• Acid detergent solution - add 100 g cetyl trimethylammonium bromide (CTAB), also called hexadecyltrimethylammonium bromide, technical grade, to 2.5 l of 1 M sulphuric acid, stir to dissolve and make up to 5 l
• Sulphuric acid, 1 M - add carefully 56 ml sulphuric acid (approximately 98% m/m H2SO4 and 18 M) to about 400 ml water in an 800-ml beaker and stir to dissolve. Transfer with washings to a 1-l volumetric flask, make up to the mark and mix
• Octan-2-ol (also called 2-octanol; capryl alcohol)
• Acetone, commercial 'drum' grade
Procedure. Weigh to the nearest 0.001 g approximately 1 g of oven-dried (^65°C) plant material (or air-dried animal feed), ground to 1 mm, into a 500ml short-neck round-bottomed flask with a ground-glass socket size 34/35. Add 100 ml acid detergent solution, and, if excess foaming is likely to occur, add 1-2 drops octan-2-ol. Place on a macro Kjeldahl heating unit, connect a coil condenser with size 34/35 ground-glass cone, turn on a steady supply of water, and bring to the boil on full heat, then turn the regulator down and allow to simmer for 1 h. Occasionally swirl the flask to wash any sample particles from the flask wall back into the detergent; also, if any bumping has caused sample particles to enter the condenser, squirt a wash-bottle into the top of the condenser to wash them back into the detergent using the minimum amount of water. Filter on a previously dried (100°C) and weighed Porosity 1 sintered glass crucible with gentle suction. The mat of residue is broken up with a small rounded end glass rod and washed twice with water near to boiling point (wear heat resisting gloves). Wash any residue from the sides of the crucible, then wash with successive portions of acetone until no further colour is removed. Suck the residue dry of acetone and allow to stand, preferably in a fume cupboard, until no smell of acetone can be detected, then dry overnight at 100°C. Cool in a desiccator and weigh the crucible plus ADF. Retain the residue for lignin and cellulose determination if required.
Calculation. Subtract the weight of the empty crucible from that of the crucible plus ADF to obtain the weight of ADF. Divide by the sample weight and multiply by 100 to obtain the % ADF in the sample DM (air-dry matter for animal feeds).
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