International Microbiology

, Volume 21, Issue 4, pp 197–205 | Cite as

Extracellular lipase from Pseudomonas aeruginosa JCM5962(T): Isolation, identification, and characterization

  • Shweta Sachan
  • Mohammed Shariq Iqbal
  • Aditi Singh
Original Article


The study was done to isolate, identify, and characterize a good lipolytic strain from soil. Lipolytic strain isolation was done using tributyrin agar medium. The biochemical testing and 16S rRNA gene sequencing analysis was done for identification. The enzyme was purified using ammonium sulfate precipitation and column chromatography. Results have shown a novel high lipolytic strain of P. aeruginosa JCM5962(T), isolated from soil of sugarcane field. The 16S rRNA sequence analysis confirmed the strain as P. aeruginosa JCM5962(T); further, the sequence was submitted to Genbank (KX946966.1). The isolate produced an extracellular lipase which was purified as single band of 31 kDa. Maximum lipase activity was observed at 50 °C and pH 8.0. Activity was enhanced in the presence of cobalt and benzene solvent, whereas mercury, sodium dodecyl sulfate, and chloroform inhibited it. The enzyme’s marked stability and activity at high temperature, alkaline pH and organic solvents suggest that this can be effectively used in a variety of applications in industries and as biotechnological tools.


Extracellular lipase Pseudomonas aeruginosa Metal ions Cobalt Benzene 



The authors are thankful to Chairperson, Pro VC, Amity University Lucknow campus and Head, Amity Institute of Biotechnology for providing support and all necessary facilities to conduct the work. Authors are also thankful to IMTECH, Chandigarh, for 16S rRNA sequencing.


