Journal of Chemical Ecology

, Volume 17, Issue 1, pp 123–134 | Cite as

Methods and pitfalls of extracting condensed tannins and other phenolics from plants: Insights from investigations onEucalyptus leaves

  • Steven J. Cork
  • Andrew K. Krockenberger


Optimal conditions for extraction of tannins and other phenolics from tree foliage and their subsequent storage rarely have been investigated. We investigated methods of drying leaves, optimal solvents, and the effects of light and temperature on the extractability and stability of condensed tannins (proanthocyanidins) and total phenolics from leaves ofEucalyptus trees. Aqueous acetone was a better solvent than aqueous methanol for condensed tannins and total phenolics, but condensed tannins were less stable in aqueous acetone than aqueous methanol. Stability of condensed tannins also was decreased substantially by room temperature versus 4°C and by exposure to indirect sunlight, although the assay for total phenolics was unaffected. For quantitative estimation of condensed tannins, extraction with 50% acetone was better than methods of direct analysis of leaf tissue. The highest estimates of total condensed tannins were obtained by exhaustive extraction with 50% acetone followed by direct analysis of the residue. Lyophilization of fresh leaf increased yield of condensed tannin (although usually by less than 10%). Lyophilization and subsequent storage of extracts had little effect on assays for condensed tannins or total phenolics.

