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1 Supporting information Aspterpenacids A and B, Two Sesterterpenoid from a Mangrove Endophytic Fungus Aspergillus terreus H010 Zhaoming Liu a, Yan Chen a, Senhua Chen a, Yayue Liu a, Yongjun Lu b,c, Dongni Chen b, Yongcheng, Lin a, Xishan Huang a,* a, b,* and Zhigang She a College of Chemistry and Chemical Engineering, Sun Yat-sen University, Guangdong , China. b School of Life Sciences and Biomedical Center, Sun Yat-Sen University, Guangzhou , P. R. China. c Key Laboratory of Functional Molecules from Oceanic Microorganism (Sun Yat-sen Uni -versity), Department of Education of Guangdong Province, Guangzhou , China. Table of contents: Experimental details. General experimental procedures. Fungal material. Fermentation, extraction and isolation. Biological assays. X-ray crystallographic data for asterpenacide A (1) Computational details. Methods. Result. Table S1. Energy Analysis for the Conformers of 1 and 2. Figure S1. B3LYP/6-31G(d) optimized low-energy conformers of 1 and 2. Figure S2. Comparison of the experimental ECD spectra of 1 with the B3lYP/6-311+g(2d,p) calculated spectrum of 1 and ent-1 in MeOH. σ = 0.30 ev. Figure S3. Comparison of the experimental ECD spectra of 2 with the B3lYP/6-311+g(2d,p) calculated spectrum of 2 and ent-2 in MeOH. σ = 0.30 ev. References. Figure S4. 1 H NMR spectrum of compound 1 in CDCl 3 (600 MHz). Figure S5. 13 C NMR spectrum of compound 1 in CDCl 3 (150 MHz). 1

2 Figure S6. DEPT-90 spectrum of compound 1 in CDCl 3 (150 MHz). Figure S7. DEPT-135 spectrum of compound 1 in CDCl 3 (150 MHz). Figure S8. 1 H- 1 H COSY spectrum of compound 1 in CDCl 3 (600 MHz). Figure S9. HSQC spectrum of compound 1 in CDCl 3 (600 MHz). Figure S10. HMBC spectrum of compound 1 in CDCl 3 (600 MHz). Figure S11. NOESY spectrum of compound 1 in CDCl 3 (600 MHz). Figure S12. Expanded NOESY spectrum of compound 1 in CDCl 3 (600 MHz). Figure S13. 1 H NMR spectrum of 1 in DMSO (600 MHz). Figure S C NMR spectrum of 1 in DMSO (600 MHz). Figure S15. DEPT-90 spectrum of compound 1 in DMSO (150 MHz). Figure S16. DEPT-135 spectrum of compound 1 in DMSO (150 MHz). Figure S17. HSQC spectrum of 1 in DMSO (600 MHz). Figure S18. 1 H- 1 H COSY spectrum of 1 in DMSO (600 MHz). Figure S19. NOESY spectrum of 1 in DMSO (600 MHz). Figure S10. HMBC spectrum of 1 in DMSO (600 MHz). Figure S21. 1 H NMR spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). Figure S C NMR spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (150 MHz). Figure S23. DEPT-90 spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (150 MHz). Figure S24. DEPT-135 spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (150 MHz). Figure S25. 1 H- 1 H COSY spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). Figure S26. HSQC spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). Figure S27. HMBC spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). Figure S28. NOESY spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). Figure S28. Expanded NOESY spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). Figure S29. 1 H NMR spectrum of 2 in DMSO (600 MHz). Figure S C NMR spectrum of 2 in DMSO (600 MHz). Figure S31. DEPT-90 spectrum of 2 in DMSO (600 MHz). Figure S32. DEPT-135 spectrum of 2 in DMSO (600 MHz). Figure S33. HSQC spectrum of 2 in DMSO (600 MHz). Figure S34. 1 H- 1 H COSY spectrum of 2 in DMSO (600 MHz). Figure S35. HMBC spectrum of 2 in DMSO (600 MHz). Figure S36. NOESY spectrum of 2 in DMSO (600 MHz). Figure S37. HRESI TOF MS spectrum of compound 1. Figure S38. HRESI TOF MS spectrum of compound 2. 2