  1. Ahmed EH, Raghavendra T, Madamwar D (2010) A thermostable alkaline lipase from a local isolate Bacillus subtilis EH 37: characterization, partial purification and application in organic synthesis. Appl Biochem Biotechnol 160(7):2102–2113CrossRefGoogle Scholar
  2. Anbu P, Hur BK (2014) Isolation of an organic solvent-tolerant bacterium Bacillus licheniformis PAL05 that is able to secrete solvent-stable lipase. Biotechnol Appl Biochem 61(5):528–534CrossRefGoogle Scholar
  3. Arpigny JL, Jaeger KE (1999) Bacterial lipolytic enzymes: classification and properties. Biochem J 343:177–183CrossRefGoogle Scholar
  4. Bisht D, Yadav SK, Gautam P, Darmwal NS (2013) Simultaneous production of alkaline lipase and protease by antibiotic and heavy metal tolerant Pseudomonas aeruginosa. J Basic Microbiol 53(9):715–722CrossRefGoogle Scholar
  5. Bisht D, Yadav SK, Darmwal NS (2014) An oxidant and organic solvent tolerant alkaline lipase by P. aeruginosa mutant: downstream processing and biochemical characterization. Braz J Microbiol 44(4):1305–1314CrossRefGoogle Scholar
  6. Boonsinthai B, Phutraku S (1999) Effect of metal ions, inhibitors and denaturants on extracellular lipases from three thermophile isolates and their clones. Chiang Mai J Sci 26(1):25–43Google Scholar
  7. Collee JG, Fraser AG, Marmion BP, Simmons A, Mackie MC (1996) Practical medical microbiology, 14th edn. Churchill Livingstone, LondonGoogle Scholar
  8. Davis BJ (1964) Disc electrophoresis II. Method and application to human serum proteins. Ann N Y Acad Sci 121:321–349Google Scholar
  9. Davranov K (1994) Microbial lipases in biotechnology (review). Appl Biochem Microbiol 30:427–432Google Scholar
  10. Demain AL (2000) Microbial biotechnology. Trends Biotechnol 18:26–31CrossRefGoogle Scholar
  11. Ghori MI, Iqbal MJ, Hameed A (2011) Characterization of a novel lipase from Bacillus sp. isolated from tannery wastes. Braz J Microbiol 42(1):22–29CrossRefGoogle Scholar
  12. Gokbulut AA, Arslanoglu A (2013) Purification and biochemical characterization of an extracellular lipase from psychrotolerant Pseudomonas fluorescens KE38. Turk J Biol 37:538–546CrossRefGoogle Scholar
  13. Guncheva M, Zhiryakova D (2011) Catalytic properties and potential applications of Bacillus lipases. J Mol Catal B Enzym 68(1):1–21CrossRefGoogle Scholar
  14. Guncheva M, Tashev E, Zhiryakova D, Tosheva T, Tzokova N (2011) Immobilization of lipase from Candida rugosa on novel phosphorous-containing polyurethanes: application in wax ester synthesis. Process Biochem 46(4):923–930CrossRefGoogle Scholar
  15. Gupta R, Gupta N, Rathi P (2004) Bacterial lipases: an overview of production, purification and biotechnological properties. Appl Microbiol Biotechnol 64:763–781CrossRefGoogle Scholar
  16. Gupta A, Joseph B, Mani A, Thomas G (2008) Biosynthesis and properties of an extracellular thermostable serine alkaline protease from Virgibacillus pantothenticus. World J Microbiol Biotechnol 24:237–243CrossRefGoogle Scholar
  17. Hasan F, Shah AA, Hameed A (2006) Industrial applications of microbial lipases. Enzym Microb Technol 39:235–251CrossRefGoogle Scholar
  18. Jaeger KE, Eggert T (2002) Lipases for biotechnology. Curr Opin Biotechnol 13(4):390–397CrossRefGoogle Scholar
  19. Ji Q, Xiao S, He B, Liu X (2010) Purification and characterization of an organic solvent-tolerant lipase from Pseudomonas aeruginosa LX1 and its application for biodiesel production. J Mol Catal B Enzym 66:264–269CrossRefGoogle Scholar
  20. Kakugawa K, Shobayashi M, Suzuki O, Miyakawa T (2002) Purification and characterization of a lipase from the glycolipid-producing yeast Kurtzmanomyces sp. I-11. Biosci Biotechnol Biochem 66(5):978–985CrossRefGoogle Scholar
  21. Kanderi DK, Yadav U, Satyanarayana SV, Verma S (2014) Characterization of partially purified lipase from Saccharomyces cerevisiae. Int J Pharm Pharm Sci 6(8):514–517Google Scholar
  22. Karadzic I, Masui A, Zivkovic LI, Fujiwara N (2006) Purification and characterization of an alkaline lipase from Pseudomonas aeruginosa isolated from putrid mineral cutting oil as component of metal working fluid. J Biosci Bioeng 102(2):82–89CrossRefGoogle Scholar
  23. Kumar A, Dhar K, Kanwar SS, Arora PK (2016) Lipase catalysis in organic solvents: advantages and applications. Biol Proced Online 18(2):1–11Google Scholar
