Biotechnology Letters

, Volume 41, Issue 2, pp 203–220 | Cite as

Solvent stable microbial lipases: current understanding and biotechnological applications

  • Priyanka Priyanka
  • Yeqi Tan
  • Gemma K. Kinsella
  • Gary T. Henehan
  • Barry J. RyanEmail author



This review examines on our current understanding of microbial lipase solvent tolerance, with a specific focus on the molecular strategies employed to improve lipase stability in a non-aqueous environment.


It provides an overview of known solvent tolerant lipases and of approaches to improving solvent stability such as; enhancing stabilising interactions, modification of residue flexibility and surface charge alteration. It shows that judicious selection of lipase source supplemented by appropriate enzyme stabilisation, can lead to a wide application spectrum for lipases.


Organic solvent stable lipases are, and will continue to be, versatile and adaptable biocatalytic workhorses commonly employed for industrial applications in the food, pharmaceutical and green manufacturing industries.


Industrial biocatalysis Lipase Lipase engineering Organic solvent stability Organic synthesis 



This work was supported by the Dublin Institute of Technology under the Fiosraigh Scholarship (PP and YT).

Author contributions

Conceived study (PP, YT, GKK, GTH, BJR), Performed research (PP, YT), Analyzed data (PP, YT), Contributed methods (GKK, GTH, BJR), Wrote the paper (PP, YT, GKK, GTH, BJR).


This work was supported by the Dublin Institute of Technology under the Fiosraigh Scholarship (PP and YT).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.


