Arthritis is a common chronic joint disorder, with general symptoms including stiffness and joint pain. β-methylphenylalanine is a well-known non-proteogenic unnatural amino acid. This study analyzes the anti-arthritic activity of β-methylphenylalanine in experimental rats. The experimental groups were as follows: group I, sham; group II, control; group III, 100 mg/kg of β-methylphenylalanine; and group IV, 200 mg/kg of β-methylphenylalanine. Lipid peroxidation, glutathione peroxidase (Gpx), reduced glutathione (GSH), superoxide dismutase (SOD), catalase, prostaglandin E₂ (PGE₂), matrix metalloproteinase-3 (MMP-3), ceruloplasmin, zinc, copper, mRNA, and protein expression of inducible nitric oxide synthase (iNOS) and nuclear factor-kappa B (NF-κB) were determined. Supplementation with β-methylphenylalanine significantly reduced lipid peroxidation, copper, PGE₂ and MMP-3 levels, whereas GSH, Gpx, catalase, SOD and zinc levels were increased. Supplementation with β-methylphenylalanine significantly reduced NF-κB mRNA expression by 26% and 47.8% in groups III and IV, respectively (P < 0.045), while iNOS mRNA expression was reduced by 14.3 and 47.6% in groups III and IV, respectively. NF-κB and iNOS protein expression increased by 160% and 120% respectively, in the control rats compared to the sham rats. However, supplementation with β-methylphenylalanine significantly reduced NF-κB protein expression by 27% and 50% in groups III and IV, respectively, while iNOS protein expression was reduced by 22.7% and 45.4% in groups III and IV, respectively. Taken together, our data show that supplementation of β-methylphenylalanine was effective against arthritis in a rat model.