Jordan has a high prevalence of physical inactivity, a major risk factor for non-communicable diseases. Working mothers are a high-risk group who face significant barriers to physical activity and experience a decline in their quality of life. This study aims to evaluate the potential impact of physical activity adherence on health-related quality of life and physical fitness among a sample of working mothers. A Correlational, retrospective analytical design, was employed among a sample of working mothers (n=120), from Amman and was divided into two comparison groups. Data collection included a self-reported questionnaire using a demographic questionnaire, a health perception scale, the International Physical Activity Questionnaire short form, the International Fitness Scale, and the Quality of Life Brief questionnaire. The statistical analysis was independent t-test analysis, analysis of variance (One-way ANOVA), Pearson r correlation and regression used to estimate the prediction of dependent variables based on independent variables. The results found a significant positive relationship between physical activity and overall quality of life and its domains (physical, psychological, and environmental); however, there was no statistically significant difference in the social domain. Adherence to physical activity was a significant positive predictor of quality of life. Moreover, the health perception level was a significant positive predictor. A t-test revealed a statistically significant difference between the physically active group and a sedentary group of working mothers regarding changes in Health-Related Quality of Life; physically active mothers are more likely to have better physical (p < 0.024), psychological (p < 0.001), and overall quality of life (p < 0.011). Furthermore, Physical fitness significantly predicted physical activity in the study results. This study confirms the existence of positive correlations between physical activity and working women's quality of life and physical fitness. Mothers with higher levels of physical activity are more likely to have better physical, psychological, and environmental domains of quality of life and better physical fitness.