Polyhydroxyalkanoate (PHA) Production Using Waste Vegetable Oil by Pseudomonas sp. Strain DR2

  • Song, Jin-Hwan (Division of Environmental Science and Ecological Engineering, Korea University) ;
  • Jeon, Che-Ok (Department of Life Science, Chung-Ang University) ;
  • Choi, Mun-Hwan (Biomaterials Science Laboratory, Division of Life Science at the College of Natural Sciences, Gyeongsang National University) ;
  • Yoon, Sung-Chul (Biomaterials Science Laboratory, Division of Life Science at the College of Natural Sciences, Gyeongsang National University) ;
  • Park, Woo-Jun (Division of Environmental Science and Ecological Engineering, Korea University)
  • Published : 2008.08.31

Abstract

To produce polyhydroxyalkanoate (PHA) from inexpensive substrates by bacteria, vegetable-oil-degrading bacteria were isolated from a rice field using enrichment cultivation. The isolated Pseudomonas sp. strain DR2 showed clear orange or red spots of accumulated PHA granules when grown on phosphate and nitrogen limited medium containing vegetable oil as the sole carbon source and stained with Nile blue A. Up to 37.34% (w/w) of intracellular PHA was produced from corn oil, which consisted of three major 3-hydroxyalkanoates; octanoic (C8:0, 37.75% of the total 3-hydroxyalkanoate content of PHA), decanoic (C10:0, 36.74%), and dodecanoic (C12:0, 11.36%). Pseudomonas sp. strain DR2 accumulated up to 23.52% (w/w) of $PHA_{MCL}$ from waste vegetable oil. The proportion of 3-hydroxyalkanoate of the waste vegetable-oil-derived PHA [hexanoic (5.86%), octanoic (45.67%), decanoic (34.88%), tetradecanoic (8.35%), and hexadecanoic (5.24%)] showed a composition ratio different from that of the corn-oil-derived PHA. Strain DR2 used three major fatty acids in the same ratio, and linoleic acid was the major source of PHA production. Interestingly, the production of PHA in Pseudomonas sp. strain DR2 could not occur in either acetate- or butyrate-amended media. Pseudomonas sp. strain DR2 accumulated a greater amount of PHA than other well-studied strains (Chromobacterium violaceum and Ralstonia eutropha H16) when grown on vegetable oil. The data showed that Pseudomonas sp. strain DR2 was capable of producing PHA from waste vegetable oil.

