Korean Journal of Environmental Biology (환경생물)

Korean Society of Environmental Biology (한국환경생물학회)
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Purification and Characterization of Bioflocculant Producing from $Lactobacillus$ $jensenii$ YW-33

$Lactobacillus$ $jensenii$ YW-33이 생산하는 생물응집제의 정제 및 특성

  • Seo, Ho-Chan (Department of Brain Education, University of Brain Education)
  • 서호찬 (국제뇌교육종합대학원대학교 뇌교육학과)
  • Received : 2011.10.31
  • Accepted : 2011.11.11
  • Published : 2011.11.30
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Abstract

The distribution of flocculating activity of culture broth was examined and the major constituent with flocculating activity was identified. Most of flocculating activity was found in culture broth without cells. As the activity was maintained by the digestion with pronase, it suggests that the activity is due to the polysaccharide. The bioflocculant obtained from $Lactobacillus$ $jensenii$ YW-33 was precipitated by 60~80% EtOH fractionation (LJ-80). LJ-80 was separated by ion-exchange chromatography using DEAE-Toyopearl 650C and LJ-80-II showed more potent flocculating activity than those of other fractions. The major activity fraction LJ-80-II was further purified on the gel permeation using Sepharose CL-6B to LJ-80-II-1. GPC (Sepharose CL-6B) and HPLC were used to determine whether LJ-80-II-1 has a homogenecity. The molecular weight of purified LJ-80-II-1 was estimated over 800,000 dalton by gel permeation chromatography. Purified LJ-80-II-1 contained 98.4% total sugar, 0.6% protein. Main sugar of purified LJ-80-II-1 was composed of mannose : galactose : glucose with a molar ratio of 1.61 : 0.25 : 1.00.

$Lactobacillus$ $jensenii$ YW-33이 생산하는 생물응집제의 활성본체를 규명하고자 응집물질을 분리, 정제하고 그 특성을 검토하고자 하였다. 생물응집제의 활성 본체를 조사하기 위하여 배양액을 원심분리하여 균체 자체와 균체를 제거한 상등액을 비교한 결과 균체를 제거한 상등액이 86%로 높은 응집활성을 나타냈다. 또한 pronase 처리와 periodate 산화를 행한 결과 pronase로 처리한 시료는 무처리군과 비교하여 차이가 없었던 반면 periodate로 산화시킨 시료는 응집활성이 크게 감소함에 따라 다당에 기인되는 것으로 추정되었다. 균체가 제거된 상등액을 감압, 농축하여 EtOH 60~80%의 농도로 분획한 결과 LJ-80을 얻었다. 분리된 LJ-80을 DEAE-Toyopearl 650C chromatography, Sepharose CL-6B chromatography을 이용하여 최종 정제된 LJ-80-II-1을 분리하였으며 HPLC 분석에 의해 순도를 확인했다. 정제된 LJ-80-II-1의 분자량은 약 800,000 이상이며 총당이 98.4%, 단백질이 0.6%으로 mannose : galactose : glucose가 1.61 : 0.25 : 1.00의 molecule ratio을 가지고 있었다.

