DOI QR코드

DOI QR Code

Conversion of Ginsenoside Rb1 and Taxonomical Characterization of Stenotrophomonas sp. 4KR4 from Ginseng Rhizosphere Soil

인삼 근권 토양에서 분리한 Stenotrophomonas sp. 4KR4의 Ginsenoside Rb1 전환능 및 분류학적 특성

  • Jeon, In-Hwa (Department of Microbial & Nano Materials, Mokwon University) ;
  • Cho, Geon-Yeong (Department of Microbial & Nano Materials, Mokwon University) ;
  • Han, Song-Ih (Department of Microbial & Nano Materials, Mokwon University) ;
  • Yoo, Sun Kyun (Department of Food Science & Biotechnology, Joongbu University) ;
  • Whang, Kyung-Sook (Department of Microbial & Nano Materials, Mokwon University)
  • 전인화 (목원대학교 미생물나노소재학과) ;
  • 조건영 (목원대학교 미생물나노소재학과) ;
  • 한송이 (목원대학교 미생물나노소재학과) ;
  • 유선균 (중부대학교 식품생명과학과) ;
  • 황경숙 (목원대학교 미생물나노소재학과)
  • Received : 2013.12.02
  • Accepted : 2013.12.19
  • Published : 2013.12.31

Abstract

We isolated the ${\beta}$-glucosidase producing bacteria (BGB) in ginseng root system (rhizosphere soil, rhizoplane, inside of root). Phylogenetic analysis of the 28 BGB based on the 16S rRNA gene sequences, BGB from rhizosphere soil belong to genus Stenotrophomonas (3 strains), Bacillus (1 strain), and Pseudoxanthomonas (1 strain). BGB isolates from rhizoplane were Stenotrophomonas (16 strains), Streptomyces (1 strain) and Microbacterium (1 strain). BGB from inside of root were categorized into Stenotrophomonas (3 strains) and Lysobacter (2 strains). Especially, Stenotrophomonas comprised the largest portion (approximately 90%) of total isolates and Stenotrophomonas was a dominant group of the ${\beta}$-glucosidase producing bacteria. We selected strain 4KR4, which had high ${\beta}$-glucosidase activity (108.17 unit), could transform ginsenoside Rb1 into Rd, Rg3, and Rh2 ginsenosides. In determining its relationship on the basis of 16S rRNA sequence, 4KR4 strain was most closely related to Stenotrophomonas rhizophila e-$p10^T$ (AJ293463) (99.62%). Therefore, on the basis of these polyphasic taxonomic evidence, the ginsenoside Rb1 converting bacteria 4KR4 was identified as Stenotrophomonas sp. 4KR4 (=KACC 17635).

인삼 근계(근권, 근면, 근내부)로부터 ginsenoside Rb1 전환효소인 ${\beta}$-glucosidase 생산 균주(BGB)를 분리하였다. 인삼 근계부터 분리된 BGB 28균주의 계통학적 특성을 확인한 결과, 근권에서 Stenotrophomonas 속(3균주), Pseudoxanthomonas 속(1균주), Bacillus 속(1균주)로 확인되었다. 근면로부터 분리된 BGB는 Stenotrophomonas 속(16균주), Streptomyces 속(1균주), Microbacterium 속(1균주)이며, 근내부는 Stenotrophomonas 속(3균주), Lysobacter 속(2균주)를 포함하는 다양한 계통군이 확인 되었다. 특히 인삼 근계로부터 분리된 BGB 균주의 90%가 Stenotrophomonas 계통군에 속하는 특징을 나타내었다. 근권으로부터 분리된 4KR4 균주는 108.17 unit의 ${\beta}$-glucosidase 활성을 나타내었으며, ginsenoside Rb1을 Rd, Rg3 그리고 minor ginsenoside Rh2로 전환되었다. 4KR4 균주는 Stenotrophomonas rhizophila e-$p10^T$ (AJ293463)와 99.65%의 높은 상동성을 나타내었다. 본 연구에서 분리된 ginsenoside 전환세균 4KR4 균주의 계통학적 위치와 표현형적 특징, 균체 지방산조성, 생리 생화학적 특성을 검토한 결과, Stenotrophomonas sp. 4KR4 (=KACC 17635) 균주로 확인되었다.

