ARS | Publication request: Acetone Enhances the Direct Analysis of Total Condensed Tannins in Forage Legumes by the Butanol-HCl Assay

Submitted to: ASA-CSSA-SSSA Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: May 12, 2009
Publication Date: November 1, 2009
Citation: Grabber, J.H. 2009. Acetone Enhances the Direct Analysis of Total Condensed Tannins in Forage Legumes by the Butanol-HCl Assay. ASA-CSSA-SSSA Proceedings. Abstract 54581.

Technical Abstract: Depending on concentration, condensed tannins (CT) in forages have no effect, enhance, or impede protein utilization and performance of ruminants. Defining optimal forage CT levels has been elusive, partly because current methods for estimating total soluble plus insoluble CT are laborious or inaccurate. In this study, various co-solvents were evaluated for enhancing CT recovery and quantitation during rapid direct analysis of plant tissues by the widely used spectrophotometric butanol-HCl method. Among the co-solvents evaluated, acetone proved most effective for consistently increasing anthrocyanidin yields from CT standards and particularly plant tissues. When used in conjunction with appropriate CT standards, amending butanol-HCl reagent with 400 mL/L acetone increased tissue CT estimates from 13.3 to 34.2 g/kg in Lotus corniculatus and from 21.4 to 69.4 g/kg in Lotus uliginosus. Additional acetone had no effect on anthrocyanidin yields from L. corniculatus tissue, but adversely affected standards. Conversely, additional acetone increased anthrocyanidin yields from L. uliginosus tissue without affecting standards. Amending butanol-HCl with acetone did not alter CT estimates for Lotus neutral-detergent fiber (NDF). Thus acetone may partly act by enhancing CT accessibility during butanol-HCl hydrolysis, which is consistent with the widespread use of aqueous acetone for isolating soluble CT from plant tissue. Among other variables examined, running L. uliginosus CT standards with tannin-free tissue or NDF from Medicago sativa slightly reduced anthrocyanidin yields as did running assays of tissues from either Lotus species with a smaller volume of reagent. Thus, conditions for CT quantitation should be optimized for each type of plant material.