Keywords

References

  1. Angelova, N. and D. Hunkeler. 1999. Rationalizing the design of polymeric biomaterials. Trends. Biotechnol. 17: 409-421 https://doi.org/10.1016/S0167-7799(99)01356-6
  2. Anderson, A. J. and E. A. Dawes. 1990. Occurrence, metabolism, metabolic role, and industrial uses of bacterial polyhydroxyalkanoates. Microbiol. Rev. 54: 450-472
  3. Chen, G. Q. and Q. Wu. 2005. The application of polyhydroxyalkanoates as tissue engineering materials. Biomaterials 26: 6565-6578 https://doi.org/10.1016/j.biomaterials.2005.04.036
  4. Choi, M. H. and S. C. Yoon. 1994. Polyester biosynthesis characteristics of Pseudomonas citronellolis grown on various carbon sources including 3-methyl-branched substrates. Appl. Environ. Microbiol. 60: 3245-3254
  5. Day, A. P. and J. D. Oliver. 2004. Changes in membrane fatty acid composition during entry of Vibrio vulnificus into the viable but nonculturable state. J. Microbiol. 42: 69-73
  6. Fernandez-Castillo, R., F. Rodriguez-Valera, J. Gonzalez-Ramos, and F. Ruiz-Berraquero. 1986. Accumulation of poly(R-3-hydroxybutyrate) by halobacteria. Appl. Envir. Microbiol. 51:214-216
  7. Godoy, F., M. Vancanneyt, M. Martinez, A. Steinbuchel, J. Swings, and B. H. Rehm. 2003. Sphingopyxis chilensis sp. nov, a chlorophenol-degrading bacterium that accumulates polyhydroxyalkanoate and transfer of Sphingomonas alaskensis to Sphingopyxis alaskensis comb. nov. Int. J. Syst. Evol. Microbiol. 53: 473-477 https://doi.org/10.1099/ijs.0.02375-0
  8. Graner, G., M. Hamberg, and J. Meijer. 2003. Screening of oxylipins for control of oilseed rape (Brassica napus) fungal pathogens. Phytochemistry 63: 89-95 https://doi.org/10.1016/S0031-9422(02)00724-0
  9. Gurieff, N. and P. Lant. 2007. Comparative life cycle assessment and financial analysis of mixed culture polyhydroxyalkanoate production. Bioresour. Technol. 98: 3393-3403 https://doi.org/10.1016/j.biortech.2006.10.046
  10. Hadi, R. S., S. M. Mousavi, H. M. Yeganeh, and I. Marc. 2007. Fatty acid and carotenoid production by Sporobolomyces ruberrimus when using technical glycerol and ammonium sulfate. J. Microbiol. Biotechnol. 17: 1591-1597
  11. Huang, T. Y., K. J. Duan, S. Y. Huang, and C. W. Chen. 2006. Production of polyhydroxyalkanoates from inexpensive extruded rice bran and starch by Haloferax mediterranei. J. Ind. Microbiol. Biotechnol. 33: 701-706 https://doi.org/10.1007/s10295-006-0098-z
  12. Ishizaki, A., K. Tanaka, and N. Taga. 2001. Microbial production of poly-D-3-hydroxybutyrate from $CO_2$. Appl. Microbiol. Biotechnol. 57: 6-12 https://doi.org/10.1007/s002530100775
  13. Kim, D. Y., H. W. Kim, M. G. Chung, and Y. H. Rhee. 2007. Biosynthesis, modification, and biodegradation of bacterial medium-chain-length polyhydroxyalkanoates. J. Microbiol. 45: 87-97
  14. Kim, T. K., M. T. Vo, H. D. Shin, and Y. H. Lee. 2005. Molecular structure of the PHA synthesis gene cluster from new mcl-PHA producer Pseudomonas putida KCTC1639. J. Microbiol. Biotechnol. 15: 1120-1124
  15. Kolibachuk, D., A. Miller, and D. Dennis. 1999. Cloning molecular analysis and expression of the polyhydroxyalkanoic acid synthase (phaC) gene from Chromobacterium violaceum. Appl. Environ. Microbiol. 65: 3561-3565
  16. Koller, M., P. Hesse, R. Bona,C. Kutschera, A. Atlie, and G. Braunegg. 2007. Potential of various archae- and eubacterial strains as industrial polyhydroxyalkanoate producers from whey. Macromol. Biosci. 7: 218-226 https://doi.org/10.1002/mabi.200600211
  17. Lang, S. and D. Wullbrandt. 1999. Rhamnose lipids-biosynthesis, microbial production and application potential. Appl. Microbiol. Biotechnol. 51: 22-32 https://doi.org/10.1007/s002530051358
  18. Li, R., Q. Chen, and P. G. Wang. 2007. A novel-designed Eshcherichia coli for the production of various polyhydroxyalkanoates from inexpensive substrate mixture. Appl. Microbiol. Biotechnol. 75: 1103-1109 https://doi.org/10.1007/s00253-007-0903-2