3 Experimental details General experimental procedures. Melting points were determined on a Fisher-Johns hot-stage apparatus and were uncorrected. UV data were measured on a UV-240 spectrophotometer (Shimadzu, Beijing, China). HRMS (ESI) were determined with a Q-TOF high-resolution mass spectrometer (Waters). IR spectrum was recorded using Bruker Vector spectrophotometer 22. Optical rotation was recorded using a an MCP300 (Anton Paar, Shanghai, China). CD data were recorded with a J-810 spectropolarimeter (JASCO, Tokyo, Japan). The NMR data were recorded on a Bruker Avance 600 spectrometer (Bruker, Beijing, China) at 600 MHz for 1 H and 125 MHz for 13 C, respectively. All chemical shifts (δ) are given in ppm with reference to TMS, and coupling constants (J) are given in Hz. Column chromatography (CC) was carried out on silica gel ( mesh, Marine Chemical Factory, Qingdao, China) and sephadex LH-20 (Amersham Pharmacia, Piscataway, NJ, USA). Solvents were distilled prior to use. Semipreparative HPLC was performed on a Waters Breeze HPLC system using a Phenomenex Luna (Phenomenex, Torrance, CA, USA) C 18 column ( mm, 5 μm), flow rate, 2.0 ml/min. Fungal material. The fungus used in this study was isolated from healthy leaf of the marine mangrove Kandelia obovata, which were collected in April 2012 from Zhanjiang Mangrove Nature Reserve in Guangdong Province, China. It was obtained using the standard protocol for the isolation. Fungal identification was carried out using a molecular biological protocol by DNA amplification and sequencing of the ITS region. The sequence data obtained from the fungal strain have been deposited at GenBank with accession no. KU A BLAST search result showed that the sequence was the most similar (100%) to the sequence of aspergillus terreus (compared to KT KP ). A voucher strain was deposited in School of Chemistry and Chemical Engineering, Sun Yat-Sen University, Guangzhou, China. 3

4 Fermentation, extraction and isolation. The fungus was grown on liquid cultured medium (composed of maltose (20.0 g/l), mannitol (20.0 g/l), glucose (10.0 g/l), monosodium glutamate (10.0 g/l), MgSO4 7H2O (0.3 g/l), KH2PO4 (0.5 g/l), yeast extract (3.0 g/l), corn steep liquor (1.0 g/l)) in 80 Erlenmeyer flasks for 45 days at room temperature under static condition. After fermentation, The former was extracted with EtOAc and concentrated under reduced pressure to yield residual gum in 4.7 g. The residue was subjected to silica gel CC using gradient elution with petroleum ether-etoac from 90:10 to 0:100 (v/v) to give twelve fractions (Frs.1 12). Fr. 4 (116 mg) was further purified by silica gel CC using CHCl3/MeOH (97:3) to afford six subfractions (Frs ). Fr. 5.5 (20.1 mg) was applied to Sephedx LH-20 CC, eluted with MeOH to obtain 1 (4.3 mg). After crystallization in MeOH and purification by HPLC, Fr. 5.2 (30.2 mg) afforded 2 (5.7 mg). Aspterpenacid A (1): Colorless crystal; m.p. 218 ~ 220 C. [α] (c = 0.01, MeOH). UV (MeOH) λ max : 275 nm. IR (KBr): 3481, 2953, 2673, 2597, 1716, 1679, 1265, 1021 cm -1. HRESI TOF MS m/z [M H] (calcd for C 27 H 42 O ). 1 H and 13 C NMR data: see table 1. D Aspterpenacid B (2): Colorless crystal; m.p. 236 ~ 238 C. [α] (c = 0.01, MeOH). UV (MeOH) λ max : 275 nm. IR (KBr): 3294, 2953, 2870, 2678, 2599, 1682, 1458, 1311, 1021 cm -1. HRESI TOF MS m/z [M H] (calcd for C 25 H 40 O ). 1 H and 13 C NMR data: see table 1. D Biological assays. Antibacterial activity were assayed according to established procedure (Appendino, G.; et. al. J. Nat. Prod. 2008, 71, ). Three Gram-positive bacterial (S. aureus, S. epidermidis and B. subtilis) and three Gram-negative bacterial (E. coli, K. peneumoniae and A. calcoaceticus) were used. 4