Keywords

References

  1. Benishin, C.G. 1992. Actions of ginsenoside Rb1 on choline uptake in central cholinergic nerve endings. Neurochem. Int. 21, 1-5. https://doi.org/10.1016/0197-0186(92)90061-U
  2. Bernardet, J.F., Nakagawa, Y., and Holmes, B. 2002. Proposed minimal standards for describing new taxa of the family Flavobacteriaceae and emended description of the family. Int. J. Syst. Evol. Microbiol. 52, 1049-1070. https://doi.org/10.1099/ijs.0.02136-0
  3. Chen, Y.J., Nose, M., and Ogihara, Y. 1987. Alkaline cleavage of ginsenoside. Chem. Pharm. Bull. 35, 1653-1655 https://doi.org/10.1248/cpb.35.1653
  4. Cheng, L.Q., Na, J.R., Bang, M.H., Kim, M.K., and Yang, D.C. 2008. Conversion of major ginsenoside Rb1 to 20(S)-ginsenoside Rg3 by Microbacterium sp.GS514. Phytochemistry 69, 218-224 https://doi.org/10.1016/j.phytochem.2007.06.035
  5. Cheng, L.Q. 2008. Isolation and characterization of $\beta$-glucosidase active Microbacterium esteraromaticum GS514 and production of 20(S)-ginsenoside Rg3. Unpublished master's thesis, Kyunghee University.
  6. Choi, M.J. 2010. Biotransformation of ginsenosides by Leifsonia sp. GAL45 isolated from ginseng field in Anseong, Korea. Unpublished master's thesis, Chungang University.
  7. Chi, H., Lee, B.H., You, H.J., Park, M.S., and Ji, G.E. 2006. Differential transformation of ginseoside from Panax ginseng by lactic acid bacteria. J. Microbiol. Biotechnol. 16, 1629-1633.
  8. Dong, A.M., Guo, Y.H., Zheng, J., and Guo, D. 2003 Microbial transformation of ginsenoside Rb1 by Rhizopus stolonifer and Curvularia lunata. Biotechnol. Lett. 25, 339-344. https://doi.org/10.1023/A:1022320824000
  9. Finkmann, W., Alterdorf, K., Stackebrandt, E., and Lipski, A. 2000. Characterization of $N_2O$-producing Xanthomonas-like isolates from biofilters as Stenotrophomonas nitritireducens sp. nov., Luteimonas mephitis gen. nov., sp. nov. and Pseudoxanthomonas broegbernensis gen. nov., sp. nov. Int. J. Syst. Evol. Microbiol. 50, 273-282. https://doi.org/10.1099/00207713-50-1-273
  10. Hadden, J.F. and Black, L.L. 1986. The status of ripe rot in Louisiana peppers. National Pepper Conferences. 14.
  11. Han, B.H., Park, M.H., Han, N., Woo, K., Sankawa, U., Yanara, S., and Tanaka, O. 1982. Degradation of ginseng saponins under mild acidic conditions. Planta Med. 44, 146-149. https://doi.org/10.1055/s-2007-971425
  12. Jang, M.H. and Kim, M.D. 2010. Exploration of $\beta$-glucosidase activity of lactic acid bacteria isolated from Kimchi. Food Engineer. Prog. 14, 243-248.
  13. Kikuchi, Y., Sasa, H., Kita, T., Hirata, J., and Tode, T. 1991. Inhibition of human ovarian cancer cell proliferation in vitro by ginsenoside-Rh2 and adjuvant effects of cisplatin in vivo. Anticancer Drugs (England) 2, 63-67. https://doi.org/10.1097/00001813-199102000-00009
  14. Kim, D.J., Seong, G.S., Kim, D.W., Go, S.L., and Jang, J.C. 2004. Antioxidative effects of red ginseng saponins on paraquat-induced oxidative stress. Korean J. Ginseng Res. 28, 5-10. https://doi.org/10.5142/JGR.2004.28.1.005
  15. Kim, D.H. 2012. Isolation and identification of the ginsenoside converting bacteria and analysis of saponin from products. Unpublished master's thesis, Wonkwang University.
  16. Kim, M.K., Lee, J.W., Lee, K.Y., and Yang, D.C. 2005. Microbial conversion of major ginsenoside Rb1 to pharm aceutically activeminor ginsenoside Rd. J. Microbiol. 43, 456-462.
  17. Kim, M.W., Ko, S.R., Choi, K.J., and Kim, S.C. 1987. Distribution of saponin in various sections of Panax ginseng root and changes of its contents according to root age. Korean J. Ginseng Sci. 11, 10-16.
  18. Kim, N.D., Kim, E.M., Kang, K.W., Cho, M.K., Choi, S.Y., and Kim, S.G. 2003. Ginsenoside Rg3 inhibits phenylephrine-induced vascular contraction through induction of nitric oxide synthase. Br. J. Pharmacol. 140, 661-670. https://doi.org/10.1038/sj.bjp.0705490
  19. Kim, S.D. and Seu, J.H. 1982. Conversion of ginseng saponin with the enzyme produced by Rhizopus sp. Kor. J. Appl. Microbiol. Bioeng. 10, 267-273,
  20. Kim, Y.C., Kim, S.R., Markelonis, G.J., and Oh, T.H. 1998. Ginsenoside Rb1 and Rg3 attenuate glutamate-induced neurotoxicity in primary cultures of rat cortical cells. Korean J. Ginseng Sci. 9, 47-56.
