Spectrophotometric Investigations on Lignin in Wheat (Triticum aestivum L.): Influence of Cell Wall Preparation, Solvent, and Standard

Absorbance at 280 nm was used to determine lignin concentrations in solutions of cell walls from winter wheat whole-crop silage, stems, and straw. Three different cell wall preparations (hot water−organic solvent fiber, ethanol−benzene residue, neutral detergent fiber) were prepared and dissolved in either HCl−triethylene glycol or acetyl bromide. Three phenolic standard solutions were used to calculate lignin concentrations in the wheat materials from absorbance readings: mixtures of phenolic monomers based on aldehydes (HIGUCHI), aldehydes and acids (REEVES), and washed Indulin (WIND). Gravimetric Klason lignin determination on the cell wall preparations consistently gave higher values than spectrophotometric lignin analysis. Material recovered as cell wall was slightly greater for hot water−organic solvent fiber than for neutral detergent fiber and was lowest for ethanol−benzene residue, probably due to partial dissolution of cell wall components with the latter procedure. Acetyl bromide shifted UV spectra about 10 nm to lower wavelengths, which complicates quantification of absorbance readings. Additionally, triethylene glycol is easier to handle and less toxic. Lignin concentrations related to the REEVES standard were more consistent across cell wall preparations and solvents than other standards. It appears that the REEVES standard was best suited for winter wheat whole-crop silage, stems, and straw.