5 Cytotoxic activity were determined using the MTT method according to established procedures (Scudiero, D. A. et. al. Cancer Research 1988, 48, 4827.). Two human cancer cell lines (hela and MCF-7) were used in cytotoxicity bioassay. X-ray crystallographic data for aspterpenacid A (1) Crystal data and structure refinement for 1 are as follows: Empirical formula 2(C 27 H 42 O 5 ), H 2 O 1 Formula weight Temperature Wavelengh Crystal system 293(2) K Å Monoclinic Space group C Unit cell dimensions a= (5) Å α= 90 b= (10) Å β= (4) c= (12) Å γ= 90 Volume (4) Å 3 Z 4 Density (calculated) Mg/m 3 Absorption coefficient mm 1 F(000) 1968 Crystal size mm 3 Theta range for data collection to Index ranges 13 <= h <= 13, 21 <= k <= 21, 30 <= l <= 31 Reflections collected Independent reflections 9570 [R(int)= 0.03] 5

6 Completeness to theta = % Absorption correction Semi-empirical from equivalents Max. and min. Transmission to Refinement method Full-matrix least-squares on F 2 Data / restraints / parameters 9670 /6 /581 Goodness-of-fit on F Final R indices [I>2sigma(I)] R1= , ωr2= R indices (all data) R1= , ωr2= Absolute structure parameter 0.1(3) Data were collected on Agilent Xcalibur Nova single-crystal diffractometer using Cu Kα radiation. The crystal structure was refined by full-matrix least-squares calculation. Crystallographic data for the structure of 1 have been deposited in the Cambridge Crystallographic Data Centre (deposition number: CCDC ). Copies of these data can be obtained free of charge via from the Cambridge Crystallographic Data Centre, 12, Union Road, Cambridge CB21EZ, UK.; fax: (t44) ; or deposit@ccd.cam.ac.uk). Computational details Methods. Molecular Merck force field (MMFF) and DFT/TD-DFT calculations were carried out with Spartan 14 software (Wavefunction Inc., Irvine, CA, USA) and Gaussian 09 program, respectively. 1 Conformers within 10 kcal/mol energy window were generated and optimized using DFT calculations at B3LYP/6-31G(d) level. Frequency calculations were performed at the same level to confirm that each optimized conformer was true minimum and to estimate their relative thermal free energy ( G) at K. Conformers with Bolzmann distribution over 1% were chosen for ECD calculations in methanol at B3lYP/6-311+g(2d,p) level. Solvent 6

effects were taken into consideration using the Self-consistent Reaction Field method (SCRF) with Polarizable continuum model (PCM).

Table S1. Energy Analysis for the Conformers of 1 and 2. compounds Conformation G (Hartree) G (Kcal/mol) G (Kcal/mol) Boltzmann Dist (%) 1 1-a 1430.27316748 897500.5604 0.3246 30.

46 ent-1 ent-1-a 1430.26943310 897498.2171 0.1334 38.59 ent-1-b 1430.26841095 897497.5756 0.7748 13.06 ent-1-c 1430.26964572 897498.3505 0.0000 48.34 ent-1-d 1430.26119481 897493.

7 effects were taken into consideration using the Self-consistent Reaction Field method (SCRF) with Polarizable continuum model (PCM). 2 The ECD spectrum were generated by program SpecDis 3 using a Gaussian band shape with 0.30 ev exponential half-width from dipole-length dipolar and rotational strengths. Result. Table S1. Energy Analysis for the Conformers of 1 and 2. compounds Conformation G (Hartree) G (Kcal/mol) G (Kcal/mol) Boltzmann Dist (%) 1 1-a b c d e ent-1 ent-1-a ent-1-b ent-1-c ent-1-d a b ent-2 ent-2-a ent-2-b ent-2-c a 1-b 1-c 7

8 1-d 1-e ent-1-a ent-1-b ent-1-c ent-1-d 2-a 2-b ent-2-a ent-2-b ent-2-c Figure S1. B3LYP/6-31G(d) optimized low-energy conformers of 1 and 2. 8

9 Figure S2. Comparison of the experimental ECD spectra of 1 with the B3lYP/6-311+g(2d,p) calculated spectrum of 1 and ent-1 in MeOH. σ = 0.30 ev. Figure S3. Comparison of the experimental ECD spectra of 2 with the B3lYP/6-311+g(2d,p) calculated spectrum of 2 and ent-2 in MeOH. σ = 0.30 ev. References. (1) Frisch, M. J.; Trucks, G. W.; Schlegel, H. B.; Scuseria, G. E.; Robb, M. A.; Cheeseman, J. R.; Scalmani, G.; Barone, V.; Mennucci, B.; Petersson, G. A.; Nakatsuji, H.; Caricato, M.; Li, X.; Hratchian, H. P.; Izmaylov, A. F.; Bloino, J.; Zheng, G.; Sonnenberg, J. L.; Hada, M.; Ehara, M.; Toyota, K.; Fukuda, R.; Hasegawa, J.; Ishida, M.; Nakajima, T.; Honda, Y.; Kitao, O.; Nakai, H.; Vreven, T.; Montgomery, J. A.; Peralta, J. E.; Ogliaro, F.; Bearpark, M.; Heyd, J. J.; Brothers, E.; 9

