Abstract
Fructooligosaccharides and levan have a wide range of applications in the food industry due to their physiological and functional properties. The enzymatic synthesis of these molecules exhibits great advantages when compared with microbial fermentation. In this study, the production of levansucrase from Bacillus subtilis natto and its utilization in fructooligosaccharides and levan syntheses using different reaction conditions were described. The best condition for levansucrase production was 420.7 g L−1 of sucrose at pH 7.0, which reached 23.9 U ml−1 of transfructosylation activity. In a bioreactor, the highest production of fructooligosaccharides was 41.3 g L−1 using a medium containing 350 g L−1 sucrose at 35 °C for 36 h. The enzymatic synthesis of levan resulted in 86.9 g L−1 when conditions similar to those used for fructooligosaccharides synthesis were applied. These results indicate that the levansucrase from B. subtilis natto could be applied for the co-production of fructooligosaccharides and levan, which are biomolecules that have health benefits and are used successfully in the food industry.
![](http://media.springernature.com/m312/springer-static/image/art%3A10.1007%2Fs12010-017-2587-0/MediaObjects/12010_2017_2587_Fig1_HTML.gif)
![](http://media.springernature.com/m312/springer-static/image/art%3A10.1007%2Fs12010-017-2587-0/MediaObjects/12010_2017_2587_Fig2_HTML.gif)
Similar content being viewed by others
References
Cantarel, B. L., Coutinho, P. M., Rancurel, C., Bernard, T., Lombard, V., & Henrissat, B. (2009). The carbohydrate active enzymes database (CAZy): an expert resource for Glycogenomics. Nucleic Acids Research, 37, 233–238.
Wuerges, J., Caputi, L., Cianci, M., Boivin, S., Meijers, R., & Benini, S. (2015). The crystal structure of Erwinia amylovora levansucrase provides a snapshot of the products of sucrose hydrolysis trapped into the active site. Journal of Structural Biology, 190, 290–298.
Guio, F., Rugeles, L. D., Rojas, S. E., Palomino, M. P., Camargo, M. C., & Sánchez, O. F. (2012). Kinetic modeling of fructooligosaccharide production using Aspergillus oryzae N74. Applied Biochemistry and Biotechnology, 167, 142–163.
Linde, D., Rodríguez-Colinas, B., Estévez, M., Poveda, A., Plou, F. J., & Fernández Lobato, M. (2012). Analysis of neofructooligosaccharides production mediated by the extracellular β-fructofuranosidase from Xanthophyllomyces dendrorhous. Bioresource Technology, 109, 123–130.
Santos-Moriano, P., Fernandez-Arrojo, L., Poveda, A., Jimenez-Barbero, J., Ballesteros, A. O., & Plou, F. J. (2015). Levan versus fructooligosaccharide synthesis using the levansucrase from Zymomonas mobilis: effect of reaction conditions. Journal of Molecular Catalysis B: Enzymatic, 119, 18–25.
Srikanth, R., Reddy, C. H. S. S. S., Siddartha, G., Ramaiah, M. J., & Uppuluri, K. B. (2015). Review on production , characterization and applications of microbial levan. Carbohydrate Polymers, 120, 102–114.
Shih, I. L., Chen, L. D., Wang, T. C., Wu, J. Y., & Liaw, K. S. (2010). Tandem production of levan and ethanol by microbial fermentation. Green Chemistry, 12, 1242–1247.
Franken, J., Brandt, B. A., Tai, S. L., & Bauer, F. F. (2013). Biosynthesis of levan, a bacterial extracellular polysaccharide, in the yeast Saccharomyces cerevisiae. PloS One, 8, 1–14.
Mussatto, S. I., & Mancilha, I. M. (2007). Non-digestible oligosaccharides: a review. Carbohydrate Polymers, 68, 587–597.
Roberfroid, M. (2007). Prebiotics : the concept revisited. Journal of Nutrition, 137, 830–837.
Saad, N., Delattre, C., Urdaci, M., Schmitter, J. M., & Bressollier, P. (2013). An overview of the last advances in probiotic and prebiotic field. LWT - Food Science and Technology, 50, 1–16.
Hangit, Y. W., & Clarke, M. A. (1990). Production and characterization of microbial levan. Advances in Applied Microbiology, 35, 393–396.
Jang, K., Song, K., Kim, C. H., Chung, B. H., Kang, S. A., Choue, R. W., & Rhee, S. (2001). Comparison of characteristics of levan produced by different preparations of levansucrase from Zymomonas mobilis. Enzyme and Microbial Technology, 19, 339–344.
Yun, J. W. (1996). Fructooligosaccharides—occurrence, preparation, and application. Enzyme and Microbial Technology, 19, 107–117.
Shih, I.-L., Chen, L.-D., & Wu, J.-Y. (2010). Levan production using Bacillus subtilis natto cells immobilized on alginate. Carbohydrate Polymers, 82, 111–117.
Calazans, G., Lima, C., França, F. P., & Lopes, C. E. (2000). Molecular weight and antitumour activity of Zymomonas mobilis levans. International Journal of Biological Macromolecules, 27, 245–247.
Silva, P., Borsato, D., & Celligoi, M. A. P. C. (2014). High production of fructooligosaccharides by levansucrase from Bacillus subtilis natto CCT 7712. African Journal of Biotechnology, 13, 2734–2740.
