The current study explores an emerging cardiac metric, heart rate fragmentation (HRF), as a novel biomarker for allostatic load (AL). HRF may better address the limitations of existing cardiac biomarkers (e.g., confounds and interpretation consistency) in applied research settings, with nonclinical samples. The study’s objectives were: 1) can HRF represent response to psychological stress and 2) can resting HRF be used as a measure of predicting subclinical mental health symptoms. One hundred and fifty-six (n = 156; 75% female) undergraduate students were fitted with a chest band to monitor cardiovascular activity, and completed online demographic and psychosocial surveys in which they were grouped as healthy or displaying probable mental health symptoms (pMH; n = 94, 60.25%) based on respective inventory thresholds for depression, anxiety, and posttraumatic stress disorder. Cardiovascular activity was measured capturing the three R’s of cardiac vagal control: a resting baseline, a reactive acute stressor task, and a paced breathing recovery. Results supported the first hypothesis, in that that HRF significantly differentiated between each RRR condition (p < 0.001). While healthy and pMH individuals did not significantly differ within individual conditions, exploratory analyses revealed healthy individuals displayed significantly larger change in HRF reactivity between conditions (p’s < 0.001) in comparison to pMH, which displayed a more blunted pattern. Overall, this study establishes associations between HRF and mental health, and serves as a promising new biomarker that may identify AL in samples that may be otherwise considered “healthy”, while addressing the limitations of prior biomarkers in non-clinical studies.