Pilot-scale Purification of Lipopeptide from Marine-derived Bacillus marinus
(2.Shanghai Zeyuan Marine Biotechnology Co., Ltd., Shanghai, China 200237)
【Abstract】This research was aimed at establishing the pilot-scale purification technology of lipopeptide from marine-derived Bacillus marinus. We studied lipopeptide surfactivity interferences on scale-up unit technologies including acid precipitation, methanol extraction, solvent precipitation, salting out, extraction, silica gel column chromatography and HZ806 macroporous absorption resin column chromatography. Then, the unit technologies were combined in a certain order, to remove the impurities gradually, and to gain purified lipopeptide finally, with high recovery rate throughout the whole process. The novel pilot-scale purification technology could effectively isolate and purify lipopeptide with 87.51% to 100% purity in hectograms from 1 ton of Bacillus marinus B-9987 fermentation broth with more than 81.73% recovery rate. The first practical hectogram production of highly purified lipopeptide derived from Bacillus marinus was achieved. With this new purification method, using complex media became possible in fermentation process to reduce the fermentation cost and scale-up the purification for lipopeptide production. For practicability and economy, foaming problem resulting from massive water evaporation was avoided in this technology.
【Keywords】 Bacillus marinus; lipopeptide; pilot-scale purification; hectogram; scale-up;
(Translated by SONG Junfeng)
Arima K, Kakinuma A, Tamura G. Surfactin, a crystalline peptidelipid surfactant produced by Bacillus subtilis: isolation, characterization and its inhibition of fibrin clot formation. Biochem Biophys Res Commun, 1968, 31(3): 488-494.
LüYN, Yang SZ, Mu BZ. Progress of lipopeptide biosurfactants. Biotechnol Bull, 2004(6): 11-16(in Chinese).
Deleu M, Paquot M. From renewable vegetables resources to microorganisms: new trends in surfactants. C R Chim, 2004, 7(6/7): 641-646.
Lai CC, Huang YC, Wei YH, et al. Biosurfactantenhanced removal of total petroleum hydrocarbons from contaminated soil. J Hazard Mater, 2009, 167(1/3): 609-614.
Scheibenbogen K, Zytner RG, Lee H, et al. Enhanced removal of selected hydrocarbons from soil by Pseudomonas aeruginosa UG2 biosurfactants and some chemical surfactants. J Chem Technol Biotechnol, 1994, 59(1): 53-59.
Brown MJ. Biosurfactants for cosmetic applications. Int J Cosmet Sci, 1991, 13(2): 61-64.
Gandhimathi R, Seghal Kiran G, Hema TA, et al. Production and characterization of lipopeptide biosurfactant by a sponge-associated marine actinomycetes Nocardiopsis alba MSA10. Bioprocess Biosyst Eng, 2009, 32(6): 825-835.
Donio MBS, Ronica SFA, Thanga Viji V, et al. Isolation and characterization of halophilic Bacillus sp. BS3 able to produce pharmacologically important biosurfactants. Asian Pac J Trop Med, 2013, 6(11): 876-883.
Gong QW, Zhang C, Lu FX, et al. Identification of bacillomycin D from Bacillus subtilis fmb J and its inhibition effects against Aspergillus flavus. Food Control, 2014, 36(1): 8-14.
Saravanakumari P, Mani K. Structural characterization of a novel xylolipid biosurfactant from Lactococcus lactis and analysis of antibacterial activity against multi-drug resistant pathogens. Bioresour Technol, 2010, 101(22): 8851-8854.
Pabel CT, Vater J, Wilde C, et al. Antimicrobial activities and matrix-assisted laser desorption/ionization mass spectrometry of Bacillus isolates from the marine sponge Aplysina aerophoba. Mar Biotechnol, 2003, 5(5): 424-434.
Shaala LA, Youssef DTA, Mc Phail KL, et al. Malyngamide 4, a new lipopeptide from the Red Sea marine cyanobacterium Moorea producens(formerly Lyngbya majuscula). Phytochem Lett, 2013, 6(2): 183-188.
Hajare SN, Subramanian M, Gautam S, et al. Induction of apoptosis in human cancer cells by a Bacillus lipopeptide bacillomycin D. Biochimie, 2013, 95(9): 1722-1731.
Baltz RH, Miao V, Wrigley SK. Natural products to drugs: daptomycin and related lipopeptide antibiotics. Nat Prod Rep, 2005, 22(6): 717-741.
Ongena M, Jacques P. Bacillus lipopeptides: versatile weapons for plant disease biocontrol. Trends Microbiol, 2008, 16(3): 115-125.
Liu RF, Zhang DJ, Li YG, et al. A new antifungal cyclic lipopeptide from Bacillus marinus B-9987. Helv Chim Acta, 2010, 93(12): 2419-2425.
