Applied Biological Chemistry

The Korean Society for Applied Biological Chemistry (한국응용생명화학회)
권호 표지

Isolation and Identification of Inhibitory Compound from Crataegi Fructus on ${\alpha}-amylase$ and ${\alpha}-glucosidase$

산사(Crataegi Fructus) 추출물로부터 ${\alpha}-amylase$와 ${\alpha}-glucosidase$ 저해 물질 분리 및 동정

  • Published : 2007.09.30
Download PDF
⟨ Previous Next ⟩

Abstract

The objective of this research was to evaluate the inhibitory activities of phenolic compounds isolated from Crataegi Fructus on ${\alpha}-amylase$ and ${\alpha}-glucosidase$. The content of total phenolic compounds of water extract from Crataegi Fructus was 22.5 mg/g. The inhibitory activity of the water extract (200 ${\mu}g/ml$) from Crataegi Fructus on ${\alpha}-amylase$ and ${\alpha}-glucosidase$ was determined to be 100% and 82.6%, respectively. Isolation of inhibitory compounds was carried out on Sephadex LH-20 and MCI-gel CHP-20 column chromatography using a gradient elution procedure of increasing MeOH in $H_2O$. The chemical structure of the inhibitory compound against ${\alpha}-amylase$ and ${\alpha}-glucosidase$ was confirmed as chlorogenic acid by spectroscopic analysis of FAB-MS, NMR and IR spectrum.

산사 추출물로부터 ${\alpha}-amylase$, ${\alpha}-glucosidase$ 저해활성을 탐색하고 Sephadex LH-20($5{\times}45$ cm)와 MCI-gel CHP-20를 이용하여 저해활성 물질을 정제하고 구조분석을 하였다. 산사 추출물의 페놀화합물의 함량은 물 추출물에서 22.5 mg/g, 60% ethanol 추출물에서 19.2 mg/g으로 나타났으며, 산사 물 추출물 (200 ${\mu}g/ml$)에서 ${\alpha}-amylase$, ${\alpha}-glucosidase$ 저해활성이 각각 100%와 82.6%로 나타났으나, 60% ethanol 추출물(200 ${\mu}g/ml$)에서는 ${\alpha}-glucosidase$ 저해활성이 12%로 낮은 저해활성을 나타내어 산사 물 추출물을 Sephadex LH-20를 이용하여 분획한 결과 I, II, III, IV의 4가지 분획물을 획득하고 다시 Sephadex LH-20과 MCI-gel을 이용하여 nomal phase type인 $EtOH{\rightarrow}H_2O$와 reverse phase type인 $H_2O{\rightarrow}MeOH$로 유기용매의 농도를 증가 시키며 용출하여 A, B, C, D의 compounds를 얻었다. 4가지 compound 중 compound C(200 ${\mu}g/ml$)가 ${\alpha}$-amylase, ${\alpha}$-glucosidase 저해활성이 각각 97.6%, 97.7%로 가장 높게 나타났으며, FAB-MS, NMR과 IR spectrum을 분석한 결과 chlorogenic acid로 동정되었다.

