Two New Bromophenols with Radical Scavenging Activity from Marine Red Alga Symphyocladia latiuscula

Chemical investigation of a Chinese collection of marine red alga Symphyocladia latiuscula yielded two new highly brominated phenols. The structures of the new compounds were elucidated by detailed spectroscopic analysis, including HRMS, 1D and 2D NMR and MS methods. Compounds 1 and 2 were evaluated for radical scavenging capability by 1,1-diphenyl-2-picrylhydrazuyl (DPPH) radical with the IC50 value of 14.5 and 20.5 μg/mL, respectively.


Introduction
Symphyocladia latiuscula (Harvey) Yamada is a marine red alga distributed along the coasts of Northern China, Korea, and Japan [1]. This red alga is a rich source of bromophenols with high chemical diversity and various biological activities. Previous chemical studies on this species have resulted in the characterization of 25 monoaryl and diaryl bromophenols with a variety of bioactivities,

General Experimental Procedures
NMR spectra were recorded on a Varian Inova 600 MHz spectrometer at 600 MHz for 1 H and 125 MHz for 13 C in DMSO-d 6 and Methanol-d 4 using solvent signals (DMSO: δ H 2.50/δ C 39.51; Methanol: δ H 3.31/δ C 49.15) as reference; the coupling constants were in Hz. ESIMS spectra were recorded with a ABI Mariner ESI-TOF. Column chromatography was performed with silica gel (200-300 mesh, Qingdao Haiyang Chemical Factory) and Sephadex LH-20 (Pharmacia Co., Sweden) columns. HPLC was performed using an Agilent 1100 Series separations module equipped with Agilent 1100 Series diode array detector and performed using an Agilent Zorbax Eclipse XDB-C8 (5 μm) semipreparative column (9.4 × 250 mm).

Extraction and Isolation
The air-dried red alga Symphyocladia latiuscula (4.3 kg) was extracted with 95% EtOH at room temperature (3 × 72 h). After the solvent was removed under reduced pressure at <40 °C, a dark residue (610 g) was obtained. The residue was partitioned between EtOAc and H 2 O and the EtOAc-soluble partition (320 g) was chromatographed over silica gel, eluting with a gradient of 0%-100% Me 2 CO/petroleum ether [4]. The fraction eluted by 30% Me 2 CO/petroleum ether was further fractionated over Sephadex LH-20 using petroleum ether-CHCl 3 -MeOH (5:5:1) to afford 18 fractions. The ninth fraction from the LH-20 column was further fractionated by ODS column, which was eluted with a stepwise gradient of 0%-100% MeOH/H 2 O to afford 11 subfractions. The forth ODS subfraction was subjected to HPLC fractionation (Zorbax Eclipse XDB-C8 5 μm 250 × 9.4 mm column) to yield compound 1. The 13th fraction from the LH-20 column was further fractionated by ODS column, which was eluted with a stepwise gradient of 0%-100% MeOH/H 2 O to afford 11 subfractions. The third ODS subfraction was subjected to HPLC fractionation (Zorbax Eclipse XDB-C8 5 μm 250 × 9.4 mm column) to yield compound 2.

Scavenging Ability on 1,1-Diphenyl-2-picrylhydrazuyl (DPPH) Radical
Each power (0.1-20 mg/mL, 4.0 mL) in deionized water was mixed with 1.0 mL of methanolic solution containing DPPH (Sigma) radicals, resulting in a final concentration of 0.2 mM DPPH. The mixture was shaken vigorously and left to stand for 30 min in the dark, and the absorbance was then measured at 517 nm against a blank [14]. The scavenging ability was calculated as follows: scavenging ability (%) = [(A517 of control − A517 of sample)/A517 of control] × 100 (1) Vitamin C was used for positive control.

Conclusions
Symphyocladia latiuscula is a rich source of bromophenols with specific subunit of 2,3,6-tribromo-4, 5-dihydroxybenzene. During the course of our systematic search for new biologically active bromophenols from this marine red alga, two new bromophenols (1 and 2) with radical scavenging activity were characterized by mass spectrum guided fractionation. Compounds 1 and 2 exhibited radical scavenging capability on 1,1-diphenyl-2-picrylhydrazuyl (DPPH) radical with the IC 50 value of 14.5 and 20.5 μg/mL, respectively.