Abstract
Levan is a homopolymer of fructose with many outstanding properties like high solubility in oil and water, strong adhesiveness, good biocompatibility, and film-forming ability. However, its industrial use has long been hampered by costly production processes which rely on mesophilic bacteria and plants. Recently, Halomonas sp. AAD6 halophilic bacteria were found to be the only extremophilic species producing levan at high titers in semi-chemical medium containing sucrose, and in this study, pretreated sugar beet molasses and starch molasses were both found to be feasible substitutes for sucrose. Five different pretreatment methods and their combinations were applied to both molasses types. Biomass and levan concentrations reached by the Halomonas sp. AAD6 cells cultivated on 30 g/L of pretreated beet molasses were 6.09 g dry cells/L and 12.4 g/L, respectively. When compared with literature, Halomonas sp. was found to stand out with its exceptionally high levan production yields on available fructose. Molecular characterization and monosaccharide composition studies confirmed levan-type fructan structure of the biopolymers. Rheological properties under different conditions pointed to the typical characteristics of low viscosity and pseudoplastic behaviors of the levan polymers. Moreover, levan polymer produced from molasses showed high biocompatibility and affinity with both cancerous and non-cancerous cell lines.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Ahmadi M, Vahabzadeh F, Bonakdarpour B, Mehranian M (2006) Empirical modeling of olive oil mill wastewater treatment using loofa-immobilized Phanerochaete chrysosporium. Process Biochem 41:1148–1154. doi:10.1016/j.procbio.2005.12.012
Bae IY, Oh IK, Lee S, Yoo SH, Lee HG (2008) Rheological characterization of levan polysaccharides from Microbacterium laevaniformans. Int J Biol Macromol 42:10–13. doi:10.1016/j.ijbiomac.2007.08.006
Banik RM, Santhiagu A, Upadhyay SN (2007) Optimization of nutrients for gellan gum production by Sphingomonas paucimobilis ATCC-31461 in molasses based medium using response surface methodology. Bioresour Technol 98(4):792–797. doi:10.1016/j.biortech.2006.03.012
Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254. doi:10.1016/0003-2697(76)90527-3
Dubois MA, Gilles KA, Hamilton JK, Robers PA, Smith F (1956) Colorimetric method for determination of sugars and related substances. Anal Chem 28:350–356. doi:10.1021/ac60111a017
Han YW, Watson MA (1992) Production of microbial levan from sucrose, sugarcane juice and beet molasses. J Ind Microbiol 9:257–260. doi:10.1007/BF01569633
Jiang L, Wang J, Liang S, Wang X, Cen P, Xu Z (2009) Butyric acid fermentation in a fibrous bed bioreactor with immobilized Clostridium tyrobutyricum from cane molasses. Bioresour Technol 100:3403–3409. doi:10.1016/j.biortech.2009.02.032
Kalogiannis SG, Iakovidou M, Liakopoulou-Kyriakides DA, Kyriakidis, Skaracis GN (2003) Optimization of xanthan gum production by Xantomonas campestris grown in molasses. Process Biochem 39:249–256. doi:10.1016/S0032-9592(03)00067-0
Kang SA, Jang KH, Seo JW et al (2009) Levan: applications and perspectives. In: Rehm BHA (ed) Microbial production of biopolymers and polymer precursors. Caister Academic Press, Norwich, pp 145–161
Kundu S, Panda T, Majumdar SK, Guha B, Bandyopadhyay KK (1984) Pretreatment of Indian cane molasses for increased production of citric acid. Biotechnol Bioeng 26:1114–1121. doi:10.1002/bit.260260915
Lee SY, Park SJ, Lee Y, Lee SH (2003) Economic aspects of biopolymer production. In: Steinbuchel A (ed) Biopolymers. VCH-Wiley, Weinheim, pp 307–337
Liu YP, Zheng P, Sun ZH, Ni Y, Dong JJ, Zhu LL (2008) Economical succinic acid production from cane molasses by Actinobacillus succinogenes. Bioresour Technol 99:1736–1742. doi:10.1016/j.biortech.2007.03.044
Liu J, Luo H, Ye Y, Sun Z, Zeng LuX (2010) Medium optimization and structural characterization of exopolysaccharides from endophytic bacterium Paenibacillus polymyxa EJS-3. Carbohydr Polym 79:206–213. doi:10.1016/j.carbpol.2009.07.055
Livingston DP, Hincha DK, Heyer AG (2009) Fructan and its relationship to abiotic stress tolerance in plants. Cell Mol Life Sci 66:2007–2023. doi:10.1007/s00018-009-0002-x
Lotfy WA, Ghanem KM, El-Helow ER (2007) Citric acid production by a novel Aspergillus niger isolate: I. Mutagenesis and cost reduction studies. Bioresour Technol 98:3464–3469. doi:10.1016/j.biortech.2006.11.007
