4-Methylthio-butanyl derivatives from the seeds of Raphanus sativus and their biological evaluation on anti-inflammatory and antitumor activities.
BACKGROUND
Raphanus sativus seeds (Brassicaceae) known as Raphani Semen have long been used as anti-cancer and/or anti-inflammatory agents in Korean traditional medicine. This study was designed to isolate the bioactive constituents from the seed extracts of Raphanus sativus and evaluate their anti-inflammatory and antitumor activities.


METHODS
Bioassay-guided fractionation and chemical investigation of a methanolic extract of the seeds of Raphanus sativus led to the isolation and identification of seven 4-methylthio-butanyl derivatives. Structural elucidation of the isolated compounds was carried out using 1D and 2D nuclear magnetic resonance (NMR) spectroscopy techniques ((1)H, (13)C, COSY, HMQC and HMBC experiments) and mass spectrometry.


RESULTS
The isolated compounds were characterized as in the following: three new 4-methylthio-butanyl derivatives, sinapoyl desulfoglucoraphenin (1), (E)-5-(methylsulfinyl)pent-4-enoxylimidic acid methyl ester (2), and (S)-5-((methylsulfinyl)methyl)pyrrolidine-2-thione (3), together with four known compounds, 5-(methylsulfinyl)-4-pentenenitrile (4), 5-(methylsulfinyl)-pentanenitrile (5), sulforaphene (6), and sulforaphane (7). Full NMR data assignments of the three known compounds 4-6 were also reported for the first time. We evaluated the anti-neuroinflammatory effect of 1-7 in lipopolysaccharide-stimulated murine microglia BV2 cells. Compound 1 significantly inhibited nitrite oxide production with IC50 values of 45.36 μM. Moreover, it also reduced the protein expression of inducible nitric oxide synthase. All isolates were also evaluated for their antiproliferative activities against four human tumor cell lines (A549, SK-OV-3, SK-MEL-2, and HCT-15), and all of them showed antiproliferative activity against the HCT-15 cell, with IC50 values of 8.49-23.97 μM.


CONCLUSIONS
4-Methylthio-butanyl derivatives were one of the main compositions of Raphanus sativus seeds, and activities demonstrated by the isolated compounds support the ethnopharmacological use of Raphanus sativus seeds (Brassicaceae) as anti-cancer and/or anti-inflammatory agents.
Natural Products Laboratory, School of Pharmacy, Sungkyunkwan University, 300 Chonchon-dong, Jangan-ku, Suwon, Gyeonggi-do 440-746, Republic of Korea.
Ki Hyun
KH
Kim
College of Pharmacy, Gachon University, #191 Hambakmoero, Yeonsu-gu, Incheon 406-799, Republic of Korea.
Eunjung
E
Moon
College of Pharmacy, Gachon University, #191 Hambakmoero, Yeonsu-gu, Incheon 406-799, Republic of Korea; Gachon Medical Research Institute, Gil Medical Center, Inchon 405-760, Republic of Korea.
Sun Yeou
SY
Kim
Korea Research Institute of Chemical Technology, Teajeon 305-600, Republic of Korea.
Sang Un
SU
Choi
College of Oriental Medicine, Dongguk University, Gyeong Ju 780-714, Republic of Korea.
Jei Hyun
JH
Lee
Natural Products Laboratory, School of Pharmacy, Sungkyunkwan University, 300 Chonchon-dong, Jangan-ku, Suwon, Gyeonggi-do 440-746, Republic of Korea. Electronic address: krlee@skku.ac.kr.
Kang Ro
KR
Lee
D000818:Animals; D000893:Anti-Inflammatory Agents / Q000737:chemistry / Q000494:pharmacology*; D000972:Antineoplastic Agents, Phytogenic / Q000737:chemistry / Q000494:pharmacology*; D002073:Butanes / Q000737:chemistry*; D002460:Cell Line; D002470:Cell Survival; D006801:Humans; D015394:Molecular Structure; D031224:Raphanus / Q000737:chemistry*; D012639:Seeds / Q000737:chemistry*; D013438:Sulfhydryl Compounds / Q000737:chemistry*
D000893:Anti-Inflammatory Agents; D000972:Antineoplastic Agents, Phytogenic; D002073:Butanes; D013438:Sulfhydryl Compounds
3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide; 4-Methylthio-butanyl derivatives; Anti-inflammation; Brassicaceae; CC; COSY; Cytotoxicity; DMEM; Doxorubicin (PubChem CID: 31703); Dulbecco's modified Eagle medium; ECL; ESI; FBS; HMBC; HMQC; HPLC; HR; HRP; IR; L-NMMA; LPLC; LPS; MS; MTT; N(G)-monomethyl-L-arginine; N(G)-monomethyl-L-arginine (PubChem CID: 135242); NCI; NMR; NO; NOS; Nitric oxide; RP; Raphanus sativus; SDS-PAGE; SRB; Sulforaphane (PubChem CID: 5350); Sulforaphene (PubChem CID: 6433206); TLC; UV; column chromatography; correlation spectroscopy; electrospray ionization; enhanced chemiluminescence; fetal bovine serum; heteronuclear multiple bond correlation; heteronuclear multiple quantum coherence; high resolution; horseradish peroxidase; iNOS; inducible nitric oxide synthase; infrared; lipopolysaccharide; low-pressure liquid chromatography; mass spectrometry; national cancer institute; nitric oxide; nitric oxide synthase; nuclear magnetic resonance; preparative high performance liquid chromatography; reversed-phase; sodium dodecyl sulfate polyacrylamide gel electrophoresis; sulforhodamine B; thin-layer chromatography; ultraviolet