Deep eutectic solvents in the extraction of plant metabolites have found many advantages, such as low toxicity, biodegradability, low cost and ease of preparation over the conventional methods. This work aims to compare natural deep eutectic solvents in extraction and optimization of oleoresin from Ferula gummosa and determining its chemical and structure properties. Box-Behnken design was applied to optimize the extraction of oleoresin from Ferula gummosa using eutectic solvents. The variables of extraction were extraction time, temperature, and ratio of eutectic solvents. Six mixtures of eutectic solvents including choline chloride/urea, acetic acid, lactic acid, formic acid, formamide and glycerol at ratios of 2:1 and 3:1 were evaluated. The highest yields were obtained for choline chloride/formic acid, choline chloride/formamide. The quadratic regression equation was set up as a predictive model with an R value of 0.85. The optimum condition was 6 h, 40 °C, and ratio 12.5% (w/v). No significant difference was found between the predicted and experimental yield. The main components of the oleoresin were β-pinene (40.27%), cylcofenchen (11.93%) and α-pinene (7.53%) as characterized by gas chromatography-mass spectrometry. The chemical structure study by spectroscopy showed that no solvents remained in the oleoresin. Therefore, F. gummosa oleoresin can be explored as a novel promising natural pharmaceutical ingredient extracted with eutectic solvents.