These enzymes have been a key focus for the Huvepharma R&D team for over a decade and, in conjunction with external research groups, they have been striving to improve the understanding of the mode of action of fibre-degrading enzymes in the monogastric gastrointestinal tract (GIT). One of these collaborations, a Huvepharma-sponsored PhD project at the Laboratory of Food Chemistry at Wageningen University and Research (WUR), explored, on a molecular level, the impact of fibre-degrading enzymes (FDEs) on the structure of NSPs and what this means for nutrient digestion and hindgut fermentation in broilers.
The findings of this recently concluded project provide clear answers on the in vitro mode and mechanisms of action of FDEs, shedding new light on their prebiotic effect, a field Huvepharma sees as so far lacking concrete evidence.
In this Industry Perspectives article, Feedinfo talks to the project lead, WUR researcher Dr. Dimitrios Kouzounis and Global Product Manager Enzymes at Huvepharma, Natalia Soares MSc, about the importance of their research and takes a closer look at the findings and what they mean for animal nutrition science and the wider animal nutrition sector.
[Feedinfo] Ms Soares, can you tell us why Huvepharma initiated this research project?
[Natalia Soares] Cereal NSPs play an important role in animal nutrition with both negative and positive implications. Consequently, fibre manipulation to reduce the negative effects of NSPs for nutrient digestion and increase the positive effects of NSPs in fermentation is of particular interest to improve animal health and performance, while minimising costs. The main means of achieving this is the supplementation of diets with FDEs that can depolymerise polysaccharides.
With the increasing interest in the inclusion of high-fibre agricultural by-products in animal diets the role of FDEs will become even more important in the future, making the full understanding of their mode of action and efficiency highly relevant to maximising animal performance and profitability.
[Feedinfo] Dr. Kouzounis, can you briefly explain what exactly you explored in your research for this project? What were the areas that interested you?
[Dimitrios Kouzounis] The supplementation of monogastric animal diets with FDEs has become common practice, but the underlying mechanisms of the enzyme action have not yet been fully elucidated.
The beneficial impact of FDEs in animal performance is made up of three main distinct mechanisms:
- viscosity reduction
- release of prebiotic oligosaccharides
- de-encapsulation of entrapped nutrients in the cell wall matrix
The viscosity reduction mechanism is well documented and studied, but the other two mechanisms, especially the release of prebiotic compounds, remained elusive. Despite several studies pointing to the in vivo release in the small intestine of arabinoxylan oligosaccharides (AXOS), the prebiotic compounds formed by the use of FDEs, and the detection of those AXOS in animal digesta was poorly reported and there was no direct evidence of their formation.
My work focused on cereal NSP degradation by FDEs in broilers, looking at their impact on oligosaccharide formation, carbohydrate fermentation patterns and nutrient utilisation in the GIT. It also encompassed the impact of FDEs on the caeca microbiota composition of broilers. This research narrowed in on the in vivo release of AXOS, the fermentation of arabinoxylan and AXOS to short chain fatty acids (SCFAs), the transit behaviour and degradation profile along the GIT of cell wall components, and the associations between AXOS fermentation and the proliferation of beneficial gut microflora populations known to ferment AXOS to SCFAs.
[Feedinfo] So what were some of the main findings and conclusions of this work and can you elaborate on its relevance to the animal nutrition sector?
[Dimitrios Kouzounis] My research has shown that FDEs, specifically endo-xylanase (from Huvepharma) releases AXOS of diverse structures in vivo, leading to a pronounced arabinoxylan and AXOS fermentability to SCFAs, to alterations in caeca ecology, and ultimately to pronounced nutrient digestion and animal growth.
NSP degradation and the fate of NSP components in vivo have been poorly characterised so far due to analytical constraints and method-related limitations. The in vivo formation of AXOS by endo-xylanase alters arabinoxylan utilisation in broilers and this research used highly advanced chromatographic and mass spectrometric methods to portray the structural variability of AXOS and followed their utilisation in the hindgut. Moreover, a newly developed analytical strategy used in my research allowed us to separate and identify the produced AXOS, showing for the first time the action of the enzyme in the animal digestive tract and that arabinoxylan was degraded in the upper GIT. It also allowed us to measure and characterise the formed AXOS. In addition, the bio-functionality of AXOS was investigated by monitoring the AXOS fate throughout the GIT in a quantitative manner.
