A New Hydroxychavicol Dimer from the Roots of Piper betle

A new hydroxychavicol dimer, 2-(γ'-hydroxychavicol)-hydroxychavicol (1), was isolated from the roots of Piper betle Linn. along with five known compounds, hydroxychavicol (2), aristololactam A II (3), aristololactam B II (4), piperolactam A (5) and cepharadione A (6). The structures of these isolated compounds were elucidated by spectroscopic methods. Compounds 1 and 2 exhibited inhibitory effects on the generation of superoxide anion and the release of elastase by human neutrophils.

Neutrophils play a pivotal role in the defense of the human body against infections.However, activated human neutrophils are known to cause tissue damage and to play a critical role in a variety of acute and chronic inflammatory diseases [14,15].For example, high concentrations of reactive oxygen species and elastase produced by activated neutrophils in the sputum of patients with airway mucus hypersecretion has been implicated in the pathogenesis of many pulmonary diseases including asthma, chronic obstructive pulmonary disease, cystic fibrosis and acute respiratory distress syndrome [16][17][18][19].In a search for suitable new anti-neutrophilic inflammatory agents from natural sources, the inhibition of O 2 •− production and elastase release in human neutrophil by compounds 1-6 were assayed.This paper describes the isolation, the determination of the structure of the new compound and the anti-inflammatory activity of the isolated compounds.
In the HMBC spectrum of 1 (Table 1 and Figure 2), the methylene proton signal at  H 3.34 (H-') showed correlations with carbon signals at  C 117.33 (C-3) and 129.79 (C-1), which also correlated to the olefinic methane proton signal at  H 5.92 (H-β) clearly suggested that the allyl group and C-' were connected to C-2 and C-1 of the tetrasubstrate benzene ring, respectively.Forthemore, the olefinic methane proton signal at  H 6.09 (H-β') displayed correlations with two aromatic quaternary carbon signals at  C 130.46 (C-2) and 130.98 (C-1'), and the signals at  H 6.24 (H-α') correlated with the signals of C-2' and C-6', indicated that C-α' was located at C-1'.The coupling constant (J α'-β' = 15.6 Hz) indicated a trans configuration between H-α and H-β.From the above data, the structure of 1 was identified as 2-('-isohydroxychavicol)hydroxychavicol.The in vitro anti-inflammatory effects of compounds 1-6 were tested (Table 2).Compound 2 (hydroxychavicol monomer) showed significant inhibitory effects in superoxide anion generation and elastase release (IC 50 0.27 and 5.78 μM; Table 2 and Figure 3).Compound 1 (a dimer of hydroxychavicol) also showed moderate effects in both assays (IC 50 8.59 and 13.14 μM; Table 2 and Figure 4).These findings suggest that compounds 1 and 2 merit further investigation as potential anti-inflammatory compounds.(A) (B)

General
Melting points were determined using a Yanaco MP-I3 micro melting point apparatus and the thermometer was used without correction.Mass spectra were recorded using a Finnigan MAT GCQ spectrometer (EIMS). 1 H, 13 C, and 2D-NMR spectra were measured with a Varian VNMRS 600 MHz spectrometer.

Plant Material
The roots of P. betle Linn.were collected from Taitung County, Taiwan, in April 2011, and was identified by a taxonomist, Mr. Jun-Chih Ou.A voucher specimen (No.20110401) was deposited in the Department of Plant Industry, National Pingtung University of Science and Technology.

Table 2 .
Effects of compounds on superoxide anion generation and elastase release by human neutrophils in response to FMLP/CB.