Key Words

Tannin extraction plant phenolics Eucalyptus browse analysis 


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  1. Asquith, T.N., andButler, L.G. 1985. Use of dye-labeled protein as spectrophotometric assay for protein precipitants such as tannin.J. Chem. Ecol. 11:1535–1544.Google Scholar
  2. Bate-Smith, E.C. 1973. Tannins of herbaceous leguminoseae.Phytochemistry 12:1809–1812.Google Scholar
  3. Bate-Smith, E.C. 1975. Phytochemistry of proanthocyanidins.Phytochemistry 14:1107–1113.Google Scholar
  4. Chippendale, G.M. 1988.Eucalyptus, Angophora (Myrtaceae), Flora of Australia, Vol. 19. Australian Government Publishing Service, Canberra.Google Scholar
  5. Cork, S.J. 1984. Utilization ofEucalyptus foliage by arboreal marsupials.Proc. Natr. Soc. Aust. 9:88–97.Google Scholar
  6. Cork, S.J., andPahl, L. 1984. The possible influence of nutritional factors on diet and habitat selection by the ringtail possum (Pseudocheirus peregrinus), pp. 269–276,in A.P. Smith and I.D. Hume (eds.). Possums and Gliders. Australian Mammal Society, Sydney.Google Scholar
  7. Cork, S.J., Hume, I.D., andDawson, T.J. 1983. Digestion and metabolism of a natural foliar diet (Eucalyptus punctata) by an arboreal marsupial, the koala (Phascolarctos cinereus).J. Comp. Physiol. 153:181–190.Google Scholar
  8. Foo, L.Y., andPorter, L.J. 1980. The phytochemistry of proanthocyanidin polymers.Phytochemistry 19:1747–1754.Google Scholar
  9. Fox, L.R., andMacauley, B.J. 1977. Insect grazing onEucalyptus in response to variation in leaf tannins and nitrogen.Oecologia 29:145–162.Google Scholar
  10. Gartlan, J.S., McKey, D.B., Waterman, P.G., Mbi, C.N., andStruhsaker, T.T. 1980. A comparative study of the phytochemistry of two African rain forests.Biochem. Syst. Ecol. 8:401–422.Google Scholar
  11. Goldstein, J.L., andSwain, T. 1963. Changes in tannins in ripening fruits.Phytochemistry 2:371–383.Google Scholar
  12. Hagerman, A.E. 1988. Extraction of tannin from fresh and preserved leaves.J. Chem. Ecol. 14:453–461.Google Scholar
  13. Hagerman, A.E., andButler, L.G. 1989. Choosing appropriate methods and standards for assaying tannin.J. Chem. Ecol. 15:1795–1810.Google Scholar
  14. Hagerman, A.E., andRobbins, C.T. 1987. Implications of soluble tannin-protein complexes for tannin analysis and plant defense mechanisms.J. Chem. Ecol. 13:1243–1259.Google Scholar
  15. Haslam, E., Haworth, R.D., Mills, S.D., Rogers, H.J., Armitage, R., andSearle, T. 1961. Gallotannins. Part II. Some esters and depsides of gallic acid.J. Chem. Soc. 1961:1836–1842.Google Scholar
  16. Hillis, W.E. 1966. Variation in polyphenol composition within species ofEucalyptus L'Herit.Phytochemistry 5:541–556.Google Scholar
  17. Inoue, K.H., andHagerman, A.E. 1988. Determination of gallotannin with rhodanine.Anal. Biochem. 169:363–369.Google Scholar
  18. Jones, W.T., Broadhurst, R.B., andLyttleton, J.W. 1976. The condensed tannins of pasture legume species.Phytochemistry 15:1407–1409.Google Scholar
  19. Lindroth, R.L., andPajutee, M.S. 1987. Chemical analysis of phenolic glycosides: Art, facts, and artifacts.Oecologia 74:144–148.Google Scholar
  20. Martin, J.S., andMartin, M.M. 1983. Tannin assays in ecological studies. Precipitation of ribulose-1,5-biphosphate carboxylase/oxygenase by tannic acid, quebracho, and oak foliage extracts.J. Chem. Ecol. 9:285–294.Google Scholar
  21. Martin, M.M., andMartin, J.S. 1984. Surfactants: Their role in preventing the precipitation of proteins by tannins in insect guts.Oecologia 61:342–345.Google Scholar
  22. McArthur, C. 1988. Influences of tannins on digestion of eucalypt foliage in common ringtail possums (Pseudocheirus peregrinus) and an analysis of some chemical assays in the presence of condensed tannins. PhD thesis. Zoology Department, Monash University, Melbourne.Google Scholar
  23. Mole, S., andWaterman, P.G. 1987a. A critical analysis of techniques for measuring tannins in ecological studies. I. Techniques for chemically defining tannins.Oecologia 72:137–147.Google Scholar
  24. Mole, S., andWaterman, P.G. 1987b. A critical analysis of techniques for measuring tannins in ecological studies. II. Techniques for biochemically defining tannins.Oecologia 72:148–156.Google Scholar
  25. Mole, S., Butler, L.G., Hagerman, A.E., andWaterman, P.G. 1989. Ecological tannin assays: A critique.Oecologia 78:93–96.Google Scholar
  26. Porter, L.J., Hrstich, L.N., andChan, B.G. 1986. The conversion of procyanidins and prodelphinidins to cyanidin and delphinidin.Phytochemistry 25:223–230.Google Scholar
  27. Price, M.L., Stromberg, A.M., andButler, L.G. 1979. Tannin content as a function of grain maturity and drying conditions in several varieties ofSorghum bicolor (L.) Moench.J. Agric. Food Chem. 27:1270–1274.Google Scholar
  28. Reed, J.D., McDowell, R.E., Van Soest, P.J., andHorvath, P.J. 1982. Condensed tannins: A factor limiting the use of cassava forage.J. Sci. Food Agric. 33:213–220.Google Scholar
  29. Singleton, V.L., andRossi, J.A. 1965. Colorimetry of total phenolics and phosphomolybdicphosphotungstic acid reagents.Am. J. Enol. Vitic. 16:144–158.Google Scholar
  30. Stafford, H.A., andCheng, T.-Y. 1980. The procyanidins of douglas fir seedlings, callus and cell suspension cultures derived from cotyledons.Phytochemistry 19:131–135.Google Scholar
  31. Swain, T. 1979. Tannins and lignins, pp. 657–682,in G.A. Rosenthal and D.H. Janzen (eds.). Herbivores: Their Interaction with Secondary Plant Metabolites. Academic Press, New York.Google Scholar
  32. Swain, T., andHillis, W.E. 1959. The phenolic constituents ofPrunus domestica I. The quantitative analysis of phenolic constituents.J. Sci. Food Agric. 10:63–68.Google Scholar
  33. Watterson, J.J., andButler, L.G. 1983. Occurrence of an unusual leucoanthocyanidin and absence of proanthocyanidins in sorghum leaves.J. Agric. Food Chem. 31:41–45.Google Scholar
  34. Zar, J.H. 1984. Biostatistical Analysis. Prentice-Hall, Englewood Cliffs, New Jersey.Google Scholar

Copyright information

© Plenum Publishing Corporation 1991

Authors and Affiliations

  • Steven J. Cork
    • 1
  • Andrew K. Krockenberger
    • 1
  1. 1.Division of Wildlife & EcologyCSIROLynehamAustralia

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