Abstract
In this research, single-factor and response surface experiments were conducted in the fed-batch fermentation process to improve the yield of iturin A. The effect of adding various concentrations of precursor amino acids l-asparagine (Asn), l-aspartic acid (Asp), l-glutamic acid (Glu), l-glutamine (Gln), l-Serine (Ser) and l-proline (Pro) at different adding times (3 and 12 h) on iturin A production and cell growth was studied. The respective addition of amino acids (Asp 0.28 g/L; Asn 0.36 g/L; Glu 0.20, 0.28 and 0.360 g/L; Gln 0.20, 0.28 and 0.36 g/L; Pro 0.12, 0.20, 0.28 and 0.36 g/L) at 3 h was shown to improve cell growth but did not affect the yield of iturin A. Meanwhile, the individual addition of the same amino acids at 12 h improved cell growth and increased the yield of iturin A. Excellent correlation was obtained between the predicted and measured values, suggesting that the regression model was accurate and reliable; highly significant (P < 0.0001), and the determination coefficient (R2 = 0.975). When 0.0752 g/L Asn; 0.1992 g/L Gln and 0.1464 g/L Pro were added at 12 h, the yield of iturin A reached 0.85 g/L, which is 32.81%-fold higher than that of the initial process. Therefore, this study obtained optimal parameters for iturin A production by the experimental method, and process validation gave high iturin A yields (0.85 g/L) during a 60 h fermentation. These findings could guide an up-scaling of the fermentation process.
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source was soybean protein (total nitrogen concentration 4.0 g/L). The fermentation conditions were as follows: initial pH 8.0, temperature 30 °C, rotating speed 150 rpm, fermentation time 60 h. The samples were taken at 0, 3, 6, 9, 12, 15, 18, 21, 24, 28, 32, 36, 42, 48, 54 and 60 h, and stored at −20 °C
source was soybean protein (total nitrogen concentration 4.0 g/L). The fermentation conditions were as follows: initial pH 8.0, temperature 30℃, rotating speed 150 rpm, fermentation time 60 h. The samples were taken at 0, 3, 6, 9, 12, 15, 18, 21, 24, 28, 32, 36, 42, 48, 54 and 60 h, and stored at −20 °C
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Aharonowitz Y (1980) Nitrogen metabolite regulation of antibiotic biosynthesis. Annu Rev Microbiol 34:209. https://doi.org/10.1146/annurev.mi.34.100180.001233
Ambati P, Ayyanna C (2001) Optimizing medium constituents and fermentation conditions for citric acid production from palmyra jaggery using response surface method. World J Microb Biot 17:331–335. https://doi.org/10.1023/A:1016613322396
Besson F, Peypoux F, Michel G, Delcambe L (1978) Identification of antibiotics of iturin group in various strains of Bacillus subtilis. J Antibiot 31:284–288. https://doi.org/10.7164/antibiotics.31.284
Besson F, Chevanet C, Michel G (1987) Influence of the culture medium on the production of iturin A by Bacillus subtilis. J Gen Microbiol 133:767–772. https://doi.org/10.1099/00221287-133-3-767
Cho SJ, Lee SK, Cha BJ, Kim YH, Shin KS (2003) Detection and characterization of the Gloeosporium gloeosporioides growth inhibitory compound iturin A from Bacillus subtilis strain KS03. FEMS Microbiol Lett 223:47–51. https://doi.org/10.1016/S0378-1097(03)00329-X
Das P, Mukherjee S, Sen R (2008) Genetic regulations of the biosynthesis of microbial surfactants: an overview. Biotechnol Genet Eng 25:165–185. https://doi.org/10.5661/bger-25-165
Fravel DR (2005) Commercialization and implementation of biocontrol. Annu Rev Phytopathol 43:337–359. https://doi.org/10.1146/annurev.phyto.43.032904.092924
