2.4.4. Determination of Lignin

1g of extractive-free sample was placed in the conical flask and 12 ml of 72% sulfuric acid was added. 100 mL of distilled water was added to the mixture. After distilled water addition, the mixture was boiled for 2 hours and cooled at room temperature. The insoluble material (lignin) was filtered by using Whatman paper (no. 42). The lignin was washed with distilled water until neutralized with H₂SO₄ and then oven dried at 105°C for 10 hours at a constant weight and cooled down in a desiccator and weighed [7, 24]. Lignin content was calculated as follows:

where W₁ = oven dried sample (1 g); W₂ = extracted residues.