The Cost of Deamination in Reduced-Crude Protein Broiler Diets

Reductions in crude protein (CP) contents of broiler diets have been realised for decades by routine additions of unbound (synthetic or crystalline) methionine, lysine and threonine, and are likely to continue as inclusion costs for the balance of amino acids become more feasible. Reduced-CP diets have the potential to provide environmental advantages from attenuated nitrogen and ammonia outputs (Nahm, 2007), bird welfare, from enhanced litter quality and lower incidences of foot-pad dermatitis (Dunlop et al., 2016), and flock health, from less undigested protein entering the large intestine to fuel the proliferation of potential pathogens (Wilkie et al., 2005). However, there appears to be a threshold where tangible CP reductions of more than 3 to 4 percentage units negatively influence growth performance, especially FCR, and this is associated with increased fat deposition (Belloir et al., 2017). Many possible explanations have been advanced and numerous strategies evaluated in attempts to lower this threshold, but the problem of compromised growth performance, especially in wheat-based diets, remains. However, the ‘cost of deamination’ may be contributing towards compromised growth performance in birds offered reduced-CP diets.

 

Dietary amino acid imbalances generated inferior growth performance in the Snetsinger and Scott (1961) study. This negative impact was partially alleviated by glycine, which was attributed to glycine enhancing the excretion of excess nitrogen (N) via the uric acid cycle. However, the effects of glycine and glutamic acid were shown to be additive in this context (Maddy et al., 1960). Additions of imbalanced amino acid mixtures to low protein diets was investigated by Hill and Olsen (1963) who concluded that the resultant depressions in weight gain stemmed from deamination of relatively large quantities of amino acids. The blend of unbound and protein-bound amino acids in reduced-CP diets almost certainly leads to amino acid imbalances at sites of protein synthesis, and any surplus of amino acids require deamination. The principal mechanism for this is oxidative deamination in the liver, which generates ammonia that demands detoxification. Ammonia detoxification is a condensation reaction in which ammonia and glutamic acid are converted to glutamine. The reaction is driven by glutamine synthetase, which is present in both mammalian (Hakvoort et al., 2017) and avian (Watford and Wu, 2005) species. In poultry, glutamine is incorporated into the Krebs uric acid cycle and N is excreted as uric acid, which is an energy consuming process that requires glycine inputs (Salway, 2018), where serine, and possibly threonine, may serve as glycine precursors.

Interestingly, Mapes and Krebs (1978) investigated the rate-limiting factors in hepatic uric acid synthesis in chickens and suggested that glutamate and glutamine are fundamental to ammonia detoxification and N excretion. Energy considerations are also involved in these pathways as synthesis and excretion of one molecule of uric acid involves the loss of one glycine molecule, which has the potential to yield 12.5 molecules of ATP (Salway, 2018).

Free amino acid concentrations in portal and systemic plasma in birds offered 21.5% and 16.5% CP wheat-based diets were determined by Yin et al. (2019). In this instructive study, portal concentrations were higher than systemic concentrations, but both followed similar patterns. The reduction in dietary CP generated an average 30.9% increase in glutamine levels, which may have resulted from increased condensations of ammonia plus glutamic acid into glutamine. Concentrations of glycine equivalents declined by 23.6% which may reflect increased inputs of glycine and serine into the Krebs uric acid cycle. In contrast, threonine concentrations rose by 28.0%, which is not indicative of threonine being a glycine precursor as glycine levels decreased by 27.7% in this study.

Ammonia intoxication arises when an excess of ammonia is produced or its removal is retarded and ammonia interferes with metabolism. Indeed, high blood ammonia levels depressed feed intakes via mechanisms involving the central nervous system in rats offered amino acid imbalanced diets (Noda, 1975; Noda and Chikamori, 1976). Thus, there is the possibility that amino acid imbalances and inadequacies in reduced-CP diets result in deamination of excess amino acids and, in turn, lead to the accumulation of ammonia in birds, which has negative impacts. Reduced-CP diets contain substantially less glutamic acid, glutamine and glycine than conventional diets so one implication is that inadequate glutamic acid concentrations in reduced-CP diets are impeding ammonia detoxification and inadequate concentrations of glycine, or glycine equivalents, are retarding the Krebs uric acid cycle and uric acid excretion. Alternatively, imbalances of unbound and protein-bound amino acids in reduced-CP diets may be generating substantial excesses of amino acids that require deamination that is not being adequately met.

