Method 75 Determination of neutral detergent fibre NDF or plant cellwall constituents

This is based on the method by Van Soest and Wine (1967) which has been modified according to subsequent recommendations. It is the only fibre determination suitable for non-ruminants. The residue consists of the plant cell-wall constituents: cellulose, hemicellulose, lignin, cutin, NDF-insoluble tannin and ash. See the article by Cherney (2000) for current modifications; these include the use of amylase to aid in the removal of starch from forages containing grain (Van Soest et al., 1991), which has been adopted by MAFF

(1993c). Also, the use of sodium sulphite and decalin has been eliminated; see Chapter 4 for further discussion.

Reagents.

• Neutral detergent solution - add 30 g of sodium dodecyl sulphate (sodium lauryl sulphate, CH3(CH2)11OSO3Na), 18.61 g of EDTA disodium salt, 6.81 g of sodium borate decahydrate, and 4.56 g of disodium hydrogen phosphate (anhydrous), to 1 l water, and stir to dissolve. Adjust the pH to 6.9-7.1 if necessary. Note: sodium dodecyl sulphate dust irritates the lungs, therefore wear dust masks when weighing and use dust extraction fans. If foaming is a problem, also add 10 ml triethylene glycol (HO(CH2)2O(CH2)2O(CH2)2OH); the 2-ethoxyethanol previously used is toxic and should be avoided.

• Acetone, commercial 'drum' grade.

Procedure. Weigh 0.5 g of the freeze-dried (or air-dried, but not oven dried) sample, ground to 1 mm, and transfer to a 500-ml short-neck round-bottom flask with ground glass socket size 34/35. (Note: if the sample is an animal feed with ^10% oil content, it should be defatted with petroleum spirit (40-60°C) before proceeding.) At the same time, weigh a sample for dry matter determination. Add 100 ml neutral detergent solution (ambient temperature), place on a macro Kjeldahl heating unit and 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 (place in cool muffle, raise to 500°C and maintain for 30 min) and weighed Porosity 1 sintered glass crucible with gentle suction (excessive suction compresses the mat and hinders the efficiency of the washing/filtration). Rinse the sample particles from the flask with the minimum amount of very hot water (80-90°C). Remove the vacuum, break up the mat of sample fibres with a small glass rod (approximately 4 mm diam. with ends rounded in a flame) and fill the crucible with very hot water. Apply just sufficient suction to filter, and repeat the washing process. Wash twice with acetone using the same technique and suck dry. Allow to air-dry in a fume cupboard until no smell of acetone is detectable, then dry overnight in an oven at 100°C, cool in a desiccator, and weigh.

Place the crucible and its contents of NDF in a cool muffle furnace. Increase the temperature of the furnace to 500 ±5°C; keep at this temperature (for approximately 3 h) until ashing is complete. It is important not to overshoot the temperature, because the sintered disc will be damaged at ^515°C. Remove the crucible from the muffle furnace, cool in a desiccator and weigh.

Calculation. Subtract the weight of the empty crucible from that of the crucible plus NDF to obtain the weight of NDF in 0.5 g sample. Divide by the sample weight and multiply by 100 to obtain the % NDF in the freeze-dried sample. Multiply this figure by 100/(100-moisture content) to obtain the % NDF in DM. Subtract the empty crucible weight from the weight of crucible plus ash and multiply by 100/weight of NDF to obtain the % ash in the NDF.

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