Skip to main content
Log in

Enzymatic clarification of fruit juices (Apple, Pineapple, and Tomato) using purified Bacillus pumilus SV-85S xylanase

  • Research Paper
  • Published:
Biotechnology and Bioprocess Engineering Aims and scope Submit manuscript

Abstract

The xylanase obtained from a hyper-producer Bacillus pumilus SV-85S was purified and characterized to evaluate its potential in industrial applications. Xylanase was purified to homogeneity 25.3-fold with 63.2% recovery using cation-exchange chromatography through CMSephadex C-50. The purified xylanase showed a single band on Native-PAGE and a single peak in RP-HPLC confirming its homogeneity. The purified enzyme revealed a single band on SDS-PAGE with a molecular mass of 23.6 kDa, which was confirmed with gel filtration chromatography through Sepharose 6B. The Km and Vmax of the purified xylanase was 1.0 mg/mL and 333.3 IU/mL, respectively. The temperature and pH profiles of the purified xylanase revealed that it was thermo and alkali stable. In recent years due to the overall increase in natural fruit juice consumption, juices have become important from a consumption point of view. However, raw juice is turbid and viscous which tends to settle during storage. Therefore, it must be clarified before commercialization. The efficacy of absolutely purified xylanase was studied on juice enrichment of apples (Malus domestica), pineapples (Ananas comosus L.) and tomatos (Lycopersicum esculentum). The treatment with xylanase lead to an increased juice yield by 23.53% (apple), 10.78% (pineapple), and 20.78% (tomato) as well as having a significant effect on juice clarity by an increase of % transmittance of 22.20, 19.80, and 14.30, respectively. The turbidity and viscosity was also decreased without affecting acid neutrality significantly.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  1. Goulart, A. J., E. C. Carmona, and R. Monti (2005) Partial purification and properties of cellulase-free alkaline xylanase produced by Rhizopus stolonifer in solid-state fermentation. Braz. Arch. Biol. Technol. 48: 327–333.

    CAS  Google Scholar 

  2. Badhan, A. K., B. S. Chadha, J. Kaur, H. S. Saini, and M. K. Bhat (2007) Production of multiple xylanolytic and cellulolytic enzymes by thermophilic fungus Myceliophthora sp. IMI 387099. Biores. Technol. 98: 504–510.

    Article  CAS  Google Scholar 

  3. Polizeli, M. L. T. M., A. C. S. Rizzatti, R. Monti, H. F. Terenzi, J. A. Jorge, and D. S. Amori (2005) Xylanases from fungi: Properties and industrial applications. Appl. Microbiol. Biotechnol. 67: 577–591.

    Article  CAS  Google Scholar 

  4. Dhiman, S. S., J. Sharma, and B. Battan (2008) Industrial applications and future prospects of microbial xylanases: A review. BioResou. 3: 1377–1402.

    Google Scholar 

  5. Ninawe, S., M. Kapoor, and R. C. Kuhad (2007) Purification and characterization of extracellular xylanase from Streptomyces cyaneus SN32. Bioresour. Technol. 99: 1252–1258.

    Article  Google Scholar 

  6. Nagar, S., A. Mittal, D. Kumar, and V. K. Gupta (2012) Production of alkali tolerant cellulase free xylanase in high levels by Bacillus pumilus SV-205. Int. J. Biol. Macromol. 50: 412–420.

    Article  Google Scholar 

  7. Lee, W. C., S. Yusof, N. S. A. Hamid, and B. S. Baharin (2006) Optimizing conditions for enzymatic clarification of banana juice using response surface methodology (RSM). J. Food Engg. 73: 55–63.

    Article  CAS  Google Scholar 

  8. Bhat, M. K. (2000) Cellulases and related enzymes in biotechnology. Biotechnol. Advan. 18: 355–383.

    Article  CAS  Google Scholar 

  9. Sims, C. A. and P. Bates (1994) Challenges to processing tropical fruit juices: Banana as an example. Proc. Florida State Horticul. Soc. 107: 315–319.

    Google Scholar 

  10. Dawes, H., P. Struebi, and J. Keene (1994) Kiwifruit juice clarification using a fungal proteolytic enzyme. J. Food Sci. 59: 858–861.

    Article  CAS  Google Scholar 

  11. Dhiman, S. S., G. Sharma, J. Mahajan, and R. Methoxy (2011) Characterization of statistically produced xylanase for enrichment of fruit juice clarification process. New Biotechnol. 28: 746–755.

