Atractylodis Macrocephalae Rhizoma (AMR) is one of the commonly used medicinal and edible herbs in China. To prevent infestation of moths and mildew, chemical-fumigation is applied during post-harvest processing, which would produce harmful substances. Ultralow oxygen (ULO) storage, as a potential controlled atmosphere preservation method for Traditional Chinese Medicines (TCMs), has no residue of chemical hazards, but its impact on chemical composition of TCMs is unclear. Here, to evaluate the impact of ULO preservation on the quality of AMR, a UPLC-QTOF-MS/MS-based non-targeted metabolomics approach was employed to screen out the differential components after AMR had been stored under ULO conditions for 18 months. The differential components were identified, and further semi-quantified with peak areas. Additionally, the differential components with known activity were quantified using HPLC-DAD and GC-TQ-MS/MS analysis. The molecular weight distribution and monosaccharide composition of polysaccharides were analyzed using HPGPC-ELSD and HPLC-DAD to assess the impact of ULO on the carbohydrates. As it showed, 20 differential components of AMR under ULO were screened and identified when compared to normal atmosphere (NA) with 21 % oxygen concentration. As storage duration increased, the levels of atractylenolides I, II, and III in both ULO and NA storage conditions increased, with significant differences in their total content (16.06 mg/g in ULO vs. 47.57 mg/g in NA). Confirmatory experiments revealed that atractylone is the precursor compound of atractylenolides and the total content of atractylenolides I, II, and III produced from atractylone in ULO storage was only 62.12 % of that in NA at 18th month. The total sugar content of AMR polysaccharides in ULO storage was 107.28 % of that in NA and the molar ratios of monosaccharides were also relatively constant. All the results indicated that ULO preservation slowed down the chemical transformation of AMR, including small molecules and polysaccharides. Therefore, the quality of AMR stored in ULO is more stable and it is proposed to be an efficient preservation method for AMR.