Background: Glycine is an important ammoniagenic amino acid, which is increased in acute liver failure (ALF). We have previously shown that ornithine phenylacetate (OP) attenuates ammonia rise and intracranial pressure in pigs suffering from ALF, but failed to demonstrate a stoichiometric relationship between change in plasma ammonia levels and excretion of phenylacetylglutamine in urine. The aim was to investigate the impact of OP treatment on the phenylacetylglycine pathway as an additional ammonia- lowering pathway. Methods: A well-validated and characterized large porcine model of ALF (portacaval anastomosis followed by hepatic artery ligation), which recapitulates the cardinal features of human ALF was used. Twenty-four female pigs were randomised into three groups: (i) Sham operated + vehicle, (ii) ALF + vehicle, and (iii) ALF + OP. Results: There was a significant increase in arterial glycine concentration in ALF (p< 0.001 compared with sham), with a 3-fold increase in glycine release into the systemic circulation from the kidney compared to the sham group. This increase was attenuated in both blood and the brain of the OP treated animals (p<0.001 and p<0.05, respectively), and the attenuation was associated with renal removal of glycine through excretion of the conjugation product phenylacetylglycine in urine (ALF+vehicle: 1060 ± 106 μmol/L, ALF+OP: 27625 ± 2670 μmol/L, P<0.003). Conclusion: Data from this study provides solid evidence for the existence of a novel, and additional pathway for ammonia removal in ALF, that involves glycine production and removal, which is targeted by OP.