  24. Martinez A, Soberon CG (2001) Characterization of the lipA gene encoding the major lipase from Pseudomonas aeruginosa strain IGB83. Appl Microbiol Biotechnol 56:731–735CrossRefGoogle Scholar
  25. Mukke VK, Chinte DN (2012) Impact of heavy metal induced alterations in lipase activity of fresh water crab, Barytelphusa guerini. J Chem Pharm Res 4(5):2763–2766Google Scholar
  26. Nielsen PM, Brask J, Fjerbaek L (2008) Enzymatic biodiesel production: technical and economical considerations. Eur J Lipid Sci Technol 110:692–700CrossRefGoogle Scholar
  27. Paje ML, Neilan BA, Couperwhite I (1997) A Rhodococcus species that thrives on medium saturated with liquid benzene. Microbiol 143:2975–2981CrossRefGoogle Scholar
  28. Pourahmad JR, Ebadi R (2013) Study the expression of mara gene in ciprofloxacin and tetracycline resistant mutants of Esherichia coli. Iran J Pharm Res 12:923–928Google Scholar
  29. Pramanik K, Saren S, Mitra S, Ghosh PK, Maiti TK (2018) Computational elucidation of phylogenetic, structural and functional characteristics of Pseudomonas lipases. Comput Biol Chem 74:190–200CrossRefGoogle Scholar
  30. Reetz MT (2002) Lipases as practical biocatalysts. Curr Opin Chem Biol 6(2):145–150CrossRefGoogle Scholar
  31. Romero CM, Pera LM, Loto F, Vallejos C, Castro G, Baigori M (2012) Purification of an organic solvent-tolerant lipase from Aspergillus niger MYA 135 and its application in ester synthesis. Biocatal Agric Biotechnol 1(1):25–31CrossRefGoogle Scholar
  32. Sachan S, Singh A (2015) Lipase enzyme and its diverse role in food processing industry. Everyman’s Sci 4:214–218Google Scholar
  33. Sachan S, Chandra VY, Yadu A, Singh A (2017) Cobalt has enhancing effect on extracellular lipases isolated from Pseudomonas aeruginosa JCM5962(T). Int J PharmTech Res 10(1):45–49CrossRefGoogle Scholar
  34. Saranya P, Kumari HS, Rao BP, Sekaran G (2014) Lipase production from a novel thermo-tolerant and extreme acidophile Bacillus pumilus using palm oil as the substrate and treatment of palm oil-containing wastewater. Environ Sci Pollut Res 21:3907–3919CrossRefGoogle Scholar
  35. Schmid RD, Verger R (1998) Lipases: interfacial enzymes with attractive applications. Angew Chem Int Ed Engl 37(12):1608–1633CrossRefGoogle Scholar
  36. Shah KR, Bhatt SA (2011) Purification and characterization of lipase from Bacillus subtilis Pa2. J Biochem Tech 3(3):292–295Google Scholar
  37. Siddiqui KS, Azhar MJ, Rashid MH, Rajoka MI (1997) Stability and identification of active-site residues of carboxym ethyl cellulases from Aspergillus niger and Cellulomonas biazotea. Folia Microbiol (Praha) 42:312–318CrossRefGoogle Scholar
  38. Stuer W, Jaeger KE, Winkler U (1986) Purification of extracellular lipase from Pseudomonas aeruginosa. J Bacteriol 168:1070–1074CrossRefGoogle Scholar
  39. Svendsen A, Borch K, Barfoed M, Nielsen T, Gormsen E, Patkar S (1995) Biochemical properties of cloned lipases from Pseudomonas family. Biochim Biophys Acta 1259:9–17CrossRefGoogle Scholar
  40. Thakur S (2012) Lipases, its sources, properties and applications: a review. Int J Sci Eng Res 3(7):1–29Google Scholar
  41. Tiwari P, Upadhyay MK, Silawat N, Verma HN (2011) Optimization and characterization of a thermo tolerant lipase from Cryptococcus albidus. Der Pharma Chemica 3(4):501–508Google Scholar
  42. Torres S, Castro GR (2003) Organic solvent resistant lipase produced by thermoresistant bacteria. New Horizons Biotechnol, Netherlands: Springer 18(2):113–122CrossRefGoogle Scholar
  43. Ulker S, Ozel A, Colak A, Karaoglu SA (2011) Isolation, production, and characterization of an extracellular lipase from Trichoderma harzianum isolated from soil. Turk J Biol 35(5):543–550Google Scholar
  44. Verma ML, Azmi W, Kanwar SS (2008) Microbial lipases: at the interface of aqueous and non-aqueous media: a review. Acta Microbiol Immunol Hung 55(3):265–294CrossRefGoogle Scholar
  45. Zouaoui B, Bouziane A, Ghalem BR (2012) Isolation, purification and properties of lipase from Pseudomonas aeruginosa. Afr J Biotechnol 11(60):12415–12421Google Scholar

Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Amity Institute of BiotechnologyAmity University Uttar PradeshLucknowIndia

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