes and tmRNA of Streptomyces aureofaciens during development and stress conditions induces by changes in temperature and the presence of antibiotics. Communicating Current Research and Educational Topics and Trends in Applied Microbiology, 11–18.

  23. Kim, Y. J., Song, J. Y., Moon, M. H., Smith, C. P., Hong, S. K., & Chang, Y. K. (2007). pH shock induces overexpression of regulatory and biosynthetic genes for actinorhodin productionin Streptomyces coelicolor A3(2). Applied Microbiology and Biotechnology, 76(5), 1119–1130.

    Article  CAS  PubMed  Google Scholar 

  24. Kim, Y. J., Moon, M. H., Song, J. Y., Smith, C. P., Hong, S. K., & Chang, Y. K. (2008). Acidic pH shock induces the expressions of a wide range of stress-response genes. BMC Genomics, 9(1), 604.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Teodoro, J. C., Baptista-Neto, A., Cruz-Hernandez, I. L., Hokka, C. O., & Badino, A. C. (2006). Influence of feeding conditions on clavulanic acid production in fed-batch cultivation with medium containing glycerol. Applied Microbiology and Biotechnology, 72(3), 450–455.

    Article  CAS  PubMed  Google Scholar 

  26. Bird, A. E., Bellis, J. M., & Gasson, B. C. (1982). Spectrophotometric assay of clavulanic acid by reaction with imidazole. Analyst, 107(1279), 1241–1245.

    Article  CAS  Google Scholar 

  27. Costa, C. L. L., & Badino, A. C. (2012). Production of clavulanic acid by Streptomyces clavuligerus in batch cultures without and with glycerol pulses under different temperature conditions. Biochemical Engineering Journal, 69, 1–7.

    Article  CAS  Google Scholar 

  28. Atala, D. I. P., Costa, A. C., Maciel, R., & Maugeri, F. (2001). Kinetics of ethanol fermentation with high biomass concentration considering the effect of temperature. Applied Biochemistry and Biotechnology, 91, 353–365.

    Article  PubMed  Google Scholar 

  29. Roubos, J. A., Krabben, P., de Laat, W., Babuska, R., & Heijnen, J. J. (2002). Clavulanic acid degradation in Streptomyces clavuligerus fed-batch cultivations. Biotechnology Progress, 18(3), 451–457.

    Article  CAS  PubMed  Google Scholar 

  30. Bersanetti, P. A., Almeida, R., Barboza, M., Araujo, M. L. G., & Hokka, C. O. (2005). Kinetic studies on clavulanic acid degradation. Biochemical Engineering Journal, 23(1), 31–36.

    Article  CAS  Google Scholar 

  31. Zeng, X., Zhao, J. J., Chen, X. S., Mao, Z. G., & Miao, W. Y. (2017). Insights into the simultaneous utilization of glucose and glycerol by Streptomyces albulus M-Z18 for high epsilon-poly-L-lysine productivity. Bioprocess and Biosystems Engineering, 40(12), 1775–1785.

    Article  CAS  PubMed  Google Scholar 

  32. Cebrian, G., Condon, S., & Manas, P. (2016). Influence of growth and treatment temperature on Staphylococcus aureus resistance to pulsed electric fields: relationship with membrane fluidity. Innovative Food Science & Emerging Technologies, 37, 161–169.

    Article  Google Scholar 

  33. Ballows, A., Truper, H., Dworkin, M., Harder, W., & Schleifer, K. (1992). The prokaryotes, a handbook on the biology of bacteria: ecophysiology, isolation, identification, applications. New York: Springer-Verlag.

    Google Scholar 

  34. Shuler, M. L., & Kargi, F. (2002). Bioprocess engineering: basic concepts. New York: Prentice Hall PTR.

    Google Scholar 

  35. Higgens, C., & Kastner, R. (1971). Streptomyces clavuligerus sp. nov., a β-lactam antibiotic producer. International Journal of Systematic and Evolutionary Microbiology, 21, 326–331.

    Google Scholar 

  36. Santos, V. C., Pereira, J. F. B., Haga, R. B., Rangel-Yagui, C. O., Teixeira, J. A. C., Converti, A., & Pessoa, A. (2009). Stability of clavulanic acid under variable pH, ionic strength and temperature conditions. A new kinetic approach. Biochemical Engineering Journal, 45(2), 89–93.

    Article  CAS  Google Scholar 

  37. Sanchez, L., & Brana, A. F. (1996). Cell density influences antibiotic biosynthesis in Streptomyces clavuligerus. Microbiology-Uk, 142(5), 1209–1220.

    Article  CAS  Google Scholar 

  38. Neves, A. A., Vieira, L. M., & Menezes, J. C. (2001). Effects of preculture variability on clavulanic acid fermentation. Biotechnology and Bioengineering, 72(6), 628–633.

    Article  CAS  PubMed  Google Scholar 

  39. Haginaka, J., Nakagawa, T., & Uno, T. (1981). Stability of clavulanic acid in aqueous solutions. Chemical and Pharmaceutical Bulletin, 29(11), 3334–3341.

    Article  CAS  Google Scholar 

  40. Ser, H. L., Law, J. W. F., Chaiyakunapruk, N., Jacob, S. A., Palanisamy, U. D., Chan, K. G., Goh, B. H., & Lee, L. H. (2016). Fermentation conditions that affect clavulanic acid production in Streptomyces clavuligerus: a systematic review. Frontiers in Microbiology, 7.

  41. Haga, R. B., Santos-Ebinuma, V. C., Silva, M. D. C., Pessoa, A., & Rangel-Yagui, C. O. (2013). Clavulanic acid partitioning in charged aqueous two-phase micellar systems. Separation and Purification Technology, 103, 273–278.

    Article  CAS  Google Scholar 

  42. Silva, M. D. C., Santos-Ebinuma, V. D., Lopes, A. M., & Rangel-Yagui, C. D. (2015). Dextran sulfate/triton X two-phase micellar systems as an alternative first purification step for clavulanic acid. Fluid Phase Equilibria, 399, 80–86.

    Article  CAS  Google Scholar 

  43. Barboza, M., Almeida, R. M., & Hokka, C. O. (2001). Kinetic studies of clavulanic acid recovery by ion exchange chromatography. Bioseparation, 10(4/5), 221–227.

    Article  CAS  PubMed  Google Scholar 

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Funding

The authors would like to thank Coordination for the Improvement of Higher Education Personnel (CAPES, process AUXPE 1363/2015) and the National Council for Scientific and Technological Development (CNPq, process #471848/2013-1) for financial support of this work. J. F. B. Pereira acknowledges the support of FAPESP (grant #2014/16424-7).

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Correspondence to M. O. Cerri.

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Rodrigues, K.C.S., Costa, C.L.L., Badino, A.C. et al. Application of Acid and Cold Stresses to Enhance the Production of Clavulanic Acid by Streptomyces clavuligerus. Appl Biochem Biotechnol 188, 706–719 (2019). https://doi.org/10.1007/s12010-019-02953-y

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