EN ISO 16472 Animal Feeds - Determination of Amylase-Treated Neutral Detergent Fibre Content (aNDF)

The standard "EN ISO 16472 Animal feeds - Determination of amylase-treated neutral detergent fibre content (aNDF)", which is based on a European standard and subsequently published by the International Organization for Standardization (ISO), describes methods for determining the amylase-treated neutral detergent insoluble fibre residue content in all types of animal feed. This standard includes a gravimetric routine method and a reference method.

For the purposes of this standard, amylase-treated neutral detergent fibre content (aNDF content) refers to the mass fraction of insoluble fibre residues determined by the procedure specified in this standard. aNDF content is expressed as a percentage by mass.

In principle, neutral detergent (ND) solution and heat-stable alpha-amylase are used to dissolve easily digestible proteins, lipids, sugars, starches and pectins in feeds, leaving an insoluble fibrous residue consisting of nitrogenous substances that are cell wall components of plant materials (cellulose, hemicellulose and lignin) and indigestible in animal products.

In addition to the usual laboratory equipment, the following equipment is used in these tests:

• Analytical balance with 0,1 mg readability, capable of weighing to the nearest 0,1 mg.
• Cyclone mill with 220 mm mesh or cutter mill with 260 mm mesh that can grind samples to obtain geometric mean particle size of 2 µm to 1 µm
• Reflux apparatus with separate heating units and cold water condensers, suitable for 600 ml vessels. Any conventional apparatus suitable for crude fibre determination is acceptable. When cold water condensers are used, the heating unit settings are calibrated to boil 50 ml of water in 4 to 5 minutes. A Fibertec type apparatus can be used and should boil 50 ml of water in 10 minutes.
• Gooch crucible with fritted disc, coarse porous crucible (pore size 40 µm to 60 µm), high form, 40 ml to 50 ml capacity or P2 (pore size 40 µm to 100 µm), 26 ml to 28 ml capacity.

New crucibles and ash are cleaned at 1 degrees for 500 hour. After each use, the crucibles are cleaned by ashing at 3 degrees for 500 hours, removing the ash, turning them upside down in detergent solution and sonicating for 7 to 10 minutes. The crucibles are rinsed in hot water and soaked in room temperature water for at least 30 minutes. A rubber stopper equipped with a trap and a port connected to a vacuum line is attached to the top of each crucible. Each crucible is backwashed with water by repeatedly immersing and withdrawing it, creating a strong rinsing effect. The filtration rate is checked occasionally. If cleaning does not improve the filtration rate, the crucible is discarded.

• Vacuum filter manifold (e.g. Fibertec type) that allows adequate wetting of fibrous debris. The manifold should provide a vacuum tight seal with the crucible to reduce foam formation in the vacuum lines. Heavy walled vacuum tubing is used to connect the manifold to a trap and vacuum source. A vacuum chamber is recommended between the trap and vacuum source to provide sufficient vacuum capacity to remove foam.
• A continuous boiling water generator or a suitable alternative is used. The instrument should provide sufficient boiling water (at least 95 degrees) for all samples being washed at one time and should be fed through a nozzle that produces a fine stream (a flow rate of 10 ml to 35 ml in 40 seconds, a 2,5 ml disposable plastic pipette tip makes an acceptable nozzle). A fine nozzle minimizes the water required to transfer particles to the crucible, but provides the water pressure required to remove debris adhering to the side of the flask. It is critical that the water be brought to a boil when water is added to the crucible, especially for samples containing starch, pectin substances, mucilage or glycoproteins. For the Fibertec type instrument, a syringe with a cone-spray nozzle is used to rinse the condensers and a 10 ml disposable syringe with a 12 cm long 60 gauge needle is used to remove debris adhering to the condensers.

Sampling is not part of the method specified in this standard. A suggested sampling method is included in ISO 6497. It is important that the laboratory obtains a sample that is truly representative and has not been damaged or altered during transport or storage.

Prepare test samples in accordance with ISO 6498. Samples should be air dried (approximately 90 percent dry matter) for ease of sample storage and grinding. Wet samples are dried at a maximum of 60 degrees to prevent the formation of artificial fibers. The amount of residue remaining after extraction is affected by the particle size of the sample. Representative samples are ground to obtain a geometric mean particle size of 220 µm to 260 µm.

Our organization, which has been trying to support businesses from every sector with its testing, measurement, analysis and evaluation studies carried out in a wide range for years, has a strong staff that closely follows the developments in the world in the field of science and technology and constantly improves itself. In this context, testing services are also provided to businesses in accordance with the standard "EN ISO 16472 Animal feeds - Determination of amylase-treated neutral detergent fiber content (aNDF)".