Abstract Background Mitochondria play a crucial role in cellular processes, such as energy metabolism, reactive oxygen species (ROS) generation and apoptosis. Mitochondrial dysfunction induced by stress has been implicated in various health conditions. Circulating cell-free mitochondrial DNA (CFC-MT-DNA) has emerged as a potential biomarker reflecting mitochondrial damage under stress. Methods To evaluate the association between CFC-MT-DNA levels and human stress through a systematic review and meta-analysis of case-control studies. A comprehensive literature search was conducted across PubMed, Web of Science and ScienceDirect databases up to September 2023. Eight eligible studies assessing CFC-MT-DNA levels in stressed vs. control individuals were included. Data were analysed using RevMan version 5.4 software. Results The meta-analysis revealed significantly elevated CFC-MT-DNA levels in individuals experiencing stress (p = 0.03), particularly in psychological stress-related conditions, such as bipolar disorder (BD) and major depressive disorder (MDD). However, no significant increase was observed in physiological stress conditions, including diabetes and sports training. High heterogeneity (I2 = 96%) was observed across studies. Conclusions CFC-MT-DNA shows promise as a non-invasive biomarker for psychological stress. Further longitudinal and mechanistic studies are needed to clarify its role across different types of stress and its potential clinical utility.