Enhanced Biosurfactant Production by Bacillus pumilus 2IR in Fed-Batch Fermentation Using 5-L Bioreactor

  • Tayebeh FooladiEmail author
  • Peyman Abdeshahian
  • Nasrin Moazami
  • Mohammad Reza Soudi
  • Abudukeremu Kadier
  • Wan Mohtar Wan Yusoff
  • Aidil Abdul Hamid
Research Paper


The enhancement of the production of lipopeptide biosurfactant by Bacillus pumilus 2IR as an oil field-isolated bacterium was investigated in the fed-batch fermentation using a 5-L bioreactor. The initial study on the culture medium composition used for biosurfactant synthesis revealed that biosurfactant production was greatly affected by glucose, crude oil, potassium nitrate and ammonium sulfate as pivotal carbon and nitrogen sources which were utilized for biosurfactant synthesis in the bioreactor. The results obtained from the batch fermentation process in the bioreactor showed that the quantities of biomass and biosurfactant produced were 4.15 and 0.98 g/L, respectively. Similar results revealed that fed-batch fermentation led to the production of 5.71 g/L biomass and 1.06 g/L biosurfactant in the bioreactor.


Biosurfactant Fed-batch fermentation Bacillus pumilus 2IR Oil field Bioreactor 



The authors sincerely acknowledge University Kebangsaan Malaysia (National University of Malaysia).