  1. Abdulmalek E, Hamidon NF, Abdul Rahman MB (2016) Optimization and characterization of lipase catalysed synthesis of xylose caproate ester in organic solvents. J Mol Catal B Enzym 132:1–4Google Scholar
  2. Abuin E, Lissi E, Jara P (2007) Effect of the organic solvent on the interfacial micropolarity of AOT -water reverse micelles. J Chil Chem Soc 52:1082–1087Google Scholar
  3. Adlercreutz P (2013) Immobilisation and application of lipases in organic media. Chem Soc Rev 42:6406–6436Google Scholar
  4. Ahmed EH, Raghavendra T, Madamwar D (2010) An alkaline lipase from organic solvent tolerant Acinetobacter sp. EH28: application for ethyl caprylate synthesis. Bioresour Technol 101:3628–3634Google Scholar
  5. Aris MH, Annuar MSM, Ling TC (2016) Lipase-mediated degradation of poly-??-caprolactone in toluene: behavior and its action mechanism. Polym Degrad Stab 133:182–191Google Scholar
  6. Arnold FH (1990) Engineering enzymes for non-aqueous solvents. Trends Biotechnol 8:244–249Google Scholar
  7. Augustyniak W, Brzezinska AA, Pijning T et al (2012) Biophysical characterization of mutants of Bacillus subtilis lipase evolved for thermostability: factors contributing to increased activity retention. Protein Sci 21:487–497Google Scholar
  8. Ayaz B, Ugur A, Boran R (2014) Purification and characterization of organic solvent-tolerant lipase from Streptomyces sp. OC119-7 for biodiesel production. Biocatal Agric Biotechnol 4(1):103–108. Google Scholar
  9. Badgujar KC, Pai PA, Bhanage BM (2016) Enhanced biocatalytic activity of immobilized Pseudomonas cepacia lipase under sonicated condition. Bioprocess Biosyst Eng 39:211–221Google Scholar
  10. Banerjee A, Chatterjee K, Madras G (2014) Enzymatic degradation of polymers: a brief review. Mater Sci Technol 30:567–573. Google Scholar
  11. Barrera-rivera KA, Flores-carreón A (2012) Synthesis of biodegradable polymers using biocatalysis with Yarrowia lipolytica. In: Lipase Chapter 28 synthesis of biodegradable polymers using biocatalysis with Yarrowia lipolytica Lipase.
  12. Ben Bacha A, Moubayed NMS, Al-Assaf A (2016) An organic solvent-stable lipase from a newly isolated Staphylococcus aureus ALA1 strain with potential for use as an industrial biocatalyst. Biotechnol Appl Biochem 63(3):378–390Google Scholar
  13. Benkovic SJ, Hammes-Schiffer S (2003) A perspective on enzyme catalysis. Science 301:1196–1202Google Scholar
  14. Benson NC, Daggett V (2012) A comparison of multiscale methods for the analysis of molecular dynamics simulations. J Phys Chem B 116:8722–8731Google Scholar
  15. Bose A, Keharia H (2013) Production, characterization and applications of organic solvent tolerant lipase by Pseudomonas aeruginosa AAU2. Biocatal Agric Biotechnol 2:255–266. Google Scholar
  16. Brady L, Brzozowski AM, Derewenda ZS et al (1990) A serine protease triad forms the catalytic centre of a triacylglycerol lipase. Nature 343:767–770. Google Scholar
  17. Cao Y et al (2012) Purification and characterization of an organic solvent-stable lipase from Pseudomonas stutzeri LC2-8 and its application for efficient resolution of (R, S)-1-phenylethanol. Biochem Eng J 64:55–60Google Scholar
  18. Chakravorty D, Parameswaran S, Dubey VK, Patra S (2012) Unraveling the rationale behind organic solvent stability of lipases. Appl Biochem Biotechnol 167:439–461Google Scholar
  19. Chen B, Hu J, Miller EM et al (2008) Candida antarctica Lipase B chemically immobilized on epoxy-activated micro- and nanobeads: catalysts for polyester synthesis. Biomacromol 9:463–471Google Scholar
  20. Cobb RE, Chao R, Zhao H (2013) Directed evolution: past, present and future. AIChE J 59:1432–1440. Google Scholar
  21. De Godoy Daiha K et al (2015) Are lipases still important biocatalysts? A study of scientific publications and patents for technological forecasting. PLoS ONE 10(6):e0131624Google Scholar
  22. De Souza TC et al (2016) Cashew apple bagasse as a support for the immobilization of lipase B from Candida antarctica: application to the chemoenzymatic production of (R)-Indanol. J Mol Catal B 130:58–69Google Scholar
  23. Díaz-García ME, Valencia-González MJ (1995) Enzyme catalysis in organic solvents: a promising field for optical biosensing. Talanta 42:1763–1773Google Scholar
  24. Dror A, Shemesh E, Dayan N, Fishman A (2014) Protein engineering by random mutagenesis and structure-guided consensus of Geobacillus stearothermophilus lipase T6 for enhanced stability in methanol. Appl Environ Microbiol 80:1515–1527Google Scholar
  25. Dror A, Kanteev M, Kagan I et al (2015) Structural insights into methanol-stable variants of lipase T6 from Geobacillus stearothermophilus. Appl Microbiol Biotechnol 99:9449–9461Google Scholar