    Article  CAS  Google Scholar 

  12. Voragen, A. G. J., R. Heutink, and W. Pilnik (1980) Solubilization of apple cell walls with polysaccharide degrading enzymes. J. Appl. Biochem. 2: 452–468.

    CAS  Google Scholar 

  13. Pal, A. and F. Khanum (2011) Efficacy of xylanase purified from Aspergillus niger DFR-5 alone and in combination with pectinase and cellulose to improve yield and clarity of pineapple juice. J. Food Sci. Technol. 48: 560–568.

    Article  CAS  Google Scholar 

  14. Grassin, C. and P. Fauquembergue (1996) Application of pectinases in beverages. Prog. Biotechnol. 14: 453–462.

    Article  CAS  Google Scholar 

  15. Nagar, S., A. Mittal, D. Kumar, L. Kumar, R. C. Kuhad, and V. K. Gupta (2011) Hyper production of alkali stable xylanase in lesser duration by Bacillus pumilus SV-85S using wheat bran under solid state fermentation. New Biotechnol. 28: 581–587.

    Article  CAS  Google Scholar 

  16. Miller, L. G. (1959) Use of dinitrosalycilic acid reagent for determination of reducing sugar. Anal. Chem. 31: 426–428.

    Article  CAS  Google Scholar 

  17. Lowry, O. H., N. J. Rosebrough, A. L. Farr, and R. J. Randall (1951) Protein measurement with the Folin Phenol reagent. J. Biological. Chem. 193: 265–275.

    CAS  Google Scholar 

  18. Davis, B. J. (1964) Disc electrophoresis. II. Method and application to human serum proteins. Ann. N. Y. Acad. Sci. 121: 404–427.

    Article  CAS  Google Scholar 

  19. Laemmli, U. K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685.

    Article  CAS  Google Scholar 

  20. Bailey, M. J., P. Biely, and K. Poutanen (1992) Laboratory testing of method for assay of xylanase activity. J. Biotechnol. 23: 257–270.

    Article  CAS  Google Scholar 

  21. Archana, A. and T. Satyanarayana (2003) Purification and characterization of cellulase-free xylanase of a moderate thermophile Bacillus licheniformis A99. World J. Microbiol. Biotechnol. 19: 53–57.

    Article  CAS  Google Scholar 

  22. Sanghi, A., N. Garg, V. K. Gupta, A. Mittal, and R. C. Kuhad (2010) One-step purification and characterization of cellulasefree xylanase produced by alkalophilic Bacillus subtilis ASH. Braz. J. Microbiol. 41: 467–476.

    Article  CAS  Google Scholar 

  23. Qureshy, A. F., L. A. Khan, and S. Khanna (2002) Cloning, regulation and purification of cellulase-free xylanase from Bacillus circulans Teri-42. Ind. J. Microbiol. 42: 35–41.

    Google Scholar 

  24. Heck, J. X., S. H. Flores, P. F. Hertz, and M. A. Z. Ayub (2006) Statistical optimization of thermo-tolerant xylanase activity from Amazon isolated Bacillus circulans on solid state cultivation. Bioresour. Technol. 97: 1902–1906.

    Article  CAS  Google Scholar 

  25. Kiddinamoorthy, J., Anceno, A. J., G. D. Haki, and S. K. Rakshit (2008) Production, purification and characterization of Bacillus sp. GRE7 xylanase and its application in eucalyptus Kraft pulp biobleaching. World J. Microbiol. Biotechnol. 24: 605–612.

    Article  CAS  Google Scholar 

  26. Maalej, I., I. Belhaj, N. F. Masmoudi, and H. Belghith (2009) Highly thermostable xylanase of the thermophilic fungus Talaromyces thermophilus: Purification and characterization. Appl. Biochem. Biotechnol. 158: 200–212.

    Article  CAS  Google Scholar 

  27. Lin, J., L. M. Ndlovu, S. Singh, and B. Pillay (1999) Purification and biochemical characteristics of α-D-xylanase from thermophilic fungus, Thermomyces lanuginosus-SSBP. Biotechnol. Appl. Biochem. 30: 73–79.

    CAS  Google Scholar 

  28. Bakir, U., S. Yavascaoglu, F. Guvenc, and A. Ersayin (2001) An endo-β-1,4-xylanase from Rhizopus oryzae: Production, partial purification and biochemical characterization. Enz. Microb. Technol. 29: 328–334.