  1. Abdel-Mawgoud AM, Aboulwafa MM, Haleem NA (2008) Optimization of surfactin production by Bacillus subtilis Isolate BS5. Appl Biochem Biotechnol 150:305–325CrossRefGoogle Scholar
  2. Abouseoud M, Maachi R, Amrane A, Boudergua S, Nabi A (2008) Evaluation of different carbon and nitrogen sources in production of biosurfactant by Pseudomonas fluorescens. Desalination 223:143–151CrossRefGoogle Scholar
  3. Amani H, Mehrnia MR, Sarrafzadeh MH, Haghighi M, Soudi MR (2010) Scale up and application of biosurfactant from Bacillus subtilis in enhanced oil recovery. Appl Biochem Biotechnol 162:510–523CrossRefGoogle Scholar
  4. Amin GA (2014) Exponential the fed-batch strategy for enhancing biosurfactant production by Bacillus subtilis. Water Sci Technol 70:234–240CrossRefGoogle Scholar
  5. Amodu OS, Ntwampe SKO, Ojumu TV (2014) Optimization of biosurfactant production by Bacillus licheniformis STK 01 grown exclusively on Beta vulgaris waste using response surface methodology. BioResour 9:5045–5065Google Scholar
  6. Aparna AG, Srinikethan SH (2012) Isolation, screening and production of biosurfactant by Bacillus clausii 5B. Res Biotechnol 3:49–56Google Scholar
  7. Armstrong RT, Wildenschild DB (2015) The effect of pore morphology on microbial enhanced oil recovery. J Pet Sci Eng 130:16–25CrossRefGoogle Scholar
  8. Avili MG, Fazaelipoor MH, Jafari SA, Ataei SA (2012) Comparison between batch and the fed-batch production of rhamnolipid by Pseudomonas aeruginosa. Iran J Biotechnol 10(4):263–269Google Scholar
  9. Bence K (2011) Bacterial production of antimicrobial biosurfactant by Bacillus subtilis. PhD thesis, University of Stellenbosch Biotechnology 150: 305–325Google Scholar
  10. Chenikher S, Guez JS, Coutte F, Pekpe M, Jacques P, Cassar JP (2010) Control of the specific growth rate of Bacillus subtilis for the production of biosurfactant lipopeptides in bioreactors with foam overflow. Process Biochem 45:1800–1807CrossRefGoogle Scholar
  11. Das K, Mukherjee AK (2007) Crude petroleum-oil biodegradation efficiency of Bacillus subtilis and Pseudomonas aeruginosa strains isolated from petroleum oil contaminated soil from north-east India. Bioresour Technol 98:1339–1345CrossRefGoogle Scholar
  12. Ebadi A, Olamaee M, Sima NAK, Nasrabadi RG, Hashemi M (2017) Isolation and characterization of biosurfactant producing and crude oil degrading bacteria from oil contaminated soils. Iran J Sci Technol Trans Sci. Google Scholar
  13. Elazzazy AM, Abdelmoneim TS, Almaghrabi OA (2015) Isolation and characterization of biosurfactant production under extreme environmental conditions by alkali-halothermophilic bacteria from Saudi Arabia. Saudi J Biol Sci 22:466–475CrossRefGoogle Scholar
  14. Fooladi T, Moazami N, Abdeshahian P, Kadier A, Ghojavand H, Yusoff WWM, Hamid AA (2016) Characterization, production and optimization of lipopeptide biosurfactant by new strain Bacillus pumilus 2IR isolated from an Iranian oil field. J Pet Sci Eng 145:510–519CrossRefGoogle Scholar
  15. Frederico KA, Cristiano BP, Denise F (2010) The fed-batch biosurfactant production in a bioreactor. Int Rev Chem Eng 2:513Google Scholar
  16. Gaytán I, Mejía MA, Hernández-Gama R, Torres LG, Escalante CA (2015) Effects of indigenous microbial consortia for enhanced oil recovery in a fragmented calcite rocks system. J Pet Sci Eng 128:65–72CrossRefGoogle Scholar
  17. Ghojavand H, Vahabzadeh F, Roayaei E, Shahraki AK (2008) Production and properties of a biosurfactant obtained from a member of the Bacillus subtilis group (PTCC 1696). J Colloid Inter Sci 324:172–176CrossRefGoogle Scholar
  18. Ghojavand H, Vahabzadeh F, Shahraki AK (2012) Enhanced oil recovery from low permeability dolomite cores using biosurfactant produced by a Bacillus mojavensis (PTCC 1696) isolated from Masjed-I Soleyman field. J Pet Sci Eng 8:24–30CrossRefGoogle Scholar
  19. Haddad NIA, Wang J, Bozhong M (2009) Identification of a biosurfactant producing strain: Bacillus subtilis HOB2. Protein Pept Lett 16:7–137CrossRefGoogle Scholar
  20. Joshi S, Bharucha C, Jha S, Yadav S, Nerurkar A, Desai AJ (2008) Biosurfactant production using molasses and whey under thermophilic conditions. Bioresour Technol 99:195–199CrossRefGoogle Scholar
  21. Joshi-Navare K, Prabhune A (2013) A biosurfactant-sophorolipid acts in synergy with antibiotics to enhance their efficiency. Biomed Res Int. Google Scholar
  22. Khopade A, Biao R, Liu X, Mahadik K, Zhang L, Kokare C (2012a) Production and stability studies of the biosurfactant isolated from marine Nocardiopsis sp. B4. Desalination 285:198–204CrossRefGoogle Scholar
  23. Khopade A, Liu XY, Mahadik K, Zhang L, Kokare C (2012b) Production and characterization of biosurfactant from marine Streptomyces species B3. J Colloid Inter Sci 367:311–318CrossRefGoogle Scholar
  24. Kiran SG, Thomas AT, Selvin J, Sabarathuhan B, Lipton AP (2010) Optimization and characterization of a new lipopeptide biosurfactant produced by marine Brevibacterium aureum MSA13 in solid state culture. Bioresour Technol 101:2389–2396CrossRefGoogle Scholar
  25. Ławniczak L, Marecik R, Chrzanowski Ł (2013) Contributions of biosurfactants to natural or induced bioremediation. Appl Microbiol Biotechnol 97:2327–2339CrossRefGoogle Scholar
  26. Lee MK, Hwang SH, Ha SD, Jang JH, Lim DJ, Kong JY (2004) Rhamnolipid production in batch and the fed-batch fermentation using Pseudomonas aeruginosa BYK-2 KCTC 18012P. Biotechnol Bioprocess Eng 9:267–273CrossRefGoogle Scholar
  27. Lotfabad TB, Shourian M, Roostaazad R, Rouholamini AN, Adelzadeh MR, Akbari Noghabi K (2009) An efficient biosurfactant producing bacterium Pseudomonas aeruginosa MR01, isolated from oil excavation areas in south of Iran. Colloid Surf B Biointer 69:183–193CrossRefGoogle Scholar
