Effects of various levels of L-ascorbic acid (L-AA) (12 and 25 mg) in ovo injected at 17 and 18 days of incubation, on embryonic and posthatch physiological variables in Ross 708 broilers were investigated. There were no significant treatment effects on the hatchability or serum concentrations of L-AA in the broilers. Eye L-AA concentrations were higher, and plasma nitric oxide levels were lower in male chicks treated in ovo with 12 mg of L-AA. While exposed to elevated atmospheric NH3 levels, the in ovo injection of 12 mg of L-AA increased the body weight gain and decreased the feed conversion ratio of the broilers from 0 to 28 days of age (doa). Decreased tracheal attenuation incidence was also experienced at 0 doa in the in ovo injected L-AA groups, and tracheal inflammation was significantly reduced at 28 doa in response to the in ovo injection of 12 mg of L-AA.