Keywords

References

  1. Park, S. Y. and Kim, J. W. (1992) Screening and isolation of the antitumor agents from medecinal plants (I). Korean J. Pharmacogn. 23, 264-267
  2. Huang, M. T., Ho, C. T. and Lee, C. Y. (1992) Phenolic compounds in food In phenolic compounds in food and their effects on health II. Maple Press, New York, pp. 2-7
  3. Park, S. W., Chung, Y. S., Yun, Y. S., Cha, B. S., Song, Y. D., Lee, H. C. and Huh, K. B. (1998) Insulin resistance and relate factors in the healthy young men. Diabetes 22, 504-512
  4. Moon, S. J. and Hong, S. M. (1996) A study on the relation between psychological stress and stress hormone, nutritional status of patients with non-insulin dependent diabetes mellitus Korean J. of Nutr. 29, 889-898
  5. Baik, S. C., Kim, J. B., Cho, M. J., Kim, Y. C., Park, C. K., Ryou, H. H., Choi, H. J. and Rhee, K. H. (1990) Prevalence of Helicobacter pylori infection among normal Korean adults (in Korean). J. Korean Soc. Mic. 25, 455-462
  6. Rhee, K. H., Youn, H. S., Baik, S. C., Lee, W. K. Cho, M. J., Choi, H.J., Maenh, K.Y. and Ko, KW. (1990) Prevalence of Helicobacter pylori infection in Korea (in Korean). J. Korean Soc. Mic. 25, 475-490
  7. Tabak, M., Armom, R., Potasman, I. and Neeman, I. (1996) In vitro inhibition of Helicobacter pylori by extracts of thyme. J. Appl. Bacteriol. 80, 667-672 https://doi.org/10.1111/j.1365-2672.1996.tb03272.x
  8. Diker, K.S. and Hascelik, G. (1994) The bactericidal activity of tea against Helicobacter pylori. Lett. Appl. Microbiol. 18, 299- 300
  9. Midolo, P.D., Lambert, J.R., Hull, R., Luo, F. and Grayson, M. S. (1995) In vitro inhibition of Helicobacter pylori NCTC 11637 by organic acids and lactic bacteria. J. Appl. Bacteriol. 79, 475-479 https://doi.org/10.1111/j.1365-2672.1995.tb03164.x
  10. Bhatia, S. J., Kochar, N., Abraham, P., Nair, N. and Mehta, A. P. (1989) Lactobacillus acidophilus inhibits growth of Campylobacter pylori in vitro. J. Clin. Microbiol. 27, 2328- 2338
  11. Feng G. H., Richardson, M., Chen, M. S., Kramer, K. J., Morgan, T. D. and Reek, G. R. (1996) Amylase inhibitors from wheat: Amino acid sequences and patterns of inhibition of insect and human ${\alpha}$-amylases. Insect Biochem. Molec. 26, 419- 426 https://doi.org/10.1016/0965-1748(95)00087-9
  12. Markwick, N. P., Laing, W. A., Cristeller, J. T., Reid, S. J. and Netwton, M. R. (1996) ${\alpha}$-amylase activities in larval midgut extraxcts from four species of Lepidoptera (Tortricidae and Gelechiidae): Response to pH and to inhibitors from wheat, barley, kidney bean, and streptomyces, J. Econ. Entomol. 89, 39-45 https://doi.org/10.1093/jee/89.1.39
  13. AOAC. (1989) Official method of analysis. 14th ed. Association of Official Analytical Chemists, Washington DC. 43, 1-3
  14. Fuwa, H. (1954) A new method for microdetermination of amylase activiy by the use of amylose as subsrate. J. Biochem. 41, 583-603 https://doi.org/10.1093/oxfordjournals.jbchem.a126476
  15. Tibbot, B. K. and skadsen, R. W (1996) Molecular cloning and characterization of a gibberellin-inducible, putative ${\alpha}$-glucosidase gene from berley. Plant Mol. Biol. 30, 229-241 https://doi.org/10.1007/BF00020110
  16. Nonaka, G. H. (1989) Isolation and structure elucidation of tannins. Pure Appl. Chem. 61, 357-363 https://doi.org/10.1351/pac198961030357
  17. Matsuo. T. and Ito, S. (1978) The chemical structure of kakitannin from immature fruit of the Persimmon (Diospyros kaki L.). Agric. Biol, Chem. 42, 1637-1643
  18. Blois, M. S. (1958) Antioxidant determination by the use of stable free radical. Nature. 26, 1198-1199
  19. Moon, J. S., Kim, S. J., Park, Y. M., Hwang. I. S. and Kim, E. H. (2004) Antimicrobial effect of methanol extracts from some medicinal herbs and the content of phenolic compounds. Korean J. Food Pre. 11, 207-213
  20. Paek, N. S. and Kim, Y. M. (1998) ${\alpha}$-Glucosidase inhibition by culture broth of streptomyces sp. NS15. Korean J. Food Nutr. 11, 640-646
  21. Mitsunnaga, T., Abe, L., Kontani, M., Ono, H. and Tanaka, T. (1997) Inhibitory effects of bark proanthocynaidins on the activities of glucosyltransferases of streptococcus sobrinus. L. Wood Shem. Korean J. Food Nutr. 17, 327-340
  22. Lee, W. Y., Ahn, J. K, Park, Y. K. and Rhee, H. I. (2004) Inhibitory effects of proanthocynidin extracted from distylium racemosum of ${\alpha}$-amylase and ${\alpha}$-glucosidase activities. Korean J. Pharmacogn. 35, 271-275
  23. Hulume, A. C. and J. D. Jones (1952) The isolation of chlorogenic acid from the apple fruit. Biochem. J. 53, 337-340
  24. Cho, Y. J., An, B. J. and Choi, C. (1993) Isolation and enzyme inhibition of tannins from korean green tea. Korean Bio. J. 26, 216-223
  25. Jin, U. H., Lee, J. Y., Kang, S. K., Kim, J. K., Park, W. H., Kim, J. G., Moon, S. K., and Kim, C. H. (2005) A phenolic compound, 5-caffeoylquinic acid (chlorogenic acid), is a new type and strong matrix metalloproteinase-9 inhibitor: Isolateion and identification from methanol extract of Euonymus alatus. Life Sic. 77, 2760-2769 https://doi.org/10.1016/j.lfs.2005.02.028