Background: Favorable health benefits of an active lifestyle have been clearly documented within the context of occupational health. However, a knowledge gap exists regarding the monitoring and comparison of micronutrient status across varying levels of physical activity (PA). This study aimed to investigate the association of PA level with micronutrient status and the associated health biomarkers among a cohort of Austrian bank employees. Methods: Using a cross-sectional design, this study involved the participation of bank employees (n = 123; average age: 43 years; 49% males) from the federal state of Tyrol, located in the western part of Austria. To assess PA levels and sedentary behavior, the Global Physical Activity Questionnaire (GPAQ; developed by the WHO) was administered. Accordingly, participants were categorized into three groups: low PA, moderate PA, and high PA. Participants’ blood samples were obtained to measure blood levels of micronutrients, homocysteine, and CoQ10. The values of vitamins and minerals in whole-blood were compared to sex-specific reference ranges and grouped into three categories: below, within, or exceeding the reference range. Results: The prevalence of a high PA level was 61%, while 18% of participants had a low PA level. Overweight/obesity was significantly less prevalent among participants with high PA levels (22%) compared to those with moderate (50%) and low (50%) PA levels (p = 0.045). No significant differences between PA levels were found for sex, age, diet type, homocysteine, or CoQ10 markers (p > 0.05). There was no significant PA-based difference in blood concentrations of most vitamins and minerals (p > 0.05), except for vitamin D (p = 0.001) among females, as well as selenium (p = 0.040) and vitamin B12 (p = 0.048) among males. Conclusion: The present findings offer initial insights into the link between PA behaviors, micronutrient status, and health, highlighting potential implications in occupational health and lifestyle, specifically in developing tailored approaches based on PA levels.