The equilibrium sublimation of L-and DL-threonine was studied using Knudsen effusion mass spectrometry in the temperature range of 371−462 K. It was found that the only component of the vapor in both cases is a monomeric molecule. The vapor pressure was determined using the Knudsen effusion method, and the combination of this method with mass spectrometric measurements of total ion currents allowed for the recommendation of pressure equations: ln(p/Pa, L-threonine) = −(17391 ± 198)/T + (38.407 ± 0.327) and ln(p/Pa, DL-threonine) = −(18022 ± 197)/T + (40.525 ± 0.331) in the temperature ranges of 385−462 and 371− 454 K, respectively. The enthalpy of sublimation of L-threonine was determined using the methods of the second and third laws of thermodynamics; based on the analysis of the obtained in this work and literature data, the value of 149.2 ± 3.2 kJ•mol −1 at 298.15 K was recommended. The enthalpy of sublimation of DL-threonine (151.1 ± 2.1 kJ•mol −1 at 298.15 K) was determined for the first time by the second law method. The obtained results provide a data basis for industrial development.