Omega-3 fatty acids (ω-3 FAs) exert anti-inflammatory effects, including the downregulation of pro-inflammatory cytokines, eicosanoids, and insulin-like growth factor-1. Therefore, they may improve acne severity as an adjunct treatment. However, there is a paucity of data regarding patients' existing deficits. The aim of this study was to determine ω-3 FA levels in acne patients in correlation with self-reported dietary preferences and clinical severity. A single-center, cross-sectional study of 100 acne patients was conducted. Patients' blood parameters, including ω-3 FAs levels, were assessed using the HS-omega-3 Index® in erythrocytes (Omegametrix® GmbH, Martinsried, Germany). Dietary preferences were assessed using a standardized food frequency questionnaire. Clinical dermatologic evaluation was performed using the Investigator Global Assessment (IGA) of acne. The values of the HS-omega-3 Index® were outside the recommended range of 8-11% in 96 patients (mean 5.15%), independent of the clinical severity or affected anatomic sites. A severe deficit (HS-omega-3 Index® < 4%) was seen more commonly in men than in women (p = 0.021). The regular consumption of legumes was significantly associated with higher ω-3 FA levels (p = 0.003), as was oral ω-3 FA supplementation (p = 0.006) and the lack of sunflower oil intake (p = 0.008). This pilot study demonstrated a deficit of ω-3 FAs in a German acne cohort. Higher ω-3 FAs levels were observed in patients with regular legume intake and oral ω-3 FAs supplementation. Further prospective studies are needed to investigate whether the clinical severity of acne improves in patients with normal HS-omega-3 Index®.