Purpose: This study examines quality of life of medical students in Yemen by evaluating validity and reliability of the World Health Organization Quality of Life questionnaire (WHOQOL) and assessing potential influencing factors. Methods: This is a single-centered cross-sectional study conducted in Hadramout University College of Medicine, Mukalla, Yemen during the academic year of 2019. The WHOQOL questionnaire was distributed among medical students. For validity, item discriminate validity and confirmatory factor analysis were assessed and for reliability, Cronbach's α test was examined. Independent sample t -test and one-way Analysis of Variance (ANOVA) were used to examine the academic level, gender, academic performance, and basic life necessities including water, electricity supply, sewage treatment and type of residence. Results: A total of 495 medical students have responded to this questionnaire which has demonstrated an adequate validity and good reliability. The mean score for students' self-rating of their quality of life in the major domains was found to be in a descending order (Mean ± SD): psychological health (55.18 ± 17.84), environmental (52.14 + 17.60), physical health (48.15 + 14.73) and social relations (45.09 ± 20.81). Demographics and basic life needs exhibit relationship with Quality of Life among medical students. Conclusion: The WHOQOL-BREF is a valid and reliable tool among medical students in Hadramout University. Demographics and basic life needs seem to impact Yemeni medical students' Quality of Life. Wellness and mentoring programs should be considered to ameliorate effects related to deteriorating medical students' Quality of Life in Hadramout University.