ABSTRACT: BACKGROUND:Using metabolomics technique to analyze the response to a dietary intervention generates valuable information concerning the effects of the prescribed diet on metabolic regulation. To determine whether low calorie diet (LCD)-induced weight reduction causes changes in plasma metabolites and metabolic characteristics. METHODS:Overweight subjects consumed a LCD (n?=?47) or a weight maintenance diet (control, n?=?50) in a randomized, controlled design study with a 12-week clinical intervention period. Plasma samples were analyzed using an UPLC-LTQ-Orbitrap MS. RESULTS:The 12-week LCD intervention resulted in significant mild weight loss, with an 8.3% and 10.6% reduction observed in the visceral fat area (VFA) at the level of the lumbar vertebrae L1 and L4, respectively. The LCD group showed a significant increase in the mean change of serum free fatty acids compared to the control group. In the LCD group, we observed a significant increase in the acylcarnitine (AC) levels, including hexanoylcarnitine, L-octanoylcarnitine, 9-decenoylcarnitine, trans-2-dodecenoylcanitine, dodecanoylcarnitine, 3,5-tetradecadiencarnitine, cis-5-tetradecenoylcarnitine, 9,12-hexadecadienoylcarnitine, and 9-hexadecenoylcarnitne at the 12-week follow-up assessment. When the plasma metabolite changes from baseline were compared between the control and LCD groups, the LCD group showed significant increases in hexanoylcarnitine, L-octanoylcarnitine, trans-2-dodecenoylcanitine, and 3,5-tetradecadiencarnitine than the control group. Additionally, the changes in these ACs in the LCD group strongly negatively correlated with the changes in the VFA at L1 and/or L4. CONCLUSION:Mild weight loss from 12-week calorie restriction increased the plasma levels of medium- and long-chain ACs. These changes were coupled with a decrease in VFA and an increase in free fatty acids. TRIAL REGISTRATION:NCT03135132 ; April 26, 2017.