Alongside my research, additional in vitro experiments were carried out on wheat arabinoxylan to further investigate the magnitude of AXOS release by dietary xylanase, using the same enzymes as used in vivo. Here the endo-xylanase was added to isolated cell wall material and arabinoxylan and AXOS were solubilised (released) under conditions mimicking digestion in the upper GIT of poultry. Released AXOS were identified and quantified. The data obtained were compared with the observations made in vivo in samples from the gizzard and ileum segments of broilers fed wheat-based diets and, overall, this in vitro study confirmed the ability of the endo-xylanase used to release AXOS under conditions simulating the upper GIT. More importantly, the same endo-xylanase presented comparable extent of arabinoxylan degradation in vivo and in vitro.
Lastly, the high-end analytical techniques, as well as the isolation and in vitro digestion procedures currently developed can greatly contribute to further research on in vivo fibre utilisation. For a more in-depth look at the research, click this link.
[Feedinfo] What is the relevance of this research for Huvepharma itself and what do the key learnings mean for your enzymes business?
[Natalia Soares] It's true that to get the most out of fibre, from an animal nutrition and health perspective, we need to be able to translate the newest science into practice. The findings of this PhD research and the collaboration with WUR brought Huvepharma several new and proven insights into fibre degradation by FDEs. Knowing what happens to the degradation products of fibre and how they are used by the animal's intestinal microbiota - and being able to measure it - means that we can control and optimise it.
This research clearly demonstrated the ability of our endo-xylanase to form AXOS of varying structures throughout the upper GIT of broilers. Moreover, it provided solid evidence regarding the beneficial impact of enzyme supplementation for animal growth, nutrient digestion and hindgut fermentation. And based on these findings, we can now show that the magnitude of AXOS release by our xylanase during feed digestion is significant and can positively impact broiler health and growth.
It is also worth mentioning that the structural analysis of released AXOS demonstrated by this research greatly improves our understanding of the ability of our xylanase to promote animal growth via a prebiotic mechanism.
In addition to this research, complementary analytical work was done revealing several aspects where our xylanase differs from others. For example, when compared to a reference xylanase of the same family (GH11), our xylanase is able to degrade more arabinoxylan (under in vitro simulation of the GIT more than 50% of pure arabinoxylan was degraded to prebiotic AXOS).
This efficiency difference is crucial when thinking about the animal digestive process, where we are highly limited by time and conditions. The expected benefit of the xylanase requires that nutrient release and AXOS formation starts during the early stages of digestion to ensure that the animal can utilise the additional released nutrients, and that AXOS are formed to be fermented by microflora, resulting in improvements in animal performance.
So, to summarise, for us the important takeaways from this research are:
- fibre breakdown products (AXOS) by the use of our xylanase were detected, identified and quantified during the animal's digestion process
- microflora changes related with AXOS fermentation in vivo by the use of our xylanase have been proven
- alterations by the use of our enzyme on the produced SCFAs were shown and measured in vivo
- the amount of arabinoxylan degraded by our enzyme was measured in in vitro conditions that mimic the in vivo digestion process and compared with a reference xylanase enzyme, showing higher arabinoxylan degradation to AXOS for our xylanase
[Feedinfo] In the current market environment, enzymes are an important nutritional tool to manage production costs. Does this research impact your recommendations on the use of fibre-degrading enzyme products?
[Natalia Soares] Knowledge and research on how to improve nutrient and fibre utilisation in monogastric diets is a must, especially during a period of rising energy costs, high raw material prices, variable raw material quality and/or low raw material availability due to geopolitical instability.
Overall, the limited grain supply means that animal nutrition must rely more and more on alternative fibre-rich raw materials. The use of such by-products has the potential to lower the production costs and contribute to a more sustainable animal farming. Nutritional tools, such as enzymes, therefore, are essential to offset the potential negative impact on animal production efficiency.
The low energy contribution of NSPs, especially in poultry, points to the need for improvements in fibre degradation. In this context, the ability of specific FDEs - which form part of Huvepharma's product range - to improve NSP fermentation and promote gut health via a prebiotic mechanism whilst mitigating the anti-nutritive effects of NSP, is expected to become more important.
This research underlines our recommendations about the use of our xylanase-based products and delivers valuable insight into the mode of action of this enzyme and its efficiency. We believe we have a nutritional tool that can help our customers improve the productivity of their operations both by optimising animal performance and through the formulation of more cost-effective diets.