    Article  CAS  Google Scholar 

  29. Jiang, Z. Q., W. Deng, X. T. Li, Z. L. Ai, L.T. Li, and I. Kusakabe (2005) Characterization of a novel, ultra-large xylanolytic complex (xylanosome) from Streptomyces olivaceoviridis E-86. Enz. Microb. Technol. 36: 923–929.

    Article  CAS  Google Scholar 

  30. Li, X. T., Z. Q. Jiang, L.T. Li, S. Q. Yang, W. Y. Feng, J. Y. Fan, and I. Kusakabe (2005) Charcterization of a cellulase-free, natural xylanase from Thermomyces lanuginosus CBS and its biobleaching effect on wheat straw pulp. Enz. Microbiol. Biotechnol. 96: 1370–1379.

    CAS  Google Scholar 

  31. Torronen, A., R. L. Mach, R. Messner, R. Gonzalez, N. Kalkkinen, A. Harkki, and C. P. Kubicek (1992) The two major xylanases from Trichoderma reesei: Characterization of both enzymes and genes. Bio-Technol. 10: 1461–1465.

    CAS  Google Scholar 

  32. Driss, D., F. Bhiri, L. Elleucha, N. Bouly, I. Stals, N. Miled, M. Blibecha, R. Ghorbela, and S. E. Chaabouni (2011) Purification and properties of an extracellular acidophilic endo-1,4-β-xylanase, naturally deleted in the “thumb”, from Penicillium occitanis Pol6. Proc. Biochem. 46: 1299–1306.

    Article  CAS  Google Scholar 

  33. Khendeparker, R. D. S. and N. B. Bhosle (2006) Isolation, purification and characterization of the xylanase produced by Arthrobacter sp. MTCC 5214 when grown in solid state fermentation. Enz. Microb. Technol. 39: 732–742.

    Article  Google Scholar 

  34. Gupta, S., B. Bhushan, and G. S. Hoondal (2000) Isolation, purification and characterization of xylanase from Staphylococcus sp. SG-13 and its application in biobleaching of kraft pulp. J. Appl. Microbiol. 88: 325–333.

    Article  CAS  Google Scholar 

  35. Sin, H. N., S. Yusof, N. S. A. Hamid, and R. Abdul Rahman (2006) Optimization of enzymatic clarification of sapodilla juice using response surface methodology. J. Food Eng. 73: 313–319.

    Article  CAS  Google Scholar 

  36. Ahmad, I., Y. K. Jha, and R. K. Anurag (2009) Optimization of enzymic extraction process forhigher yield and clarity of guava juice. J. Food Sci. Technol. 46: 307–311.

    CAS  Google Scholar 

  37. Olfa, E., M. Mondher, S. Issam, L. Ferid, and M. M. Nejib (2007) Induction, properties and application of xylanase activity from Sclerotinia sclerotiorum S2 fungus. J. Food Biochem. 31: 1096–10735.

    Article  Google Scholar 

  38. Abdullah, A.G., N. M. Sulaiman, M. K. Aroua, and M. J. Megat Mohd (2007) Response surface optimization of conditions for clarification of carambola fruit juice using a commercial enzyme. J. Food Eng. 81: 65–71.

    Article  CAS  Google Scholar 

  39. Carvalho, L. M. J., C. A. B. Silva, and A. P. T. R. Pierucci (1998) Clarification of pineapple juice (Ananas comosus L. Merryl) by ultrafiltration and microfiltration: Physicochemical evaluation of clarified juices, soft drink formulation, and sensorial evaluation. J. Agri. Food Chem. 46: 2185–2189.

    Article  Google Scholar 

  40. Jacob, N., R. K. Sukumaran, and P. Prema (2008) Optimization of enzymatic clarification of sapodilla juice: A statistical perspective. Appl. Biochem. Biotechnol. 151: 353–363.

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Vijay Kumar Gupta.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Nagar, S., Mittal, A. & Gupta, V.K. Enzymatic clarification of fruit juices (Apple, Pineapple, and Tomato) using purified Bacillus pumilus SV-85S xylanase. Biotechnol Bioproc E 17, 1165–1175 (2012). https://doi.org/10.1007/s12257-012-0375-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12257-012-0375-9

Keywords

Navigation