Fibre, particularly the woody fraction, is an essential factor in the digestive health of rabbits. Hence the need for precise characterisation of raw materials, in order to provide the right quantities in the feed. There are several methods for measuring lignin. The two most commonly used in animal nutrition are Van Soest's ADL (Acid Detergent Lignin) method and the direct method for lignin insoluble in H2SO4. A Techna France Nutrition study compared these two methods of chemical analysis of lignin with several raw materials. So which method is best suited to determining the lignin content of raw materials and therefore of rabbit feed?
Significant deviations between Van Soest’s technique and the direct method in H2SO4
The study carried out by Techna shows that Van Soest's ADL method leads to an overestimation of the lignin value for certain raw materials.
For example, for grape co-products, the ADL value is much higher (up to + 7.7 pts) than the Lignin H2SO4 value. There is a more modest difference between sunflower husks and cakes.
In order to determine the most accurate method, we analysed a mixture of fibrous raw materials, Lapilest®, and the individual raw materials that make it up. The weighted sum of the raw materials was compared with the analytical value of the blend.
The additivity is therefore validated by the H2SO4 lignin method, while the ADL analysed on the blend is higher than the expected value. This discrepancy clearly reflects an overestimation of the Van Soest method, whereas the H2SO4 lignin method restores the unit values of the raw materials.
Implications of overestimating lignin levels
According to the study cited above, the Van Soest method overestimates lignin levels for certain raw materials (grape co-products, carob, etc.). A rabbit feed containing this type of raw material is therefore likely to have a lignin deficiency when analysed using the ADL method. This can have significant consequences for the health of the animals (digestive problems, mortality, etc.).
Is the polyphenol content the key to the differences?
One explanation for the difference in results between the two methods of analysis may be linked to the polyphenol content of the raw materials: 3380µg/g for grape pulp and 5208µg/g for grape seed, with a high proportion of flavonoids (87.6% and 92.9%), a sub-family that contains tannins in particular.
Literature data shows that in the presence of tannins, the neutral detergent etching stage of the Van Soest method generates protein-tannin complexes. In addition, the various high-temperature drying stages (3 x 3h at 103°C) increase the polymerisation of these tannins and their complexation with other cellular constituents (fibres, proteins) which are not totally solubilised during the acid attacks (acid detergent then H2SO4). The ADL residue obtained by the Van Soest method would therefore contain lignin as well as polymerised tannins. Hence an overestimate of the lignin level measured.
What applications for our rabbit formulation?
On the basis of these findings, the Techna group's service company is characterising the lignin in raw materials and feeds with two nutrients: Van Soest's ADL and Lignin insoluble in H2SO4. These nutrients are derived from the results of analyses or from a specific equation.
The nutritional recommendations in our feed specifications are based on the H2SO4 lignin criterion, to ensure the most accurate formulation possible.
This study highlighted differences between the two lignin analysis methods on certain fibrous raw materials and their blends. These differences are linked to the presence of tannins, which interfere with the analysis. In the light of these conclusions, our experts recommend characterising the woody fraction of polyphenol-rich raw materials and rabbit feeds, preferably using the method for lignin insoluble in H2SO4. Raw materials with little wood and those containing few polyphenols could be analysed using Van Soest's ADL method.
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