2.1. Hericium erinaceus Extracts and Analysis of Erinacine A

Fresh dried mycelium of H. erinaceus (2 kg) was extracted using 95% ethanol. The extracted ethanol solution was concentrated and fractionated by solvent partition between ethyl acetate (EtOAc) and water to afford an H₂O layer and EtOAc layer. The EtOAc layer analysis was subjected to silica gel column chromatography according to previous studies [16,24], while the HPLC analysis of erinacine A was performed with minor modifications. The analytical column was a COSMOSIL 5C18-AR-II (250 × 4.6 mm; particle size 5 μm, Nacalai USA, Inc., Kyoto, Japan). The 5 mg/kg erinacine A in the H. erinaceus extracted with 85% ethanol was confirmed and quantified by HPLC. The chemical compounds suggested in this article, erinacine A (PubChem CID: 10410568), the HPLC chromatogram (as supporting material), and the calibration curve used are shown in Figure 1 [27].

HPLC analysis and LC-MS analysis of the ethanol Hericium erinaceus mycelium (HEM) extract. The retention time peak at 7.493 min was erinacine A (UV detection at 340 nm).