As non-vascular plants, which lack lignin for protection, bryophytes support themselves in harsh environment by producing various chemicals. In response to cold stress, lipids play a crucial role in cell adaptation and energy storage. Specifically, bryophytes survive at low temperatures by producing very long-chain polyunsaturated fatty acids (vl-PUFAs). However, a systematic knowledge and comprehensive understanding of the cold acclimation of bryophytes is limited. To overcome this obstacle, we performed lipid profiling using ultra-high performance liquid chromatography-quadrupole time of flight mass spectrometry (UHPLC-QTOF-MS) of two moss species ( Bryum pseudotriquetrum and Physcomitrium patens) cultivated at standard condition compared to those cultivated at cold stressed condition. The potential biomarkers were identified by multivariate statistical analysis in each species. In B. pseudotriquetrum, we found that the phospholipids and glycolipids increased significantly under cold stress, while storage lipids decreased. The accumulation of the lipids with high unsaturation degrees (i.e. at least one fatty acyl chain contains more than two double bonds) mostly appear in phospholipids and glycolipids. However, this trend cannot be observed in P. patens. This suggests that different moss species may undergo a different lipid metabolic pathway of cold adaptation. Our findings present a deeper understanding of how mosses are adapted to cold temperature and provide a basis for future studies.