Direct versus Sequential Analysis of Procyanidin- and Prodelphinidin-Based Condensed Tannins by the HCl-Butanol-Acetone-Iron Assay.

In this study, we optimized the HCl-butanol-acetone-iron (HBAI) assay for the analysis of B-linked procyanidin (PC) and prodelphinidin (PD) condensed tannins (CTs) by direct analysis of whole tissue and sequential analysis of acetone-water extracts and insoluble residues prepared from forage, woody plant, food, and food byproduct samples. Yields of anthocyanidins (cyanidin and delphinidin) were optimized by heating ≤0.25 mg mL-1 CT standards, 1 mg mL-1 tissue, or 1-2 mg mL-1 acetone-water fractioned tissue for 3 h at 70 °C in medium containing 5% concentrated HCl, 6.7% total water, 50% acetone, 42% n-butanol, and 0.15% ammonium iron(III) sulfate dodecahydrate. Accurate quantitation required CT standards of known purity sourced from the same tissue being analyzed. Both analysis methods provided comparable estimates of total CTs for most PD-rich samples, but only the sequential method gave good recovery and accurate estimates of CTs in most PC-rich samples.