  28. Makkar RS, Cameotra SS (2002) An update on the use of unconventional substrates for biosurfactant production and their new applications. Appl Microbiol Biotechnol 58:428–434CrossRefGoogle Scholar
  29. Matar SM, El-Kazzaz SA, Wagih EE, El-Diwany AI, Moustafa HE, El-Saadani MA, Abo-Zaid GA, Hafez EE (2009) Bioprocessing and scaling-up cultivation of Bacillus subtilis as a potential antagonist to certain plant pathogenic fungi, III. Biotechnol 8:138–143CrossRefGoogle Scholar
  30. Mukherjee S, Das P, Sivapathasekaran C, Sen R (2008) Enhanced production of biosurfactant by a marine bacterium on statistical screening of nutritional parameters. Biochem Eng J 42:254–260CrossRefGoogle Scholar
  31. Mulligan CN (2004) Environmental applications for biosurfactants. Environ Pollut 133:183–198CrossRefGoogle Scholar
  32. Najafi AR, Rahimpour MR, Jahanmiri AH, Roostaazad R, Arabian D, Soleimani M, Jamshidnejad Z (2011) Interactive optimization of biosurfactant production by Paeni bacillus alvei ARN63 isolated from an Iranian oil well. Colloids Surf B: Biointer 82:33CrossRefGoogle Scholar
  33. Nitschke M, Pastore GM (2006) Production and properties of a surfactant obtained from Bacillus subtilis grown on cassava wastewater. Bioresour Technol 97:336–341CrossRefGoogle Scholar
  34. Nur AMN, Salwa MS, Amirul AA, Ahmad RM (2012) The fed-batch cultivation of Pseudomonas aeruginosa USM-AR2 Producing rhamnolipid in bioreactor through pulse feeding strategy. Int Conf Biol Life Sci 40:45Google Scholar
  35. Oh JS, Kim BG, Park TH (2002) Importance of specific growth rate for subtilisin expression in the fed-batch cultivation of Bacillus subtilis spoIIG mutant. Enzy Microbiol Technol 30:747–751CrossRefGoogle Scholar
  36. Pacwa M, Płaza GA, Piotrowska SZ, Cameotra SS (2011) Environmental applications of biosurfactants: recent advances. Inter J Mol Sci 12:633–654CrossRefGoogle Scholar
  37. Patel J, Borgohain S, Kumar M, Rangarajan V, Somasundaran P, Sen R (2015) Recent developments in microbial enhanced oil recovery. Renew Sustain Energy Rev 52:1539–1558CrossRefGoogle Scholar
  38. Pornsunthorntawee O, Wongpanit P, Chavadej S, Abe M, Rujiravanit R (2008) Structural and physicochemical characterization of crude biosurfactant produced by Pseudomonas aeruginosa SP4 isolated from petroleum-contaminated soil. Bioresour Technol 99:1589–1595CrossRefGoogle Scholar
  39. Rahman PKSM, Gakpe E (2008) Production, characterization and application of biosurfactants—review. Biotechnol J 7:360–370CrossRefGoogle Scholar
  40. Samsu ZA, Yusof Z, Awang MS, Noh NM, Yahya ARM (2014) The fed-batch production of valuable biosurfactant, rhamnolipid, from waste cooking oil by indigenously isolate Pseudomonas aeruginosa USM-AR2. Adv Environ Biol 8:33–38Google Scholar
  41. Sarafin Y, Donio MBS, Velmurugan S, Michaelbabu M, Citarasu T (2014) Kocuria marina BS- 15 a biosurfactant producing halophilic bacteria isolated from solar salt works in India. Saudi J Biol Sci 21:511–519CrossRefGoogle Scholar
  42. Sarafzadeh P, Niazi A, Oboodi V, Ravanbakhsh M, Hezave AZ, Ayatollahi SS, Raeissi S (2014) Investigating the efficiency of MEOR processes using Enterobacter cloacae and Bacillus stearothermophilus SUCPM#14 (biosurfactant-producing strains) in carbonated reservoirs. J Pet Sci Eng 113:46–53CrossRefGoogle Scholar
  43. Sari M, Kusharyoto W, Artika IM (2014) Screening for biosurfactant-producing yeast: confirmation of biosurfactant production. Biotechnol 13:106–111CrossRefGoogle Scholar
  44. Sharma D, Saharan BS, Chauhan N, Bansal A, Procha S (2014) Production and structural characterization of Lactobacillus helveticus derived biosurfactant. Sci World J. Google Scholar
  45. Sivapathasekaran C, Sen R (2013) Performance evaluation of batch and unsteady state fed- batch reactor operations for the production of a marine microbial surfactant. J Chem Technol Biotechnol 88:719–726CrossRefGoogle Scholar
  46. Snehal RM, Bhalchandra KV, Renuka MJ, Kiran MD, Sanjay NN (2008) Use of response surface optimization for the production of biosurfactant from Rhodococcus spp. MTCC 2574. Bioresour Technol 99:7875–7880CrossRefGoogle Scholar
  47. Vaz DA, Eduardo J, Gudina EJA, Jose A, Teixeira LRR (2012) Performance of a biosurfactant produced by a Bacillus subtilis strain isolated from crude oil samples as compared to commercial chemical surfactants. Coll Surf B: Biointerface 89:167–174CrossRefGoogle Scholar
  48. White DA, Hird LC, Ali SD (2013) Production and characterization of a trehalolipid biosurfactant produced by the novel marine bacterium Rhodococcus sp., strain PML026. J Appl Microbiol 115:744–755CrossRefGoogle Scholar
  49. Youssef N, Duncan K, Nagle D, Kristen N, Savageroy MK, McInerney MJ (2004) Comparison of methods to detect biosurfactant production by diverse microorganisms. J Microbiol Method 56:339–347CrossRefGoogle Scholar
  50. Youssef N, Simpsona DR, McInerneya MJ, Duncan KE (2013) In-situ lipopeptide biosurfactant production by Bacillus strains correlates with improved oil recovery in two oil wells approaching their economic limit of production. Int Biodeterior Biodegrad 81:127–132CrossRefGoogle Scholar

Copyright information

© Shiraz University 2018

Authors and Affiliations

  1. 1.Department of Microbiology, Faculty of Biological SciencesAlzahra UniversityTehranIran
  2. 2.Department of Veterinary Medicine, Faculty of Agriculture, Shoushtar BranchIslamic Azad UniversityShoushtarIran
  3. 3.Biotechnology Center Iranian Research Organization for Science and Technology (IROST)TehranIran
  4. 4.Department of Chemical and Process Engineering, Faculty of Engineering and Built EnvironmentNational University of Malaysia (Universiti Kebangsaan Malaysia)BangiMalaysia
  5. 5.School of Biosciences and Biotechnology, Faculty of Science and TechnologyUniversiti Kebangsaan Malaysia (UKM)BangiMalaysia

Personalised recommendations