  19. Matsusaki, H., H. Abe, K. Taguchi, T. Fukui, and Y. Doi. 2000. Biosynthesis of poly(3-hydroxybutyrate-co-3-hydroxyalkanoates) by recombinant bacteria expressing the PHA synthase gene phaC1 from Pseudomonas sp. 61-3. Appl. Microbiol. Biotechnol. 53: 401-409 https://doi.org/10.1007/s002530051633
  20. Nomura, C. T. and S. Taguchi. 2007. PHA synthase engineering toward superbiocatalysts for custom-made biopolymers. Appl. Microbiol. Biotechnol. 73: 969-979
  21. Ostle, A. G. and J. G. Holt. 1982. Nile blue A as a fluorescent stain for poly-beta-hydroxybutyrate. Appl. Environ. Microbiol. 44: 238-241
  22. Park, I. J., Y. H. Rhee, N. Cho, and K. Shin. 2006. Cloning and analysis of medium-chain-length poly(3-hydroxyalkanoate) depolymerase gene of Pseudomonas luteola M13-4. J. Microbiol. Biotechnol. 16: 1935-1939
  23. Pohlmann, A., W. F. Fricke, F. Reinecke, B. Kusian, H. Liesegang, R. Cramm, et al. 2006. Genome sequence of the bioplastic-producing "Knallgas" bacterium Ralstonia eutropha H16. Nat. Biotechnol. 24: 1227-1229 https://doi.org/10.1038/nbt1006-1227
  24. Poirier, Y., C. Nawrath, and C. Somerville. 1995. Production of polyhydroxyalkanoates, a family of biodegradable plastics and elastomers in bacteria and plants. Biotechnology 13: 142-150 https://doi.org/10.1038/nbt0295-142
  25. Rehm, B. H., T. A. Mitsky, and A. Steinbuchel. 2001. Role of fatty acid de novo biosynthesis in polyhydroxyalkanoic acid (PHA) and rhamnolipid synthesis by pseudomonads: Establishment of the transacylase (PhaG)-mediated pathway for PHA biosynthesis in Escherichia coli. Appl. Environ. Microbiol. 67: 3102-3109 https://doi.org/10.1128/AEM.67.7.3102-3109.2001
  26. Sheu, D. S., Y. T. Wang, and C. Y. Lee. 2000. Rapid detection of polyhydroxyalkanoate-accumulating bacteria isolated from the environment by colony PCR. Microbiology 146: 2019-2025 https://doi.org/10.1099/00221287-146-8-2019
  27. Shin, D. S., M. S. Park, S. Jung, M. S. Lee, K. H. Lee, K. S. Bae, and S. B. Kim. 2007. Plant growth-promoting potential of endophytic bacteria isolated from roots of coastal sand dune plants. J. Microbiol. Biotechnol. 17: 1361-1368
  28. Spiekermann, P., B. H. Rehm, R. Kalscheuer, D. Baumeister, and A. Steinbüchel. 1999. A sensitive viable-colony staining method using Nile red for direct screening of bacteria that accumulate polyhydroxyalkanoic acids and other lipid storage compounds. Arch. Microbiol. 171: 73-80 https://doi.org/10.1007/s002030050681
  29. Stanier, R. Y., N. J. Palleroni, and M. Doudoroff. 1966. The aerobic pseudomonads: A taxonomic study. J. Gen. Microbiol. 43: 159-271 https://doi.org/10.1099/00221287-43-2-159
  30. Steinbuchel, A. and H. V. Valentin. 1995. Diversity of bacterial polyhydroxyalkanoic acids. FEMS Microbiol. Lett. 128: 219-228 https://doi.org/10.1111/j.1574-6968.1995.tb07528.x
  31. Suriyamongkol, P., R. Weselake, S. Narine, M. Moloney, and S. Shah. 2007. Biotechnological approaches for the production of polyhydroxyalkanoates in microorganisms and plants-A review. Biotechnol. Adv. 25: 148-175 https://doi.org/10.1016/j.biotechadv.2006.11.007
  32. Thompson, J. D., D. G. Higgins, and T. J. Gbson. 1994. Clustal W: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 22: 4673-4680 https://doi.org/10.1093/nar/22.22.4673
  33. Van Dyk, M. S., J. L. F. Kock, and A. Botha. 1994. Hydroxy long chain fatty-acids in fungi. World J. Microbiol. Biotechnol. 10: 495-504 https://doi.org/10.1007/BF00367653
  34. Ward, P. G., G. D. Roo, and K. E O'Connor. 2005. Accumulation of polyhydroxyalkanoate from styrene and phenylacetic acid by Pseudomonas putida CA-3. Appl. Environ. Microbiol. 71: 2046-2052 https://doi.org/10.1128/AEM.71.4.2046-2052.2005
  35. Yun, H. S., D. Y. Kim, C. W Chung, H. W. Kim, Y. K. Yang, and Y. H. Rhee. 2003. Characterization of a tacky poly(3-hydroxyalkanoate) produced by Pseudomonas chlororaphis HS21 from palm kernel oil. J. Microbiol. Biotechnol. 13: 64-69