A high-speed counter-current chromatography (HSCCC) method, using a two-phase solvent system composed of ethyl acetate/n-butanol/methanol/water (5:1:1:5, v/v/v/v), was successfully established to separate the five iridoid glucosides 7-O-ethyl sweroside (1), secologanin dimethylacetal (2), adinoside F (3), (7R)-secologain n-butyl methyl acetal (4) and adinoside G (5) from Lonicerae Japonicae Flos. Their purities were 96.8%, 98.5%, 93.3%, 98.0% and 99.9%, respectively. All the iridoid glucosides were identified by HR-ESI-MS, 1D and 2D NMR. Compounds 3 and 5 are new iridoid glucosides. The anti-inflammatory tests showed that compounds 1–5 all expressed moderate inhibitory effects on β-glucuronidase release in rat polymorphonuclear leukocytes (PMNs) induced by platelet-activating factor (PAF) with IC₅₀ values ranging from 4.52 to 6.50 µM, while the antibacterial assays demonstrated that all the compounds displayed mild inhibitory activities against Staphylococcus aureus ATCC 25923 with MIC values ranging from 13.7 to 26.0 µg/mL.

One new lignan, loniceralanside A (1), together with three known compounds (2–4), were purified from Lonicerae Japonicae Flos. Their structures were determined by HR-ESI-MS, 1 D and 2 D NMR data. Compounds 2–4 were separated from the plant for the first time. The biological test showed that compound 1 had significant anti-inflammatory activity with inhibiting the release of β-glucuronidase induced by platelet-activating factor (PAF) in rat polymorphonuclear leukocytes (PMNs) (IC₅₀ = 3.05 µM). Graphical Abstract