There is a need of experimental studies on biomarkers in patients with anorexia nervosa (PAN), especially in the context of stress, in order to foster understanding in illness maintenance. To this end, the cortisol response to an acute stressor was investigated in n = 26 PAN (BMI: 19.3 ± 3.4 kg/m2), age, and gender matched to n = 26 healthy controls (HC; BMI: 23.08 ± 3.3 kg/m2). For this purpose, salivary cortisol parameters were assessed in two experimental conditions: (1) rest/no intervention and (2) stress intervention (TSST; Trier Social Stress Test). In addition, psychological indicators of stress were assessed (Primary Appraisal Secondary Appraisal, Visual Analogue Scale, and Trier Inventory for the assessment of Chronic Stress), as well as psychological distress, depression, and eating disorder (ED) symptoms. A 2 × 2 × 8 ANOVA demonstrated elevated cortisol levels in PAN in the resting condition. In the stress intervention no significant group effect in terms of cortisol (F (1, 50) = 0.69; p = 0.410; [Formula: see text]). A significant condition (F (1, 50) = 20.50; p = 0.000; [Formula: see text]) and time effect (F(2.71, 135.44) = 11.27; p = 0.000; [Formula: see text]) were revealed, as well as two significant interaction effects. First: Condition × group (F (1, 50) = 4.17, p = 0.046; [Formula: see text]) and second: Condition × time (F (2.71, 135.44) = 16.07, p = 0.000, [Formula: see text]). In terms of AUCG, no significant differences between both groups were exhibited. Regardless, significant results were evinced in terms of an increase (AUCi: F(1, 50) = 20.66, p = 0.015, [Formula: see text]), baseline to peak (+20 min post-TSST: t5 = 16.51 (9.02), p = 0.029) and reactivity (MPAN = 0.73 vs. MHC = 4.25, p = 0.036). In addition, a significant correlation between AUCG and BMI: r (24) = -0.42, p = 0.027 was demonstrated, but not between AUCi and BMI (r (24) = -0.26, p = 0.20). Psychological indices suggested higher levels of chronic and perceived stress in PAN relative to HC. However, stress perception in the stress condition (VAS) was comparable. Additional analyses demonstrated that ED-symptoms are highly correlated with psychological distress and depression, but not with BMI. In addition, it could be demonstrated that reactivity is rather related to ED-symptoms and psychological burden than to BMI. In conclusion, PAN showed elevated basal cortisol levels at rest and exhibited a blunted cortisol reactivity to the TSST as evinced by salivary cortisol parameters. Further, it was shown that weight recovery influences reversibility of hypercortisolemia, i.e., cortisol levels normalize with weight gain. However, HPAA (hypothalamus-pituitary-adrenal axis) irregularities in terms of reactivity persist even at a BMI ≤ 19.3 (±3.4). Our data suggest that pronounced psychological burden in PAN, have a greater impact on the HPAA functionality (secondary to the ED) than BMI itself.