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A modified dot-blot method of protein determination applied in the tannin-protein precipitation assay to facilitate the evaluation of tannin activity in animal feeds

Published online by Cambridge University Press:  09 March 2007

E. M. Hoffmann ,
S. Muetzel  and
K. Becker
E. M. Hoffmann
Affiliation:
Institute for Animal Production in the Tropics and Subtropics (480), University of Hohenheim, Fruwirthstr. 12, D-70593 Stuttgart, Germany
S. Muetzel
Affiliation:
Institute for Animal Production in the Tropics and Subtropics (480), University of Hohenheim, Fruwirthstr. 12, D-70593 Stuttgart, Germany
K. Becker*
Affiliation:
Institute for Animal Production in the Tropics and Subtropics (480), University of Hohenheim, Fruwirthstr. 12, D-70593 Stuttgart, Germany
*
*Corresponding author: Professor Dr K. Becker, fax +49 711 459 3702, email [email protected]
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Abstract

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Tannins have received considerable attention from animal nutritionists as potential agents for modifying ruminal fermentation patterns, or for exploring new feed resources. This group of secondary plant compounds is defined by their ability to form complexes with proteins. A widely accepted method for assaying the biological activity of extracted tannins is the precipitation of bovine serum albumin. The protein carries a radioactive label (125I) to allow direct quantification from the precipitate. Tannin–protein complexes dissolve in sodium dodecylsulfate solution. A dot-blot assay for protein determination, which is based on the reversible binding of a fluorochrome, benzoxanthene yellow, to the protein spots and is not disturbed by the presence of detergents, can replace the radioactive method by a fluorimetric measurement. A novel alternative to the last part of the dot-blot assay is to scan the stained protein spots in situ using a video camera and computer image analysis. Several filter sets were tested and, within a concentration range of 0·1–2·0 mg protein/ml, each of them yielded results identical to the original method while the time required was only 30 % of the working time consumed by the original procedure. The modified dot-blot assay should be applicable to the evaluation of tannin activity in all shrub and tree foliages considered as animal feed.

Keywords

Protein determinationBenzoxanthene yellowTannin activityFeed evaluation
Type
Research Article
Information
British Journal of Nutrition , Volume 87 , Issue 5 , May 2002 , pp. 421 - 426
Copyright
Copyright © The Nutrition Society 2002

References

Barry, TN & McNabb, WC (1999) The implications of condensed tannins on the nutritive value of temperate forages fed to ruminants. British Journal of Nutrition 81, 263–272.CrossRefGoogle ScholarPubMed
Bradford, MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry 72, 248–254.CrossRefGoogle ScholarPubMed
Guttenberger, M, Neuhoff, V & Hampp, R (1991) A dot-blot assay for quantitation of nanogram amounts of protein in the presence of carrier ampholytes and other possibly interfering substances. Analytical Biochemistry 196, 99–103.CrossRefGoogle ScholarPubMed
Guttenberger, M, Schaeffer, C & Hampp, R (1994) Kinetic and electrophoretic characterization of NADP dependent dehydrogenases from root tissues of Norway spruce (Picea abies (L.) Karst) employing a rapid one-step extraction procedure. Trees 8, 191–197.CrossRefGoogle Scholar
Hagerman, AE, Rice, ME & Ritchard, NT (1998) Mechanisms of protein precipitation for two tannins, pentagalloyl glucose and epicatechin(16) (4→8) catechin (procyanidin). Journal of Agricultural and Food Chemistry 46, 2590–2595.CrossRefGoogle Scholar
Hagerman, AE, Robbins, CT, Weerasuriya, Y, Wilson, TC & Mcarthur, C (1992) Tannin chemistry in relation to digestion. Journal of Range Management 45, 57–62.CrossRefGoogle Scholar
Lowry, OH, Rosebrough, NJ, Farr, AL & Randall, RJ (1951) Protein measurement with the Folin phenol reagent. Journal of Biological Chemistry 193, 265–275.CrossRefGoogle ScholarPubMed
Makkar, HPS, Dawra, RK & Singh, B (1988) Changes in tannin content, polymerisation and protein precipitation capacity in oak (Quercus incana) leaves with maturity. Journal of the Science of Food and Agriculture 44, 301–307.CrossRefGoogle Scholar
Mueller-Harvey, I (2001) Analysis of hydrolysable tannins. Animal Feed Science and Technology 91, 3–20.CrossRefGoogle Scholar
Neuhoff, V, Philipp, K, Zimmer, HG & Mesecke, S (1979) A simple, versatile, sensitive and volume-independent method for quantitative protein determination which is independent of other external influences. Zeitung für Physiologische Chemie 360, 1657–1670.CrossRefGoogle ScholarPubMed
Porter, LJ, Hrstich, LN & Chan, BG (1986) The conversion of procyanidins and prodelphinidins to cyanidin and delphinidin. Phytochemistry 25, 223–230.CrossRefGoogle Scholar
Schofield, P, Mbugua, DM & Pell, AN (2001) Analysis of condensed tannins: a review. Animal Feed Science and Technology 91, 21–40.CrossRefGoogle Scholar
Spector, T (1978) Refinement of the coomassie blue method of protein quantitation. Analytical Biochemistry 86, 142–146.CrossRefGoogle ScholarPubMed