Zhang DJ, Liu RF, Li YG, et al. Two new antifungal cyclic lipopeptides from Bacillus marinus B-9987. Chem Pharm Bull, 2010, 58(12): 1630-1634.
Ongena M, Jourdan E, Adam A, et al. Surfactin and fengycin lipopeptides of Bacillus subtilis as elicitors of induced systemic resistance in plants. Environ Microbiol, 2007, 9(4): 1084-1090.
Mnif I, Elleuch M, Chaabouni SE, et al. Bacillus subtilis SPB1 biosurfactant: production optimization and insecticidal activity against the carob moth Ectomyelois ceratoniae. Crop Protect, 2013, 50: 66-72.
Yánez-Mendizábal V, Zeriouh H, Viñas I, et al. Biological control of peach brown rot(Monilinia spp. )by Bacillus subtilis CPA-8 is based on production of fengycin-like lipopeptides. Eur JPlant Pathol, 2012, 132(4): 609-619.
Zhao ZZ, Wang QS, Wang KM, et al. Study of the antifungal activity of Bacillus vallismortis ZZ185 in vitro and identification of its antifungal components. Bioresour Technol, 2010, 101(1): 292-297.
Moyne AL, Shelby R, Cleveland TE, et al. Bacillomycin D: an iturin with antifungal activity against Aspergillus flavus. J Appl Microbiol, 200190(4): 622-629.
Kelleher TJ, Lai JJ, Decourcey JP, et al. Process for the purification of daptomycin: EP, 1586580. 2009-11-25.
Wang ZG, Wang WY, Wu M, et al. Purification of daptomycin from fermentation broth of Streptomyces roseosporus by anion-exchange resin Pharm Clin Res, 2011, 19(4): 318-321(in Chinese).
Dhanarajan G, Rangarajan V, Sen R. Dual gradient macroporous resin column chromatography for concurrent separation and purification of three families of marine bacterial lipopeptides from cell free broth. Sep Purif Technol, 2015, 143: 72-79.
Zhang DL, Wang YF, Lu YJ, et al. An efficient method for separation of surfactin from Bacillus amyloliquefaciens fmb50 broth by flocculation. Process Biochem, 2014, 49(7): 1182-1188.
Chen HL, Chen YS, Juang RS. Separation of surfactin from fermentation broths by acid precipitation and two-stage dead-end ultrafiltration processes. J Membr Sci, 2007, 299(1/2): 114-121.
Yu JT, Tang XX, Wu XY, et al. 新编生物工艺学(上册). Beijing: Chemical Industry Press, 2003: 99-100(in Chinese).
Gao W, Tian L, Zhou JY, et al. Antifungal mechanism of Bacillus marinus B-9987. Acta Microbiol Sin, 2009, 49(11): 1494-1501(in Chinese).
Chen J, Wei HG, Luo YC, et al. Medium optimization for the production of new antifungl cyclic lipopeptide marinhysin A by Bacillus marinus B-9987. China Biotechnol, 2013, 33(1): 84-89(in Chinese).
Bezza FA, Chirwa EMN. Production and applications of lipopeptide biosurfactant for bioremediation and oil recovery by Bacillus subtilis CN2. Biochem Eng J, 2015, 101: 168-178.
de Sousa T, Bhosle S. Isolation and characterization of a lipopeptide bioemulsifier produced by Pseudomonas nitroreducens TSB. MJ10 isolated from a mangrove ecosystem. Bioresour Technol, 2012, 123: 256-262.
Ibrahim ML, Ijah UJJ, Manga SB, et al. Production and partial characterization of biosurfactant produced by crude oil degrading bacteria. Int Biodeterior Biodegrad, 2013, 81: 28-34.
Zhang XW, Yin QX, Cui PL, et al. Correlation of solubilities of hydrophilic pharmaceuticals versus dielectric constants of binary solvents. Ind Eng Chem Res, 2012, 51(19): 6933-6938.
Yu JT, Tang XX, Wu XY, et al. 新编生物工艺学(下册). Beijing: Chemical Industry Press, 2003: 37-42, 145-153(in Chinese).
Snyder LR. Classification off the solvent properties of common liquids. J Chromatogr Sci, 1978, 16(6): 223-234.
Zhang ZM, Wang SJ, Yu SY, et al. GB 15670-1995农药登记毒理学试验方法. Beijing: China Standards Press, 1996: 207-241(in Chinese).
Abdel-Mawgoud AM, Aboulwafa MM, Hassouna NAH. Optimization of surfactin production by Bacillus subtilis isolate BS5. Appl Biochem Biotechnol, 2008, 150(3): 305-325.
Wei YH, Wang LF, Chang JS. Optimizing iron supplement strategies for enhanced surfactin production with Bacillus subtilis. Biotechnol Prog, 2004, 20(3): 979-983.