Metabolism of Dietary Nitrate and its Safe Use for Mitigating Methane Emissions from Sheep

Author(s)
de Raphelis-Soissan, Victoire
Hegarty, Roger
Li, Li
Godwin, Ian
Publication Date
2017-10-27
Abstract
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Abstract
<p>Supplementing ruminants with dietary nitrate (NO<sub>3</sub>) is an effective methane mitigation strategy if it can be managed so as to not expose ruminants to any risk of clinical nitrite (NO<sub>2</sub>) toxicity. The objective of this thesis was firstly to deepen the understanding for NO<sub>3</sub> metabolism in sheep and secondly to develop practical strategies to reducing risk of NO<sub>2</sub> toxicity in sheep supplemented with dietary NO<sub>3</sub>.</p> <p>It has been previously established, that in the rumen NO<sub>3</sub> is reduced to NO<sub>2</sub> and then to NH<sub>3</sub>, and that supplementing with excessive amounts of NO<sub>3</sub> can expose ruminants to NO<sub>2</sub> toxicity due to the absorption of NO<sub>2</sub>. This thesis reports a series of five investigations of NO<sub>3</sub> metabolism by sheep and identifies:</p> <p> Nitrate, like urea, is ‘recycled’ within the ruminant. Transfer of ruminal <sup>15</sup>NO<sub>3</sub><sup>-</sup>-N into the blood and transfer of blood NO<sub>2</sub>-N into the rumen being quantified. Only 20% of rumen NO<sub>3</sub><sup>-</sup>and 30% of blood NO<sub>2</sub><sup>-</sup> were recovered in urine.</p> <p>That in hourly fed sheep approximately 90% of dietary NO<sub>3</sub><sup>-</sup> was rapidly converted to NH<sub>3</sub> in the rumen, with the remainder leaving the rumen by absorption into the bloodstream or passage to the lower gastro-intestinal tract.</p> <p> Within the rumen, the conversion of NO<sub>3</sub><sup>-</sup>to NH<sub>3</sub> is neither simple nor complete. <i>In vitro</i> and in-vivo studies showed NO<sub>3</sub><sup>-</sup>is reduced to gaseous nitrous oxide (N<sub>2</sub>O) and N<sub>2</sub>O may be further metabolised to N<sub>2</sub> gas by the rumen microbiota. Approximately 0.04% and 3.0% of dosed NO<sub>3</sub><sup>-</sup>-N was recovered over 10 h from sheep as N<sub>2</sub>O and N<sub>2</sub> respectively, and this was not affected by whether sheep had prior adaption to NO<sub>3</sub><sup>-</sup> or not, identifying denitrification as a reaction not previously reported from the rumen.</p> <p>From this understanding and a review of the literature on ruminant NO<sub>3</sub> metabolism, eight critical control points for reducing the risk of nitrite toxicity (methaemoglobinaemia), were identified and the potential for manipulating five of these evaluated. </p> <p> Reducing the rate at which NO<sub>3</sub> became available to the rumen biota by coating calcium nitrate with paraffin wax significantly reduced blood methaemoglobin level (MetHb; an indicator of NO<sub>2</sub> toxicity) in sheep supplemented with NO<sub>3</sub>.</p> <p>The extent of methaemoglobinaemia could also be reduced by the daily ration being consumed at shorter intervals rather than in a single bout, and this established that feed management is pivotal to safe feeding of NO<sub>3</sub><sup>-</sup>containing diets. </p> <p> Enhancing the rumen’s capacity to reduce potentially toxic NO<sub>2</sub> <sup>-</sup>by supplying <i>Propionibactericum acidicpropionici</i> as a direct fed microbial was ineffective in reducing blood MetHb or NO<sub>2</sub><sup>-</sup>concentration of sheep fed NO<sub>3</sub><sup>-</sup> supplemented diets.</p> <p> Attempts to increase the rate of removal of NO<sub>2</sub><sup>-</sup>from the rumen by providing a substrate (glycerol) to stimulate NADH availability in the rumen, and accelerate the nitrite reductase enzyme system did not reduce the concentration of NO<sub>2</sub> in incubations of rumen contents supplemented with NO<sub>3</sub><sup>-</sup>.</p> <p> We found no evidence that adapting sheep to dietary NO<sub>3</sub><sup>-</sup> protected them against NO<sub>2</sub><sup>-</sup> toxicity. Indeed, <i>in vitro</i> more NO<sub>2</sub><sup>-</sup> accumulated in incubation when donors where adapted to dietary NO<sub>3</sub><sup>-</sup>. Also, no signs of reduced MetHb were noticed after several weeks of NO<sub>3</sub><sup>-</sup>supplementation <i>in vivo</i>.</p> <p>Other critical control points such as regulating microbial uptake of NO<sub>3</sub> and ruminal absorption of NO<sub>3</sub> and NO<sub>2</sub> were unable to be assessed in this thesis.</p> <p>The studies reported here also confirmed the practical impacts of NO<sub>3</sub> as an effective supplement for reducing enteric methane emissions and increasing wool growth of sheep. As well as providing a better understanding of NO<sub>3</sub><sup>-</sup>metabolism, studies also showed that the greenhouse gas (GHG) abatement impact of methane mitigation may be partly offset by an associated production of the potent GHG, N<sub>2</sub>O. Discovery of the production of N<sub>2</sub>O and N<sub>2</sub> from NO<sub>3</sub><sup>-</sup>in the rumen and identification of recycling of blood NO<sub>2</sub><sup>-</sup> to the rumen has expanded our understanding of NO<sub>3</sub><sup>-</sup>metabolism. Coating NO<sub>3</sub><sup>-</sup>to decrease the rapidity of NO<sub>3</sub><sup>-</sup> release in the rumen as a strategy to reduce NO<sub>2</sub> toxicity was effective but needs further investigation. The applicability of feed grade NO<sub>3</sub><sup>-</sup>as a commercially available feed additive will also depend on the cost of NO<sub>3</sub> and the additional cost of the technology to ensure its safe feeding, compared to the cheaper alternative non-protein nitrogen source, urea.</p>
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Title
Metabolism of Dietary Nitrate and its Safe Use for Mitigating Methane Emissions from Sheep
Type of document
Thesis Doctoral
Entity Type
Publication

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