  26. Duchiron SW et al (2017) Enzymatic synthesis of poly(ε-caprolactone-co-ε-thiocaprolactone). Eur Polym J 87:147–158Google Scholar
  27. Dutta Banik S, Nordblad M, Woodley JM, Peters GH (2016) A Correlation between the activity of Candida antarctica Lipase B and differences in binding free energies of organic solvent and substrate. ACS Catal 6:6350–6361Google Scholar
  28. Fischer M, Peiker M, Thiele C, Schmid RD (2000) Lipase engineering database understanding and exploiting sequence—structure—function relationships Jurgen. J Mol Catal 10:491–508Google Scholar
  29. Grosch JH, Wagner D, Nistelkas V, Spie AC (2017) Thermodynamic activity-based intrinsic enzyme kinetic sheds light on enzyme-solvent interactions. Biotechnol Prog 33:96–103Google Scholar
  30. Halling PJ (1997) Predicting the behaviour of lipases in low-water media. Biochem Soc Trans 25:170–174Google Scholar
  31. Hasan F, Shah AA, Hameed A (2006) Industrial applications of microbial lipases. Enzyme Microb Technol 39:235–251Google Scholar
  32. Illanes A, Cauerhff A, Wilson L, Castro GR (2012) Recent trends in biocatalysis engineering. Bioresour Technol 115:48–57Google Scholar
  33. Iyer PV, Ananthanarayan L (2008) Enzyme stability and stabilization-Aqueous and non-aqueous environment. Process Biochem 43:1019–1032Google Scholar
  34. Jain D, Mishra S (2015) Multifunctional solvent stable Bacillus lipase mediated biotransformations in the context of food and fuel. J Mol Catal B 117:21–30Google Scholar
  35. Jia C et al (2010) A simple approach for the selective enzymatic synthesis of dilauroyl maltose in organic media. J Mol Catal B 62(3–4):265–269Google Scholar
  36. Jiang X et al (2013a) Synthesis of vitamin E succinate from Candida rugosa lipase in organic medium. Chem Res Chin Univ 29(2):223–226Google Scholar
  37. Jiang Z et al (2013b) Synthesis of phytosterol esters catalyzed by immobilized lipase in organic media. Chin J Catal 34(12):2255–2262Google Scholar
  38. Kamarudin NHA et al (2014) A new cold-adapted, organic solvent stable lipase from mesophilic Staphylococcus epidermidis AT2. Protein J 33(3):296–307Google Scholar
  39. Kanmani P et al (2016) Enzymatic degradation of polyhydroxyalkanoate using lipase from Bacillus subtilis. Int J Environ Sci Technol 13(6):1541–1552Google Scholar
  40. Kawata T, Ogino H (2009) Enhancement of the organic solvent-stability of the LST-03 lipase by directed evolution. Biotechnol Prog 25:1605–1611Google Scholar
  41. Kawata T, Ogino H (2010) Amino acid residues involved in organic solvent-stability of the LST-03 lipase. Biochem Biophys Res Commun 400(3):384–388Google Scholar
  42. Kazlauskas RJ (1994) Elucidating structure-mechanism relationships in lipases: prospects for predicting and engineering catalytic properties. Trends Biotechnol 12:464–472Google Scholar
  43. Khan FI, Lan D, Durrani R et al (2017) The lid domain in lipases: structural and functional determinant of enzymatic properties. Front Bioeng Biotechnol 5:1–13. Google Scholar
  44. Kirdi R et al (2017) Mycelium-bound lipase from Aspergillus oryzae as efficient biocatalyst for cis-3-hexen-1-yl acetate synthesis in organic solvent. Biocatal Agric Biotechnol 10:13–19Google Scholar
  45. Kobayashi S, Uyama H, Takamoto T (2000) Lipase-catalyzed degradation of polyesters in organic solvents. A new methodology of polymer recycling using enzyme as catalyst. Biomacaromolecules 1(1):3–5Google Scholar
  46. Korman TP, Sahachartsiri B, Charbonneau DM et al (2013) Dieselzymes: development of a stable and methanol tolerant lipase for biodiesel production by directed evolution. Biotechnol Biofuels 6:70Google Scholar
  47. Kulschewski T, Sasso F, Secundo F et al (2013) Molecular mechanism of deactivation of C. antarctica lipase B by methanol. J Biotechnol 168:462–469. Google Scholar
  48. Kumar D, Parshad R, Gupta VK (2014a) Application of a statistically enhanced, novel, organic solvent stable lipase from Bacillus safensis DVL-43. Int J Biol Macromol 66:97–107Google Scholar
  49. Kumar V et al (2014b) Engineering lipase A from mesophilic Bacillus subtilis for activity at low temperatures. Protein Eng Des Sel 27(3):73–82Google Scholar
  50. Kumar A et al (2015) Cellulose binding domain assisted immobilization of lipase (GSlip-CBD) onto cellulosic nanogel: Characterization and application in organic medium. Colloids Surf B 136:1042–1050. Google Scholar
  51. Kumar A et al (2016) Lipase catalysis in organic solvents: advantages and applications. Biol Proced Online 18:2Google Scholar
  52. Li X, Yu HY (2014) Characterization of an organic solvent-tolerant lipase from Haloarcula sp. G41 and its application for biodiesel production. Folia Microbiologica 59(6):455–463Google Scholar