Gong M, Wang JD, Zhang J, Yang H, Lu XF, Pei Y, Cheng JQ (2006) Study of the antifungal ability of Bacillus subtilis strain PY-1 in vitro and identification of its antifungal substance (iturin A). Acta Biochem Biophys Sin 38:233–240. https://doi.org/10.1111/j.1745-7270.2006.00157.x
Jin H et al (2014) Direct bio-utilization of untreated rapeseed meal for effective Iturin A production by Bacillus subtilis in submerged fermentation. PLoS ONE 9:e111171. https://doi.org/10.1371/journal.pone.0111171
Jin H, Li KP, Niu YX, Guo MA, Hu CJ, Chen SW, Huang FH (2015) Continuous enhancement of iturin A production by Bacillus subtilis with a stepwise two-stage glucose feeding strategy. BMC Biotechnol 15:53. https://doi.org/10.1186/s12896-015-0172-6
Kim PI, Ryu J, Kim YH, Chl YT (2010) Production of biosurfactant lipopeptides Iturin A, Fengycin, and Surfactin A from Bacillus subtilis CMB32 for control of Colletotrichum gloeosporioides. J Microbiol Biotechn 20:138–145. https://doi.org/10.4014/jmb.0905.05007
Klich MA, Lax AR, Bland JM (1991) Inhibition of some mycotoxigenic fungi by Iturin-A, a peptidolipid produced by Bacillus subtilis. Mycopathologia 116:77–80. https://doi.org/10.1007/Bf00436368
Larsson DGJ (2014) Antibiotics in the environment Upsala. J Med Sci 119:108–112. https://doi.org/10.3109/03009734.2014.896438
Li XH, Zhang YZ, Wei ZW, Guan ZB, Cai YJ, Liao XR (2016) Antifungal activity of isolated Bacillus amyloliquefaciens SYBC H47 for the biocontrol of Peach Gummosis. PLoS ONE 11:e0162125. https://doi.org/10.1371/journal.pone.0162125
Liao JH, Chen PY, Yang YL, Kan SC, Hsieh FC, Liu YC (2016) Clarification of the antagonistic effect of the lipopeptides produced by Bacillus amyloliquefaciens BPD1 against Pyricularia oryzae via in situ MALDI-TOF IMS analysis. Molecules 21:1670. https://doi.org/10.3390/molecules21121670
Lin HY, Rao YK, Wu WS, Tzeng YM (2008) Ferrous ion enhanced lipopeptide antibiotic iturin A production from Bacillus amyloliquefaciens B128. Int J Appl Sci Eng 5:123–132
Magetdana R, Ptak M, Peypoux F, Michel G (1985) Pore-forming properties of Iturin-A, a lipopeptide antibiotic. Biochim BiophysActa 815:405–409. https://doi.org/10.1016/0005-2736(85)90367-0
Magetdana R, Harnois I, Ptak M (1989) Interactions of the lipopeptide antifungal Iturin-A with lipids in mixed monolayers. Biochem Biophys Acta 981:309–314. https://doi.org/10.1016/0005-2736(89)90042-4
Martinez JL (2009) Environmental pollution by antibiotics and by antibiotic resistance determinants. Environ Pollut 157:2893–2902. https://doi.org/10.1016/j.envpol.2009.05.051
Meena KR, Kanwar SS (2015) Lipopeptides as the antifungal and antibacterial agents: applications in food safety and therapeutics. Biomed Res Int. https://doi.org/10.1155/2015/473050
Miller GL, Blum R, Glennon WE, Burton AL (1960) Measurement of carboxymethylcellulase activity. Anal Biochem 1:127–132. https://doi.org/10.1016/0003-2697(60)90004-X
Ongena M, Jacques P (2008) Bacillus lipopeptides: versatile weapons for plant disease biocontrol. Trends Microbiol 16:115–125. https://doi.org/10.1016/j.tim.2007.12.009
Peng WJ et al (2014) The artificial neural network approach based on uniform design to optimize the fed-batch fermentation condition: application to the production of iturin A. Microb Cell Fact. https://doi.org/10.1186/1475-2859-13-54
Perez-Garcia A, Romero D, de Vicente A (2011) Plant protection and growth stimulation by microorganisms: biotechnological applications of Bacilli in agriculture. Curr Opin Biotechnol 22:187–193. https://doi.org/10.1016/j.copbio.2010.12.003
Peypoux F, Guinand M, Michel G, Delcambe L, Das BC, Lederer E (1978) Structure Of Iturine-A, a peptidolipid antibiotic from Bacillus subtilis. Biochemistry 17:3992–3996. https://doi.org/10.1021/bi00612a018
