Honey-processed Astragalus is a dosage form of Radix Astragalus mixed with honey by a traditional Chinese medicine processing method which strengthens the tonic effect. Astragalus polysaccharide (APS), perform its immunomodulatory effects by relying on the tonic effect of Radix Astragalus, therefore, the improved pharmacological activity of honey-processed Astragalus polysaccharide (HAPS) might be due to structural changes during processing. The molecular weights of HAPS and APS were 1,695,788 Da, 2,047,756 Da, respectively, as determined by high performance gel filtration chromatography combined with evaporative light scattering detection (HPGFC-ELSD). The monosaccharide composition was determined by ultra-performance liquid chromatogram quadrupole time-of-flight mass spectrometry (UPLC/ESI-Q-TOF-MS) after pre-column derivatization with 1-phenyl-3-methyl-5-pyrazolone (PMP). The results showed that the essential components were mannose, glucose, xylose, arabinose, glucuronic acid and rhamnose, is molar ratios of 0.06:28.34:0.58:0.24:0.33:0.21 and 0.27:12.83:1.63:0.71:1.04:0.56, respectively. FT-IR and NMR analysis of HAPS results showed the presence of uronic acid and acetyl groups. The anti-inflammatory activities of HAPS were more effective than those of APS according to the NO contents and the expression of IFN-γ, IL-1β, IL-22 and TNF-α in lipopolysaccharide (LPS)-induced RAW264.7 cells. This findings suggest that the anti-inflammatory and bioactivity improvement might be associated with molecular structure changes, bearing on the potential immunomodulatory action.

A novel method has been developed for rapid analysis of the major constituents in Radix Astragali and honey-processed Astragalus within 23 minutes using ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC/ESI-Q-TOF-MS). Separation analysis was performed on HSS T3 columns with a gradient elution of acetonitrile-water containing 0.1% formic acid at a flow rate of 0.3 mL min⁻¹. Methodology validation showed that the established method had good repeatability (RSD < 8.07%), intra-day precisions (RSD < 4.20%), and stability (RSD < 7.78%). Accurate molecular weight and characteristic fragment ions obtained from Q-TOF-MS provided a reliable criterion for the structural characterization of the major compounds in raw and honey-processed Astragalus. 35 major constituents, including flavonoids, isoflavan, pterocarpan, and saponins, were identified or tentatively characterized by comparing their retention times and MS spectra with those of authentic standards and literature data. Furthermore, quantitative analysis revealed that the content changes of chemical compositions could cause the tonic effect differences between raw and honey-processed Astragalus. The established method is rapid and reliable for simultaneous analysis of constituents in an extract of Radix Astragali. Moreover, this method will simplify further studies of metabolism and pharmacodynamics and will encourage the use of HSS T3 columns in similar traditional Chinese medicine research.