New inhibitors of nitric oxide production from the seeds of Myristica fragrans

Abstract

Six dihydrobenzofuran type neolignans were isolated from the dried ripe seeds of Myristica fragrans Houtt. (family: Myristicaceae) and their chemical structures were identified as licarin B (1), 3'-methoxylicarin B (2), myrisfrageal A (3), isodihydrocainatidin (4), dehydrodiisoeugenol (5), and myrisfrageal B (6), respectively, on the basis of spectroscopic data analyses. Among them, compounds 3 and 6 are new compounds. Compounds 1-6 showed inhibition of nitric oxide production in lipopolysaccharide-activated murine monocyte-macrophage RAW264.7 with IC50 values of 53.6, 48.7, 76.0, 36.0, 33.6, and 45.0 μM, respectively. These values were compared to those of the positive controls, indomethacin and L-N(6)-(1-iminoethyl)-lysine, which have IC50 values of 65.3 and 27.1 μM, respectively. Further compounds 3, 5 and 6 suppressed LPS-induced iNOS mRNA expression in a does-dependent manner in RAW 264.7 cells assayed by real-time RT-PCR. Compounds 3, 5 and 6 may inhibit NO overproduction via inhibition of iNOS mRNA expression. The results provided valuable information for further investigation of compounds 1-6 as anti-inflammatory and chemopreventive agents.

Keywords: (1)H–(1)H COSY; (1)H–(1)H correlation spectroscopy; 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide; CC; CD; CDCl(3); CHCl(3); DMEM; DMSO; Dulbecco’s modified Eagle’s medium; EI–MS; FBS; HMBC; HR-ESI-MS; HSQC; IC(50); IND; IR; Inducible nitric oxide synthase; L-N(6)-(1-iminoethyl)-lysine; L-NIL; LPS; MTT; MeOH; Myrisfrageal A; Myrisfrageal B; Myristica fragrans; NMR; NO; NOESY; Nitric oxide; Nutmeg; RP-SP-HPLC; RT-PCR; TMS; UV; chloroform; circular dichroism; column chromatography; deuterated chloroform; dimethyl sulfoxide; electron ionization mass spectrometry; fetal bovine serum; half maximal inhibitory concentration; heteronuclear multiple-bond correlation; heteronuclear single quantum coherence; high-resolution electron spray ionization mass spectrometry; iNOS; indomethacin; inducible nitric oxide synthase; infrared; lipopolysaccharide; methanol; nitric oxide; nuclear Overhauser effect spectroscopy; nuclear magnetic resonance; reverse transcription-polymerase chain reaction; reversed phase semi-preparative high-performance liquid chromatography; tetramethylsilane; ultraviolet.