Water recovery by membrane distillation (MD) is an attractive alternative to existing urine treatment systems because it could improve the water recovery rate and reliability in space missions. However, there are few studies of urine MD, particularly on the removal of the remaining contaminants from distillate water and the assessment of its long-term performance. In this study, the influences of various operation parameters on distillate water quality and operation stability were investigated in batch mode. The low pH of feedstock reduced the conductivity and total ammonium nitrogen (TAN) in distillate water because the low pH promoted the ionization of ammonia to ammonium ions. However, the low pH also facilitated the formation of free chlorine hydride, which resulted in the minor deterioration of the conductivity in the distillate due to the increasing volatility of chlorine hydride in the feedstock. Thirty batches of vacuum membrane distillation (VMD) experiments demonstrated that the permeate flux and the distillate water quality slightly decreased due to the small range of membrane wetting but still maintained an over 94.2% and 95.8% removal efficiency of the total organic carbon (TOC) and TAN, and the conductivity was <125 μs cm−1 in the distillate water after 30 test batches. VMD is a feasible option for urine treatment in space missions.