Berté, S. D., Borsato, D., Silva, P. B., Vignoli, J. A., & Celligoi, M. A. P. C. (2013). Statistical optimization of levansucrase production from Bacillus subtilis ATCC 6633 using response surface methodology. African Journal of Microbiology Research, 7, 898–904.
Somogyi, M. (1945). A new reagent for the determination of sugars. The Journal of Biological Chemistry, 160, 61–68.
Nelson, N. (1944). A photometric adaptation of the somogyi method for the determination of glucose. The Journal of Biological Chemistry, 153, 375–380.
Ananthalakshmy, V. K., & Gunasekaran, P. (1999). Isolation and characterization of mutants from levan-producing Zymomonas mobilis. Journal of Bioscience and Bioengineering, 87, 214–217.
Laemmli, U. K. (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, 227, 680–685.
Sangeetha, P. T., Ramesh, M. N., & Prapulla, S. G. (2004). Production of fructo-oligosaccharides by fructosyl transferase from Aspergillus oryzae CFR 202 and Aureobasidium pullulans CFR 77. Process Biochemistry, 39, 755–760.
Bersaneti, G. T., Mantovan, J., Magri, A., Mali, S., & Celligoi, M. A. P. C. (2016). Edible films based on cassava starch and fructooligosaccharides produced by Bacillus subtilis natto CCT 7712. Carbohydrate Polymers, 151, 1132–1138.
Viikari, L., & Gisler, R. (1986). By-products in the fermentation of sucrose by different Zymomonas strains. Applied Microbiology and Biotechnology, 23, 240–244.
S R Core Team. R: a language and environment for statistical computing. R Foundation for Statistical Computing,Vienna, Austria, 2016. Available from: https://www.R-project.org/.
Shapiro, S. S., & Wilk, M. B. (1965). An analysis of variance test for normality (complete samples). Biometrika, 52, 591.
Breusch, T. S., & Pagan, A. R. (1979). A simple test for heteroscedasticity and random coefficient variation. Econometrica, 47, 1287–1294.
Inthanavong, L., Tian, F., Khodadadi, M., & Karboune, S. (2013). Properties of Geobacillus stearothermophilus levansucrase as potential biocatalyst for the synthesis of levan and fructooligosaccharides. Biotechnology Progress, 29, 1405–1415.
Abdel-Fattah, A. F., Mahmoud, D. a. R., & Esawy, M. a. T. (2005). Production of levansucrase from Bacillus subtilis NRC 33a and enzymic synthesis of levan and fructo-oligosaccharides. Current Microbiology, 51, 402–407.
Belghith, K. S., Dahech, I., Belghith, H., & Mejdoub, H. (2012). Microbial production of levansucrase for synthesis of fructooligosaccharides and levan. International Journal of Biological Macromolecules, 50, 451–458.
Silva, P. B., Borsato, D., & Celligoi, M. A. P. C. (2014). Optimization of high production of fructooligosacharides by sucrose fermentation of Bacillus subtilis natto CCT 7712. American Journal of Food Technology, 9, 144–150.
Khandekar, D. C., Palai, T., Agarwal, A., & Bhattacharya, P. K. (2014). Kinetics of sucrose conversion to fructooligosaccharides using enzyme (invertase) under free condition. Bioprocess and Biosystems Engineering, 37, 2529–2537.
Romano, N., Santos, M., Mobili, P., Vega, R., & Gómez-zavaglia, A. (2016). Effect of sucrose concentration on the composition of enzymatically synthesized short-chain fructooligosaccharides as determined by FTIR and multivariate analysis. Food Chemistry, 202, 467–475.
Mehmood, A., Abdallah, K., Khandekar, S., Zhurina, D., Srivastava, A., Al-karablieh, N., Alfaro-espinoza, D., Pletzer, M. S., & Ullrich, M. S. (2015). Expression of extra-cellular levansucrase in Pseudomonas syringae is controlled by the in planta fitness-promoting metabolic repressor. HexR, BMC - Microbiology, 15, 1–11.
Bakar, B., & Kaplan-Turkoz, B. (2017). Structural modelling and structure-function analysis of Zymomonas mobilis levansucrase. Journal of Natural and Applied Sciences, 21, 279–285.
Risso, F. V. A., Mazutti, M. A., Treichel, H., Costa, F., Maugeri, F., & Rodrigues, M. I. (2012). Comparison between systems for synthesis of fructooligosaccharides from sucrose using free inulinase from Kluyveromyces marxianus NRRL Y-7571. Food and Bioprocess Technology, 5, 331–337.
Wu, F., Chou, S., & Shih, I. (2013). Factors affecting the production and molecular weight of levan of Bacillus subtilis natto in batch and fed-batch culture in fermenter. Journal of the Taiwan Institute of Chemical Engineers, 615, 01–08.
Acknowledgements
The authors would like to thank the Coordination for the Improvement of Higher Education Personnel (CAPES-Brazil) and the Araucaria Foundation for the financial support.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of Interest
The authors declare that they have no conflict of interest.
Ethical Statement
This article does not contain any studies with human participants or animals performed by any of the authors.
Rights and permissions
About this article
Cite this article
Bersaneti, G.T., Pan, N.C., Baldo, C. et al. Co-production of Fructooligosaccharides and Levan by Levansucrase from Bacillus subtilis natto with Potential Application in the Food Industry. Appl Biochem Biotechnol 184, 838–851 (2018). https://doi.org/10.1007/s12010-017-2587-0
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12010-017-2587-0