Deciphering the differentiations of traditional Chinese medicine analogous formulae by parallel liquid chromatography-mass spectrometry coupled with microplate-based assays

For traditional Chinese medicine (TCM), its most common application form is TCM formulae that assemble herbs and other materials together to treat diseases. According to the shared herbs or similar indications, most TCM formulae (Fangji or FJ in Chinese) can be grouped into several categories, namely analogous TCM formulae. Similarities/dissimilarities between these analogous TCM formulae provide a unique opportunity to understand mechanisms of action of TCM and discover biologically active compounds or components, as well as rationalize clinical use of TCM formulae. In this work, high performance liquid chromatography-mass spectrometry (HPLC-MS) coupled with microplate-based rapid screening assays was used to carry out systematic chemical analysis of the Wu-Ling-San (WLS) formula group. A total of thirty-seven compounds were identified in WLS and their MS fragmentation pathways were clarified. Seven potential anti-inflammatory components were discovered in WLS and other three analogous formulae by nitric oxide assay-based screening. The differences in chemical profiles and related mechanisms of action of the WLS group were discussed, which were reported for the first time and would be helpful for further pharmacological studies. Our results showed that this method is very effective and powerful for the research into TCM analogous formulae.