  21. Kim, H.G., Kim, K.Y., and Cha, C.J. 2007. Screening for ginsengfermenting microorganisms capable of biotransforming ginsenosides. Kor. J. Microbiology. 43, 142-146.
  22. Kikuchi, Y., Sasa, H., Kita, T., Hirata, J., Tode, T., and Nagata, I. 1991. Inhibition of human ovarian cancer cell proliferation in vitro by ginsenoside Rh2 and adjuvant effects to cisplatin in vivo. Anticancer Drugs (England). 2, 63-67. https://doi.org/10.1097/00001813-199102000-00009
  23. Kitagawa, I., Yoshikawa, M., Yoshihara, M., Hayashi, T., and Taniuama, T. 1983. Chemical studies on crude drug procession. I. On the constituents of ginseng radix rudura (I). Yakugaku Zasshi. 103, 612-622. https://doi.org/10.1248/yakushi1947.103.6_612
  24. Lee, D.S., Kim, Y.S., Ko, C.N., Cho, H.K., Bae, H.S. Lee, K.S., Kim, J.J., Park, E.K., and Kim, D.H. 2002. Fecal metabolic activities of herbal components to bioactive compounds. Arch. Pharm. Res. 25, 165-169. https://doi.org/10.1007/BF02976558
  25. Lee, J.Y. and Hwang, B.K. 2002. Diversity of antifungal actinomycetes in various vegetative soils of Korea. Can. J. Microbiol. 48, 407-417. https://doi.org/10.1139/w02-025
  26. Lee, J.W. and Do, J.H. 2005 Market trend of health functional food and the prospect of ginseng market. J. Ginseng Res. 29, 206-214. https://doi.org/10.5142/JGR.2005.29.4.206
  27. Lee, K.Y., Park, J.A., Chung, E., Lee, H.H., Kim, S.I., and Lee, S.K. 1996. Ginsenoside-Rh2 blocks the cell cycle of SK-HEP-1 cells at the G1/S boundary by selectively inducing the protein expression of p27Kip1. Cancer Lett. 110, 193-200. https://doi.org/10.1016/S0304-3835(96)04502-8
  28. Lee, S.R., Park, H.J., Choi, K.J., and Kim, N.D. 1997. Inhibitory effect of ginsenoside Rg3 on platelet aggregation and its mechanism of action. Kor. J. Ginseng Sci. 21, 132-140.
  29. Mei, B., Wang, Y.E., Wu, J.X., and Chen, W.Z. 1994. Protective effect of ginsenoside on oxygen free radical induced damages of cultured vascular endothelial cells in vitro. Yao. Hsueh Hsuuh Pao. 29, 801-808.
  30. Na, J.R., Kim, Y.J., Kim, S.H., Kim, H.B., Snim, J.S., Kim, S.Y., and Yang, D.C. 2009. Conversion of ginsenoside Rb1 by ginseng soil bacterium Cellulosimicrobium sp. Gsoil 235 according to various culture broths. Kor. J. Microbiol. Biotechnol. 37, 55-61.
  31. Park, J.A., Lee, K.Y., Oh, O.I., and Kim, S.I. 1997. Activation of caspase-3 protease via a Bcl-2-insensitive pathway during the process of ginsenoside Rh2-induced apoptosis. Cancer Lett. 121, 73-81. https://doi.org/10.1016/S0304-3835(97)00333-9
  32. Saito, H. and Nishiyama, N. 1988. Effect of ginseng and its saponins on experimental amnesis in mice and on cell cultures of neurons, pp. 92-98. Proc. 5th Int'l. Ginseng Symp. Seoul, Korea.
  33. Saitou, N. and Nei, N. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol. Biol. Evol. 4, 406-425.
  34. Shibata, S., Tanaka, T., Ando, T., Sado, M., Tsushima, S., and Ohsawa, T. 1966. Chemical studies on oriental plant drugs (XIV). Protopanaxadial, a genuine sapogenin of ginseng saponins. Chem. Pharm. Bull. 14, 595-600. https://doi.org/10.1248/cpb.14.595
  35. Singh, V.K., Agarwal, S.S., and Gupta, B.M. 1984. Immunomodulatory activity of Panax ginseng extract, pp. 225-232. Proc. 4th Int'l. Ginseng Symp. Seoul, Korea.
  36. Wang, L., An, D.S., Kim, S.G., Jin, F.X., Kim, S.C., Lee, S.T., and Im, W.T. 2012. Ramilbacter ginsenosidimutans sp. nov., with ginsenoside converting activity. J. Microbial. Biotechnol. 22, 311-315. https://doi.org/10.4014/jmb.1106.06041
  37. Wolf, A., Fritze, A., Hagemann, M., and Berg, G. 2002. Stenotrophomonas rhizophila sp. nov., a novel plant-associated bacterium with antifungal properties. Int. J. Syst. Evol. Microbiol. 52, 1937-1944. https://doi.org/10.1099/ijs.0.02135-0
  38. Yeom, J.H. 2010. Conversion of ginsenoside $Rb_1$ by bacterium isolated from ginseng cultivation soil. Unpublished master's thesis, Joongbu University.
  39. Yi, H., Srinivasan, S., and Kim, M.K. 2010. Stenotrophomonas panacihumi sp. nov., isolated from soil of a ginseng field. J. Microbiol. 48, 30-35. https://doi.org/10.1007/s12275-010-0006-0