Phister TG, O’Sullivan DJ, McKay LL (2004) Identification of bacilysin, chlorotetaine, and iturin A produced by Bacillus sp. strain CS93 isolated from pozol, a Mexican fermented maize dough. Appl Environ Microb 70:631–634. https://doi.org/10.1128/Aem.70.1.631-634.2004
Ratnam BVV, Rao MN, Rao MD, Rao SS, Ayyanna C (2003) Optimization of fermentation conditions for the production of ethanol from sago starch using response surface methodology. World J Microb Biot 19:523–526. https://doi.org/10.1023/A:1025174731814
Stein T (2005) Bacillus subtilis antibiotics: structures, syntheses and specific functions. Mol Microbiol 56:845–857. https://doi.org/10.1111/j.1365-2958.2005.04587.x
Tan XU, Ren YL, Zhong J, Tan H (2012) Influence of amino acid addition on Jiean-peptide fermentation. Guangdong Agric Sci 11:109–113
Trupkin S, Levin L, Forchiassin F, Viale A (2003) Optimization of a culture medium for ligninolytic enzyme production and synthetic dye decolorization using response surface methodology. J Ind Microbiol Biot 30:682–690. https://doi.org/10.1007/s10295-003-0099-0
Wei YH, Chu IM (1998) Enhancement of surfactin production in iron-enriched media by Bacillus subtilis ATCC 21332. Enzyme Microb Tech 22:724–728. https://doi.org/10.1016/S0141-0229(98)00016-7
Wei YH, Chu IM (2002) Mn2+ improves surfactin production by Bacillus subtilis. Biotechnol Lett 24:479–482. https://doi.org/10.1023/A:1014534021276
Wu JY, Liao JH, Shieh CJ, Hsieh FC, Liu YC (2018) Kinetic analysis on precursors for iturin A production from Bacillus amyloliquefaciens BPD1. J Biosci Bioeng 126:630–635. https://doi.org/10.1016/j.jbiosc.2018.05.002
Xu H, Sun LP, Shi YZ, Wu YH, Zhang B, Zhao DQ (2008) Optimization of cultivation conditions for extracellular polysaccharide and mycelium biomass by Morchella esculenta As51620. Biochem Eng J 39:66–73. https://doi.org/10.1016/j.bej.2007.08.013
Yao DH, Ji ZX, Wang CJ, Qi GF, Zhang LL, Ma X, Chen SW (2012) Co-producing iturin A and poly-gamma-glutamic acid from rapeseed meal under solid state fermentation by the newly isolated Bacillus subtilis strain 3–10. World J Microb Biot 28:985–991. https://doi.org/10.1007/s11274-011-0896-y
Yu GY, Sinclair JB, Hartman GL, Bertagnolli BL (2002) Production of iturin A by Bacillus amyloliquefaciens suppressing Rhizoctonia solani. Soil Biol Biochem 34:955–963. https://doi.org/10.1016/S0038-0717(02)00027-5
Acknowledgements
The authors thank the Chinese Academy of Sciences (CAS) Light of West China Program (Grant No. 2017XBZG_XBQNXZ_B_006), the Key Laboratory of Environmental and Applied Microbiology of the Chengdu Institute of Biology CAS (grant No. KLCAS-2018-3) and Innovation Academy for Seed Design CAS for financial support of this work.
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HY carried out the experiments. HY and JZ established the feeding method. HY, JY, DL and JZ participated in the Separation and extraction of iturin A. HY, ZL, DS and HW participated in the statistical analysis. HT conceived of the study, and participated in the design and coordination and helped to draft the manuscript. All authors read and approved the final manuscript.
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Yue, H., Zhong, J., Li, Z. et al. Optimization of iturin A production from Bacillus subtilis ZK-H2 in submerge fermentation by response surface methodology. 3 Biotech 11, 36 (2021). https://doi.org/10.1007/s13205-020-02540-7
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DOI: https://doi.org/10.1007/s13205-020-02540-7