496 Spectral Assignments and Reference Data Received: 25 September 2009 Revised: 28 December 2009 Accepted: 4 March 2010 Published online in Wiley Interscience: 13 April 2010 (www.interscience.com) DOI 10.1002/mrc.2605 Structural elucidation and NMR assignments of four aromatic lactones from a mangrove endophytic fungus (No. GX4-1B) Hongbo Huang, a Qing Li, a Xiaojun Feng, a Bin Chen, a Jun Wang, b,c Lan Liu, a Zhigang She a,c * and Yongcheng Lin a,c * Two new aromatic lactones, 6-hydroxy-4-hydroxymethyl-8-methoxy-3- methylisocoumarin (1) and 1,10-dihydroxy-8-methyl- dibenz[b, e]oxepin-6,11-dione (2), together with two known compounds, 1,10-dihydroxy-dibenz[b, e]oxepin-6,11-dione (3) and 3-hydroxymethyl-6,8-dimethoxycoumarin (4), were isolated from a mangrove endophytic fungus (No. GX4-1B) collected from the South China Sea. Their structures were elucidated and the data of 1 H and 13 C NMR were assigned completely by HREIMS, 1D and 2D NMR experiments including HMQC, HMBC and NOESY. Copyright c  2010 John Wiley & Sons, Ltd. Keywords: NMR; 1 H NMR; 13 C NMR; 2D NMR; aromatic lactone; isocoumarin; coumarin; dibenzoxepin derivative; marine fungus Introduction Marine-derived fungi provide unique and bioactive secondary metabolites and have been the focus of natural products research. [1] Our research group has isolated several series of novel bioactive compounds from marine-derived mangrove endophytic fungi from the South China Sea in the past 10 years. [2–7] During the course of our continuing investigation of bioactive constituents from marine fungus, four aromatic lactones including one new isocoumarin (1), one new dibenzoxepin derivative (2) together with two known compounds (3) and (4) (Fig. 1) were obtained from the culture broth extract of a mangrove endophytic fungus (No. GX4-1B) collected from Bruguiera gymnoihiza (L.) Savigny., grown in the coastal salt marsh of Guangxi province, China. High-resolution EIMS (HREIMS), 1D- and 2D NMR experiments were used to determine their structures. The known compound (3) was isolated from nature first. Compounds (2) and (3) are dibenz[b,e]oxepin-6,11-dione derivative. Only several dibenz[b,e]oxepin-6,11-dione compounds obtained from nature were reported in the literature, [8 – 10] which suggested that dibenz[b,e]oxepin-6,11-dione derivative was rare as natural products. Compound (4) has been reported in the previous articles. [11] However, the 13 C NMR data of (4) was partial and the presumed assignment was mistaken in the literature. Therefore, we report the full assignment of the 1 H and 13 C NMR of compound (4) for reference purpose. Results and Discussion Compound (1) was obtained as colorless needle crystal. A molecular ion peak in HREIMS at m/z 236.0680 (calculated for 236.0679) established a molecular formula of C 12 H 12 O 5 . Then seven degrees of unsaturation were determined. The 13 H and 13 C NMR spectrums (Table 1) revealed compound (1) had nine sp2 carbons including one carboxyl group (δ C = 164.1), which suggested that this compound had a coumarin or isocoumarin skeleton. Futhermore, one methyl (δ C = 16.7, δ H = 2.23), one methoxyl (δ C = 55.7, δ H = 3.81), one hydroxymethyl (δ C = 56.2, δ H = 4.39) and a pair of meta-coupled aromatic hydrogens (δ H = 6.69, d, J = 2.0 Hz; δ H = 6.46, d, J = 2.0 Hz) existed. From the HMBC correlations of H-1 ′ to C-3, C-4, C-4a, 3-Me to C-3, C-4, C-4a, in conjunction with the interactions between H-1 ′ and H-5, H-1 ′ and 3-Me in NOESY spectrum (Fig. 2), we could deduce a 3-methyl-4-methoxyl-6,8-bisubstituted isocoumarin. The methoxyl was connected to C-8 by HMBC correlation from 8-OMe to C-8, and this connection was confirmed by the NOESY correla- tion between 7-H and 8-OMe. So compound (1) was identified as 6-hydroxy-4-hydroxymethyl-8-methoxy-3- methylisocoumarin. Compound (2) was isolated as yellow needle crystal. The molecular formula C 15 H 10 O 5 was determined by HREIMS at m/z 270.0524 (calculated 270.0523) along with the analysis of NMR data, which indicated 11 ◦ of unsaturation. The 13 C NMR spectrum (Table 2) gives rise to 15 carbon signals: one carbonyl carbon of ketone (δ C = 180.4), one ester carbonyl carbon (δ C = 169.3), seven aromatic quarternary carbons, five aromatic methines and one methyl carbon (δ C = 21.2). The 1 H NMR spectrum displayed the signals of two exchangeable protons (δ H = 12.30, brs; δ H = 13.21, brs) from two hydroxyl chelated with carboxyl, three coupled aromatic hydrogens (δ H = 7.74, t, J = 8.4 Hz; δ H = 7.08, dd, J = 8.4, 0.8 Hz; δ H = 6.84, dd, J = 8.4, 0.8 Hz), showed the ∗ Correspondence to: Zhigang She and Yongcheng Lin, School of Chemistry and Chemical Engineering, Sun Yat-Sen (Zhongshan) University, Guangzhou, PR China. E-mail: cesshzhg@mail.sysu.edu.cn; ceslyc@mail.sysu.edu.cn a School of Chemistry and Chemical Engineering, Sun Yat-Sen (Zhongshan) University, Guangzhou, 510275, PR China b School of Pharmaceutical Sciences, Sun Yat-Sen (Zhongshan) University, Guangzhou 510006, PR China c Key Laboratory of Functional Molecules from Marine Microorganisms, Sun Yat- Sen University, Education Department of Guangdong Province, Guangzhou 510080, PR China Magn. Reson. Chem. 2010, 48, 496–499 Copyright c  2010 John Wiley & Sons, Ltd.