We report for the first time either in vitro or in vivo the phase I hydroxylation and phase II conjugation metabolic pathways of an organophosphate flame retardant, triphenyl phosphate (TPHP), in addition to diphenyl phosphate (DPHP) metabolite formation. Using a chicken embryonic hepatocyte (CEH) assay, TPHP was phase I metabolized to p- and m-hydroxy-TPHP metabolites, which were largely present in the assay medium and cells as phase II conjugates with glucuronic acid. After treatment with β-glucuronidase, deconjugated p-OH-TPHP was present in both the medium and cells at increasing concentrations of 0.073 ± 0.003, 1.95 ± 0.03, and 2.10 ± 0.09 nmol/well at CEH incubation time points of 0, 12, and 36 h, respectively. Similarly, after β-glucuronidase treatment, there were increasing m-OH-TPHP concentrations of 0.0050 ± 0.0005, 0.18 ± 0.01, and 0.18 ± 0.01 nmol/well. p-OH-TPHP at 36 h accounted for 60% of the initial TPHP treatment concentration, which was 3.5- or 12-fold greater than that of the DPHP or m-OH-TPHP metabolites, respectively. Overall, in TPHP-exposed organisms, this study demonstrates the importance of phase I and II metabolic processes in the biological fate of TPHP.