 

Data generated by both Namroud et al. (2008) and Ospina-Rojas et al. (2014) provide support for the proposal that the hepatic oxidative deamination of amino acids with the liberation of ammonia in reduced-CP broiler diets may result in excessive plasma ammonia concentrations, which has negative impacts on growth performance. A significant increase in systemic plasma ammonia concentrations of 14.5% (0.71 versus 0.62 mg/100mL) following a reduction in dietary CP from 230 to 170 g/kg was reported in broiler chickens by Namroud et al. (2008). Moreover, it may be deduced from this study that there were negative linear regressions between mean systemic plasma ammonia concentrations and 28-day body weights (r = -0.982; P < 0.001) and 10 to 28-day feed intakes (r = -0.962; P < 0.001) in broiler chickens offered eight dietary treatments with different CP levels and inclusions of unbound amino acids. In addition, there was a quadratic relationship (r = 0.941; P = 0.004) between ammonia concentrations and FCR as illustrated in Figure 1.

Subsequently, Ospina-Rojas et al. (2014) recorded systemic plasma concentrations of ammonia and the transition from 220 to 190 g/kg CP diets significantly increased ammonia concentrations by 59.4% (7.27 versus 4.56 mg/dl). Ten different combinations of valine, isoleucine, arginine and glycine were added to the 190 g/kg CP diet to provide a total of twelve dietary treatments. It may be deduced that there was a quadratic relationship (r = 0.799; P = 0.015) between mean ammonia plasma concentrations and 21-day weight gain, where increasing ammonia concentrations were associated with a decline in weight gains, as shown in Figure 2.

Figure 1 - Quadratic relationship (r = 0.941; P = 0.004) between systemic NH3 levels (mg/100 mL) and 10 to 28-day FCR in broiler chickens (Namroud et al. 2008).

Figure 2 - Quadratic relationship (r = 0.799; P = 0.015) between systemic NH3 levels (mg/dl) and 21-day weight gains in chickens (Ospina-Rojas et al. 2014).

Figure 3 - Quadratic relationship (r = 0.919; P < 0.001) between sampling time and systemic NH3 levels (mg/100 ml) where y = 0.346 + 0.157*hours – 0.025*hours² (Okumura and Tasaki, 1969).

The determination of ammonia concentrations in portal or systemic plasma per se is straightforward but is complicated by the fact that plasma ammonia concentrations are volatile over the sampling time duration. Okumura and Tasaki (1969) documented the volatility of systemic portal ammonia concentrations in poultry and it may be deduced that these researchers detected a quadratic relationship (r = 0.919; P < 0.001) between elapsed sampling time and systemic ammonia plasma concentrations in birds offered diets containing 150, 200 300 and 400 g/kg casein. It may be calculated from the regression equation that ammonia plasma levels were 0.35 mg/100 ml at zero hours, peaked at 0.59 mg/100 ml after 3.14 hours and had returned to 0.39 mg/100 after six hours, as shown in Figure 3. This means that the interpretation of plasma ammonia concentrations will be best addressed by an ANCOVA, or an analysis of covariance, which is a blend of standard analyses of variance with quadratic regressions. The accurate interpretation of plasma ammonia concentrations may prove pivotal in investigations into the likely cost of deamination and, in extreme cases, the possibility of ammonia toxicity in birds offered reduced-CP diets.

ACKNOWLEDGEMENTS: The authors would like to acknowledge the encouragement, guidance and financial support provided by both AgriFutures Chicken-meat and Evonik Nutrition & Care GmbH.