  53. Li X et al (2014) Characterization of an organic solvent-tolerant lipase from Idiomarina sp. W33 and its application for biodiesel production using Jatropha oil. Extremophiles 18(1):171–178Google Scholar
  54. Maiangwa J, Mohamad Ali MS, Salleh AB et al (2017) Lid opening and conformational stability of T1 Lipase is mediated by increasing chain length polar solvents. PeerJ 5:e3341. Google Scholar
  55. Mander P, Cho SS, Simkhada JR et al (2012) An organic solvent—tolerant lipase from Streptomyces sp. CS133 for enzymatic transesterification of vegetable oils in organic media. Process Biochem 47:635–642. Google Scholar
  56. Masomian M et al (2013) A new thermostable and organic solvent-tolerant lipase from Aneurinibacillus thermoaerophilus strain HZ. Process Biochem 48(1):169–175. Google Scholar
  57. Matte CR, Bordinhaõ C, Poppe JK et al (2016) Synthesis of butyl butyrate in batch and continuous enzymatic reactors using Thermomyces lanuginosus lipase immobilized in Immobead 150. J Mol Catal B 127:67–75. Google Scholar
  58. McAuley M, Timson DJ (2016) Modulating mobility: a paradigm for protein engineering? Appl Biochem Biotechnol 181:1–8Google Scholar
  59. Miyazawa T, Hamada M, Morimoto R, Maeda Y (2014) Candida antarctica lipase B-mediated regioselective acylation of dihydroxybenzenes in organic solvents. Tetrahedron 71:3915–3923. Google Scholar
  60. Mo Q et al (2016) A novel thermostable and organic solvent-tolerant lipase from Xanthomonas oryzae pv. oryzae YB103: screening, purification and characterization. Extremophiles 20(2):157–165Google Scholar
  61. Molina-Gutiérrez M et al (2016) Green synthesis of β-sitostanol esters catalyzed by the versatile lipase/sterol esterase from Ophiostoma piceae. Food Chem. Google Scholar
  62. Monsef Shokri M, Ahmadian S, Akbari N, Khajeh K (2014) Hydrophobic substitution of surface residues affects lipase stability in organic solvents. Mol Biotechnol 56:360–368Google Scholar
  63. Nakano H, Ide Y, Tsuda T et al (1998) Improvement in the organic solvent stability of pseudomonas lipase by random mutation. Annals of the New York Academy of Sciences. Blackwell Publishing Ltd, Oxford, pp 431–434Google Scholar
  64. Narwal SK et al (2016) Green synthesis of isoamyl acetate via silica immobilized novel thermophilic lipase from Bacillus aerius. Russ J Bioorg Chem 42(1):69–73. Google Scholar
  65. Ogino H, Ishikawa H (2001) Enzymes which are stable in the presence of organic solvents. J Biosci Bioeng 91:109–116Google Scholar
  66. Öztürk Düşkünkorur H et al (2014) Lipase catalyzed synthesis of polycaprolactone and clay-based nanohybrids. Polymer (United Kingdom) 55(7):1648–1655Google Scholar
  67. Park HJ, Joo JC, Park K, Yoo YJ (2012) Stabilization of Candida antarctica lipase B in hydrophilic organic solvent by rational design of hydrogen bond. Biotechnol Bioprocess Eng 17:722–728Google Scholar
  68. Park HJ, Joo JC, Park K et al (2013a) Prediction of the solvent affecting site and the computational design of stable Candida antarctica lipase B in a hydrophilic organic solvent. J Biotechnol 163:346–352Google Scholar
  69. Park HJ, Park K, Yoo YJ (2013b) Understanding the effect of tert-butanol on Candida antarctica lipase B using molecular dynamics simulations. Mol Simul 39:653–659Google Scholar
  70. Patel V et al (2015) Synthesis of ethyl caprylate in organic media using Candida rugosa lipase immobilized on exfoliated graphene oxide: process parameters and reusability studies. Biochem Eng J 95:62–70. Google Scholar
  71. Reetz MT, Carballeira JD, Vogel A (2006) Iterative saturation mutagenesis on the basis of b factors as a strategy for increasing protein thermostability. Angew Chemie - Int Ed 45:7745–7751Google Scholar
  72. Reetz MT, Soni P, Fernández L et al (2010) Increasing the stability of an enzyme toward hostile organic solvents by directed evolution based on iterative saturation mutagenesis using the B-FIT method. Chem Commun (Camb) 46:8657–8658Google Scholar
  73. Ringborg RH, Woodley JM (2016) The application of reaction engineering to biocatalysis. React Chem Eng 1:10–22Google Scholar
  74. Romero CM et al (2014) Activity and stability of lipase preparations from Penicillium corylophilum: Potential use in biocatalysis. Chem Eng Technol 37(11):1987–1992Google Scholar
  75. Salihu A, Alam MZ (2015) Solvent tolerant lipases: a review. Process Biochem 50:86–96Google Scholar
  76. Sandoval GC, Marty A, Condoret J-S (2001) Thermodynamic activity-based enzyme kinetics: efficient tool for nonaqueous enzymology. AIChE J 47:718–726Google Scholar
  77. Schulze B, Klibanov AM (1991) Inactivation and stabilization of stabilisins in neat organic solvents. Biotechnol Bioeng 38:1001–1006Google Scholar
