Five new glycerol esters including 2-acetyl-1-coumaroyl-3-cinnamoylglycerol (13), (+)-2-acetyl-1-feruloyl-3-cinnamoylglycerol (14), (-)-2-acetyl-1-feruloyl-3-cinnamoylglycerol (15), 2-acetyl-1,3-dicinnamoylglycerol (16), and (-)-2-acetyl-1-(E)-feruloyl-3-(3″(ζ),16″)-dihydroxy-palmitoylglycerol (17) were isolated from methanolic extract of Wuhan propolis. The chemical structures of the five new compounds were confirmed by (1)H and (13)C NMR and HR-MS spectra. Twelve minor constituents in the Wuhan propolis extract were tentatively identified by UPLC-Q-TOF-MS, according to their characteristic UV spectrum, retention times, and accurate MS data. The anti-inflammatory activities of the five new compounds were studied in lipopolysaccharide (LPS)-stimulated RAW 264.7 mouse macrophage cells. All five compounds exhibited remarkable inhibitory effects on interleukin (IL)-1β, IL-6, and cyclooxygenase (COX)-2 mRNA expressions at 10 and 100 μM, suggesting that Wuhan propolis may serve as a potential anti-inflammatory functional food ingredient or nutraceutical.