Supplementary Information

Isohericenone, a newcytotoxic isoindolinone alkaloid from Hericium erinaceum

Ki Hyun Kim1,†, Hyung Jun Noh2,†, Sang UnChoi3, and Kang Ro Lee1

1Natural Products Laboratory, School of Pharmacy, SungkyunkwanUniversity, Suwon 440-746, Korea

2Department of Herbal Crop Research, National Institutes of Horticultural & Herbal Science, RDA, Eumseoung 440-746, Korea

3Bio-organic Science Division, Pharmacology Research Center, Korea Research Institute of Chemical Technology, Teajeon 305-600, Korea

†These authors contributed equally to this work

General

IR spectra were recorded on a Bruker IFS-66/S FT-IR spectrometer (Bruker, Karlsruhe, Germany). Electrospray ionization (ESI) and high-resolution (HR)-ESI mass spectra were recorded on a SI-2/LCQ DecaXP Liquid chromatography (LC)-mass spectrometer (Thermo Scientific, West Palm Beach, PL, USA). Nuclear magnetic resonance (NMR) spectra, including 1H-1H COSY, HMQC, and HMBCexperiments, were recorded on a Varian UNITY INOVA 500 NMR spectrometer (Varian, Palo Alto, CA, USA) operating at 500 MHz (1H) and 125 MHz (13C), with chemical shifts given in ppm (δ). Preparative high performance liquid chromatography (HPLC) used a Gilson 306 pump (Gilson, Middleton, WI, USA) with a Shodex refractive index detector (Shodex, New York, NY, USA). Column chromatography was performed with a silica gel 60 (Merck, Darmstadt, Germany, 70-230 mesh and 230-400 mesh). Merck precoated Silica gel F254 plates and RP-18 F254s plates (Merck, Darmstadt, Germany) were used for TLC. Spots were detected on TLC under UV light or by heating after spraying with anisaldehyde-sulfuric acid.

Mushroom Material

The fresh fruiting bodies of Hericium erinaceumwere purchased from Pochun mushroom development Co.,Ltd at Pochun-gun in Gyunggi-Do province, Korea, in July 2010.A voucher specimen (SKKU-2010-07) of the mushroom was authenticated by one of the authors (K.R.L.)and was deposited in the herbarium of the School of Pharmacy, Sungkyunkwan University, Korea.

Physico-chemical properties

Isohericenone (1).colorless gum,IR (KBr) νmax3357, 2944, 2833, 1701, 1661, 1593, 1471, 1113, 1032 cm-1. 1H- and 13C-NMR spectra: see Table 1. ESI-MS m/z456 [M+Na]+. HR-ESI-MS (positive-ion mode) m/z456.2155 ([M + Na]+, C27H31O4NNa, calcd. for 456.2151).

Test for cytotoxicity in vitro.

A sulforhodamin B bioassay (SRB) was used to determine the cytotoxicity of each compound against four cultured human cancer cell lines. The assays were performed at the Korea Research Institute of Chemical Technology. The cell lines used were A549 (non small cell lungcarcinoma), SK-OV-3 (ovary malignant ascites), SK-MEL-2 (skin melanoma), and HCT-15 (colon adenocarcinoma). Doxorubicin was used as a positive control. The cytotoxicities of doxorubicin against A549, SK-OV-3, SK-MEL-2, and HCT-15 cell lines were at IC50values of 0.001, 0.003, 0.002, and 0.081μM, respectively.

S1.The 1H NMR spectrum of 1(CD3OD, 500 MHz)

S2. The 13C NMR spectrum of 1 (CD3OD, 125 MHz)

S3. The 1H-1HCOSY spectrum of 1 (CDCl3)

S4. The HMQC spectrum of 1 (CDCl3)

S5. The HMBC spectrum of 1 (CDCl3)

S6. The NOESY spectrum of 1 (CDCl3)

S7.The 1H NMR spectrum of 2 (CDCl3, 500 MHz)

S8. The 13C NMR spectrum of 2 (CDCl3, 125 MHz)

S9. The 1H-1HCOSY spectrum of 2 (CDCl3)

S10. The HMQC spectrum of 2 (CDCl3)

S11. The HMBC spectrum of 2 (CDCl3)