Molecular Networking-Guided Isolation of Anti-Inflammatory Isolariciresinol and Seco-Isolariciresinol Type Lignans from Alnus japonica

Abstract

A comparative analysis of UPLC-DAD and MS chromatograms revealed distinct chemical profiles between the methanolic extracts of the branch and cortex of Alnus japonica. MS/MS-based molecular networking further identified unique molecular features present in the branch extract, which facilitated the targeted isolation of five previously undescribed lignan derivatives. The structures of these compounds were elucidated using 1D and 2D NMR spectroscopy, along with high-resolution mass spectrometry (HRMS). The anti-inflammatory potential of the isolated compounds was evaluated by measuring the inhibition LPS-induced nitric oxide (NO) production in RAW 264.7 macrophage cells. Notably, compounds 1-5 exhibited significant NO inhibitory activity, with IC₅₀ values of 3.7 and 7.4 μM.

1

Comparison of UPLC-DAD (A) and total ion chromatograms (TIC) (B) of the branch and cortex extracts of A. japonica. Peaks highlighted in red were exclusively detected in the branch extract.

2

Molecular network of the branch and cortex extracts of A. japonica. An additional cluster of nodes in the m/z range of 700–1000, highlighted in red, was observed exclusively in the branch extract compared to the cortex extract.

3

Molecular network of the solvent-partitioned fractions of A. japonica.

4

Chemical structures of compounds 1–5. The structures of compounds 1–4 are depicted as relative configurations.

5

Key HMBC and COSY correlations of compounds 1–4.

6

J-value based configuration analysis and key ROE correlations of compound 1 and 2.

7

J-value based configuration analysis and key ROE correlations of compound 3 and 4.

8

Key HMBC, COSY, and NOE correlations of compound 5.

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Annotation of isolated compounds 1–7 on the molecular network.