Keywords

References

  1. 서호찬, 최양문, 조홍연, 양한철. 1997. Lactobacillus jensenii YW-33이 생산하는 다당류성 생물응집제 및 생산조건. 한국산업미생물학회지. 25:328-334.
  2. Ander BN and SO Enfors. 1982. Production of extracellular polysaccharide by Zoogloea ramigera. Appl. Environ. Microbiol. 44:1231-1237.
  3. Brierly CL, DP Kelly, KJ Sea and DJ Best. 1985. In Biotechnology, pp. 186-204, Blackwell Scientific Pub., Oxford.
  4. Cripps RE and W Elizabeth. 1967. The accumlation of extracellular macromolecules by Staphylococcus aureus growth in the presence of sodium chloride and glucose. J. Gen. Microbiol. 49:127. https://doi.org/10.1099/00221287-49-1-127
  5. Crueger W and A Crueger. 1984. Biotechnology : Textbook of Industrial Microbiology. pp. 288-291. Science Tech. Inc. Madison.
  6. Deavin L, TR Jarman, CJ Lawson, RC Righelato and S Slocomb. 1977. The production of alginic acid by Azotobacter vinelandii in batch and continous culture. American Chemical Society Symp. Ser. 45:14-26.
  7. Desmond PF, F Auer and RJ Seviour. 1990. Influence of varying nitrogen sources on polysaccharide production by Aureobasidum pillulans in batch culture. Appl. Microbiol. Biotechnol. 32:63-637.
  8. Dubios M, KA Hamilton, TK Rebers and F Sonisth. 1956. Colorimetric method for determination of sugar and related substances. Anal. Chem. 28:350. https://doi.org/10.1021/ac60111a017
  9. Ikeda FH, ST Fukui and T Tomita. 1982. An extracellular polysaccharide produced by Zoogloea ramigera 115. Eur. J. Biochem. 123:437-445. https://doi.org/10.1111/j.1432-1033.1982.tb19787.x
  10. Jarman TR, L Deavin, S Slocombe and RC Righelato. 1978. Investigation of the effect of environmental conditions on the rate of exopolysaccharide synthesis in Azotobacter vinelandii. J. Gen. Microbiol. 107:59-64. https://doi.org/10.1099/00221287-107-1-59
  11. Jochen B and M Nava. 1988. Cell adsorption control by culture conditions. Appl. Microbiol. Biotechnol. 29:119. https://doi.org/10.1007/BF00939295
  12. Kazuo S and H Takahashi. 1981. DNA as a flocculation factor in Pseudomonas sp. Agric. Biol. Chem. 45:2869. https://doi.org/10.1271/bbb1961.45.2869
  13. Koizumi JI and M Takeda. 1991. Synergistic Flocculation of the Bioflocculant fix extracellulary produced by Norcadia Amara. J. Gen. Appl. Microbiol. 37:447. https://doi.org/10.2323/jgam.37.447
  14. Kurane R. 1994. Production of a bioflocculant by Rhodococcus erythropolis S-1 Grown on alcohol. Biosci. Biotech. Biochem. 58:428. https://doi.org/10.1271/bbb.58.428
  15. Kurane R. 1994. Purification and Characterization of Lipid Bioflocculant produced by Rhodococcus erythropolis. Biosci. Biotech. Biochem. 58:1977. https://doi.org/10.1271/bbb.58.1977
  16. Kurane RK, K Toeda, K Takeda and T Suzuki. 1986. Cultural conditions for production of microbial flocculant by Rhodococcus erythropolis. Agr. Biol. Chem. 50:2309-2313. https://doi.org/10.1271/bbb1961.50.2309
  17. Lowry OH, NJ Rosebrough, K Farr and RJ Rindall. 1951. Protein measurement with the folin phenol reagent. J. Biol. Chem. 193:256.
  18. McNeil B and B Kristiansen. 1990. Temperature effects on polysaccharide formation by Aureobasidium pullulans in stirred tanks. Enzyme Micro. Technol. 12:521-526. https://doi.org/10.1016/0141-0229(90)90069-3
  19. Miki BL, AN Hung, A Poon, P James and VL Seligy. 1992. Possible mechanism for flocculation interaction governed by gene FLO I in Saccharomyces cerevisiae. J. Bacteriol. 150:890-899.
  20. Nakamura J, S Miyashiro and Y Hirose. 1976. Screening, isolation and some properties of microbial cell flocculants. Agr. Biol. Chem. 40:377-382. https://doi.org/10.1271/bbb1961.40.377
  21. Nakamura J, S Myashiro and Y Hirose. 1976. Conditions for production of polysaccharide from Pullularia pullulans. Appl. Microbiol. 25:628-635.
  22. Norberg AB and S Enfors. 1982. Production of extracellular polysaccharide by Zoogloea ramigera. Appl. Environ. Microbiol. 44:1231-1237.
  23. Parker ND and CB Munn. 1984. Increased cell surface hydrophibicity associated with possesion of an additional surface protein by Aeromonas salmonicida. FEMS Microbiology Letters 21:233. https://doi.org/10.1111/j.1574-6968.1984.tb00216.x
  24. Souw P and AL Demain. 1979. Nutritional studies on xanthan production by Xanthomonas campestris NRRL B1459. Appl. Environ. Microbiol. 37:1186-1192.
  25. Suzuki K, N Nakano, Y Nagatomi, H Tominaga, N Nakazono, M Itai, M Uyeda and M Shibata. 1990. HAF hepatoma aggregation factor produced by Streptomyces sp. strain No. A-6143. Agric. Biol. Chem. 54:2061-2068. https://doi.org/10.1271/bbb1961.54.2061
  26. Takagi H and K Kadowaki. 1985. Flocculant production by Paccilomyces sp. Taxonomic studies and culture conditions for production. Agr. Bio. Chem. 49:3151-3157. https://doi.org/10.1271/bbb1961.49.3151
  27. Takagi H and K Kadowki. 1985. Purification and chemical properties of a flocculant produced by Paecilomyces sp., Agr. Biol. Chem. 49:3159-3164. https://doi.org/10.1271/bbb1961.49.3159
  28. Toeda K and R Kurane. 1991. Microbial flocculant from Alcaligenes cupidus KT201. Agr. Biol. Chem. 55:2793-2799. https://doi.org/10.1271/bbb1961.55.2793
  29. Yamada H, H Kiyohara, JC Cyong, Y Kojima, Y Kumazawa and Y Otsuka. 1984. Studies of polysaccharide from Angleica acutiloba. part I. Fractionation and biological properties of polysaccharides. Planta Med. 50:163-167. https://doi.org/10.1055/s-2007-969661
  30. Zajic JE and A Leduy. 1973. Flocculant and chemical properties of a microbial cell flocculant by Aspergillus sojae AJ7002. Agr. Biol. Chem. 40:1341-1347.