Manca MC, Lama L, Improta R, Esposito E, Gambacorta A, Nicolaus B (1996) Chemical composition of two exopolysaccharides from Bacillus thermoantarcticus. Appl Environ Microbiol 62:3265–3269
Mossman T (1983) Rapid colorimetric assay for cellular growth and survivals: application to proliferation and cytotoxicity assays. J Immunol Methods 65:55–63. doi:10.1016/0022-1759(83)90303-4
Müller B (2009) Impact of the bacterium Pseudomonas fluorescens and its genetic derivatives on vermiculite: effects on trace metals contents and clay mineralogical properties. Geoderma 153:94–103. doi:10.1016/j.geoderma.2009.07.025
Nicolaus B, Kambourova M, Toksoy Öner E (2010) Exopolysaccharides from extremophiles: from fundamentals to biotechnology. Environ Technol 31(10):1145–1158. doi:10.1080/09593330903552094
Oliveira MR, Silva RSSF, Buzato JB, Celligoi MAPC (2007) Study of levan production by Zymomonas mobilis using regional low-cost carbohydrate sources. Biochem Eng J 37:177–183. doi:10.1016/j.bej.2007.04.009
Ozimek LK, Kralj S, van der Maarel MJEC, Dijkhuizen L (2006) The levansucrase and inulosucrase enzymes of Lactobacillus reuteri 121 catalyse processive and non-processive transglycosylation reactions. Microbiology 152:1187–1196. doi:10.1099/mic.0.28484-0
Patidar SK, Tare V (2006) Effect of nutrients on biomass activity in degradation of sulfate laden organics. Process Biochem 41:489–495. doi:10.1016/j.procbio.2005.07.001
Poli A, Kazak H, Gürleyendağ B, Tommonaro G, Pieretti G, Toksoy Öner E, Nicolaus B (2009) High level synthesis of levan by a novel Halomonas species growing on defined media. Carbohydr Polym 78:651–657. doi:10.1016/j.carbpol.2009.05.031
Roukas T (1998) Pretreatment of beet molasses to increase pullulan production. Process Biochem 33:805–810. doi:10.1016/S0032-9592(98)00048-X
Schwanninger M, Rodrigues JC, Pereira H, Hinterstoisser B (2004) Effects of short-time vibratory ball milling on the shape of FT-IR spectra of wood and cellulose. Vib Spectrosc 36:23–40. doi:10.1016/j.vibspec.2004.02.003
Survase SA, Saudagar PS, Singhal RS (2007) Use of complex media for the production of scleroglucan by Sclerotium rolfsii MTCC 2156. Bioresour Technol 98(7):1509–1512. doi:10.1016/j.biortech.2006.05.022
van Hoek P, Aristidou A, Hahn JJ, Patist A (2003) Fermentation goes large scale. Chem Eng Prog 99:37–42
Vedyashkina TA, Revin VV, Gogotov IN (2005) Optimizing the conditions of dextran synthesis by the bacterium Leuconostoc mesenteroides grown in a molasses-containing medium. Appl Biochem Microbiol 41(4):361–364. doi:10.1007/s10438-005-0061-1
Wu M, Wu Y, Zhou J, Pan Y (2009) Structural characterization of a water-soluble polysaccharide with high branches from the leaves of Taxus chinensis var. mairei. Food Chem 113:1020–1024. doi:10.1016/j.foodchem.2008.08.055
Yin X, Calderin L, Stott MJ, Sayer M (2002) Density functional study of structural, electronic and vibrational properties of Mg- and Zn-doped tricalcium phosphate biomaterials. Biomaterials 23:4155–4163. doi:10.1016/S0142-9612(02)00199-0
Yoo SH, Yoon EJ, Cha J, Lee HG (2004) Antitumor activity of levan polysaccharides from selected microorganisms. Int J Biol Macromol 34:37–41. doi:10.1016/j.ijbiomac.2004.01.002
Yoon EJ, Yoo SH, Cha J, Lee HG (2004) Effect of levan’s branching structure on antitumor activity. Int J Biol Macromol 34:191–194. doi:10.1016/j.ijbiomac.2004.04.001
Zhang Y, Zhang Z, Suzuki K, Maekawa T (2003) Uptake and mass balance of trace metals for methane producing bacteria. Biomass Bioenerg 25:427–433. doi:10.1016/S0961-9534(03)00012-6
Acknowledgements
This research has been financially supported by TUBITAK through project no. MAG-108 M193 and by the Ministry of Foreign Affair (MAE, Italy). Fellowship of Faruk Küçükaşık (project no. MAG-108M193) provided by TUBITAK is gratefully acknowledged. Authors thank Bahar Gurleyendag and Dr. Seval Genc for their contribution to the rheological studies. This work is dedicated to Ayse Yarsuvat for her unforgettable support to the IBSB Research Group.
Conflict of interest
The authors declare that they have no conflict of interest.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Küçükaşik, F., Kazak, H., Güney, D. et al. Molasses as fermentation substrate for levan production by Halomonas sp.. Appl Microbiol Biotechnol 89, 1729–1740 (2011). https://doi.org/10.1007/s00253-010-3055-8
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00253-010-3055-8