In-situ disappearance and content of p-coumaric and ferulic acid in lucerne from various vegetative stages

Content of monomeric p-coumaric (PCA) and ferulic (FA) acid was determined by HPLC either in the bud stage lucerne sample after its incubation in the rumen (0, 2, 4, 8, 16, 24, 48 h) by the in-situ method (three steers equipped with rumen cannulas, fed near the maintenance level of nutrition), or in the lucerne samples cut in ten consecutive vegetative stages. PCA and FA were analysed in four lucerne fractions based on methanol extraction: free phenolic acids (F1) extracted into ether from the methanol extract, ester-bonded phenolic acids (F2) after alkaline hydrolysis of the methanol extract, glycoside-bonded phenolic acids (F4) after acid hydrolysis of the extract, and cell wall-bonded phenolic acids (F3) after alkaline hydrolysis of the solid residue after the extraction. d 98% of the total content of PCA and FA, respectively, disappeared in the rumen within the 5h interval. Total content of PCA and FA (Y) decreased linearly with increasing maturity (vegetative stage quantified by the mean stage by weight number, MSW, X): Y = 82.3 − 9.80X (r2 = 68%, P < 0.01) and Y = 97.8 − 6.53X (r2 = 41%, P < 0.05), respectively. However, PCA and FA content increased significantly (P < 0.01) to 67 and 120mg 100g−1 of dry matter, respectively, in the bud stage (MSW = 3.5). Content of the PCA fraction bound to cell wall was correlated with lucerne organic matter degradability (r2 = 66%, P < 0.01), this relationship was not significant in the case of FA (r2 = 17%, P > 0.05).