  78. Sharma S, Kanwar SS (2014) Organic solvent tolerant lipases and applications. Sci World J 2014:625258Google Scholar
  79. Sivaramakrishnan R, Incharoensakdi A (2016) Purification and characterization of solvent tolerant lipase from Bacillus sp. for methyl ester production from algal oil. J Biosci Bioeng 121(5):517–522Google Scholar
  80. Su H, Mai Z, Zhang S (2016) Cloning, expression and characterization of a lipase gene from marine bacterium Pseudoalteromonas lipolytica SCSIO 04301. J Ocean Univ China 15(6):1051–1058. Google Scholar
  81. Tamilarasan K, Kumar MD (2012) Biocatalysis and agricultural biotechnology purification and characterization of solvent tolerant lipase from Bacillus sphaericus MTCC 7542. Biocatal Agric Biotechnol 1:309–313. Google Scholar
  82. Tao J, Kazlauskas RJ, Romas J (2011) Biocatalysis for green chemistry and chemical process development. Wiley, New YorkGoogle Scholar
  83. Timasheff SN (1993) The control of protein stability and association by weak interactions with water: how do solvents affect these processes? Annu Rev Biophys Biomol Struct 22:67–97Google Scholar
  84. Tufvesson P, Lima-Ramos J, Al Haque N et al (2013) Advances in the process development of biocatalytic processes. Org Process Res Dev 17:1233–1238Google Scholar
  85. Vescovi V et al (2017) Immobilized lipases on functionalized silica particles as potential biocatalysts for the synthesis of fructose oleate in an organic solvent/water system. Molecules 22(2):212Google Scholar
  86. Villeneuve P, Muderhwa JM, Graille J, Haas MJ (2000) Customizing lipases for biocatalysis: a survey of chemical, physical and molecular biological approaches. J Mol Catal B 9:113–148. Google Scholar
  87. Vrutika P, Datta M (2015) Lipase from solvent-tolerant Pseudomonas sp. DMVR46 strain adsorb on multiwalled carbon nanotubes: application for enzymatic biotransformation in organic solvents. Appl Biochem Biotechnol 177(6):1313–1326Google Scholar
  88. Vrutika P, Shruti N, Datta M (2014) An extracellular solvent stable alkaline lipase from Pseudomonas sp. DMVR46: partial purification, characterization and application in non-aqueous environment. Process Biochem 49(10):1673–1681Google Scholar
  89. Wang R et al (2015) Enzymatic synthesis of lutein dipalmitate in organic solvents. Catal Lett 145(4):995–999Google Scholar
  90. Wang S, Meng X, Zhou H et al (2016) Enzyme stability and activity in non-aqueous reaction systems: a mini review. Catalysts 6:32Google Scholar
  91. Wen S, Tan T, Zhao H (2013) Improving the thermostability of lipase Lip2 from Yarrowia lipolytica. J Biotechnol 164:248–253Google Scholar
  92. Wenda S, Illner S, Mell A, Kragl U (2011) Industrial biotechnology—the future of green chemistry? Green Chem 13:3007Google Scholar
  93. Wongwatanapaiboon J et al (2016) Cloning, expression, and characterization of Aureobasidium melanogenum lipase in Pichia pastoris. Biosci Biotechnol Biochem 80(11):2231–2240. Google Scholar
  94. Xie C et al (2016) A lipase with broad solvent stability from Burkholderia cepacia RQ3: isolation, characteristics and application for chiral resolution of 1-phenylethanol. Bioprocess Biosyst Eng 39(1):59–66Google Scholar
  95. Xun E, Wang J, Zhang H et al (2013) Resolution of N-hydroxymethyl vince lactam catalyzed by lipase in organic solvent. J Chem Technol Biotechnol 88:904–909. Google Scholar
  96. Yagonia CFJ, Park HJ, Hong SY, Yoo YJ (2015) Simultaneous improvements in the activity and stability of Candida antarctica lipase B through multiple-site mutagenesis. Biotechnol Bioprocess Eng 20:218–224. Google Scholar
  97. Yang S, Zhou L, Tang H et al (2002) Rational design of a more stable penicillin G acylase against organic cosolvent. J Mol Catal B 18:285–290Google Scholar
  98. Yang C, Wang F, Lan D et al (2012) Effects of organic solvents on activity and conformation of recombinant Candida antarctica lipase A produced by Pichia pastoris. Process Biochem 47(3):533–537Google Scholar
  99. Yao C, Cao Y, Wu S et al (2013) An organic solvent and thermally stable lipase from Burkholderia ambifaria YCJ01: purification, characteristics and application for chiral resolution of mandelic acid. J Mol Catal B 85:105–110. Google Scholar
  100. Yedavalli P, Madhusudhana Rao N (2013) Engineering the loops in a lipase for stability in DMSO. Protein Eng Des Sel 26:317–324. Google Scholar
  101. Yuan Z, Bailey TL, Teasdale RD (2005) Prediction of protein B-factor profiles. Proteins Struct Funct Bioinforma 58:905–912. Google Scholar
  102. Zaks A, Klibanov AM (1984) Enzymatic catalysis in organic media at 100 degrees C. Science 224:1249–1251. Google Scholar

Copyright information

© Springer Media B.V. 2018

Authors and Affiliations

  1. 1.Dublin Institute of TechnologyDublin 1Ireland

Personalised recommendations