10 Kudin, K. N.; Staroverov, V. N.; Keith, T.; Kobayashi, R.; Normand, J.; Raghavachari, K.; Rendell, A.; Burant, J. C.; Iyengar, S. S.; Tomasi, J.; Cossi, M.; Rega, N.; Millam, J. M.; Klene, M.; Knox, J. E.; Cross, J. B.; Bakken, V.; Adamo, C.; Jaramillo, J.; Gomperts, R.; Stratmann, R. E.; Yazyev, O.; Austin, A. J.; Cammi, R.; Pomelli, C.; Ochterski, J. W.; Martin, R. L.; Morokuma, K.; Zakrzewski, V. G.; Voth, G. A.; Salvador, P.; Dannenberg, J. J.; Dapprich, S.; Daniels, A. D.; Farkas, O.; Foresman, J. B.; Ortiz, J. V.; Cioslowski, J.; Fox, D. J. Gaussian 09, revision C.01. Gaussian, Inc.: Wallingford CT, (2) Bruhn, T.; Schaumlöffel, A.; Hemberger, Y.; Bringmann, G. SpecDis: Quantifying the comparison of calculated and experimental electronic circular dichroism spectra. Chirality 2013, 25, (3) Wu, P.; Xue, J.; Yao, L.; Xu, L.; Li, H.; Wei, X. Org. Lett. 2015, 17, (4) Valle, P.; Figueroa, M.; Mata, R. J. Nat. Prod , ; (5) Jaivel, N.; Uvarani, C.; Rajesh, R.; Velmurugan, D.; Marimuthu, P. J. Nat. Prod ,

11 Figure S4. 1 H NMR spectrum of compound 1 in CDCl 3 (600 MHz). 11

12 Figure S5. 13 C NMR spectrum of compound 1 in CDCl 3 (150 MHz). 12

13 Figure S6. DEPT-90 spectrum of compound 1 in CDCl 3 (150 MHz). 13

14 Figure S7. DEPT-135 spectrum of compound 1 in CDCl 3 (150 MHz). 14

15 Figure S8. 1 H- 1 H COSY spectrum of compound 1 in CDCl 3 (600 MHz). 15

16 Figure S9. HSQC spectrum of compound 1 in CDCl 3 (600 MHz). 16

17 Figure S10. HMBC spectrum of compound 1 in CDCl 3 (600 MHz). 17

18 Figure S11. NOESY spectrum of compound 1 in CDCl 3 (600 MHz). 18

19 Figure S12. Expanded NOESY spectrum of compound 1 in CDCl 3 (600 MHz). (H 3-20/H 3-24) (H 3-20/H 3-19) (H-10/H-14) 19

20 Figure S13. 1 H NMR spectrum of 1 in DMSO (600 MHz). 20

21 Figure S C NMR spectrum of 1 in DMSO (600 MHz). 21

22 Figure S15. DEPT-90 spectrum of compound 1 in DMSO (150 MHz). 22

23 Figure S16. DEPT-135 spectrum of compound 1 in DMSO (150 MHz). 23

24 Figure S17. HSQC spectrum of 1 in DMSO (600 MHz). 24

25 Figure S18. 1 H- 1 H COSY spectrum of 1 in DMSO (600 MHz). 25

26 Figure S19. NOESY spectrum of 1 in DMSO (600 MHz). 26

27 Figure S20. HMBC spectrum of 1 in DMSO (600 MHz). 27

28 Figure S21. 1 H NMR spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). 28

29 Figure S C NMR spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (150 MHz). 29

30 Figure S23. DEPT-90 spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (150 MHz). 30

31 Figure S24. DEPT-135 spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (150 MHz). 31

32 Figure S25. 1 H- 1 H COSY spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). 32

33 Figure S26. HSQC spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). 33

34 Figure S27. HSQC spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). 34

35 Figure S28. NOESY spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). 35

36 Figure S29. Expanded NOESY spectrum of compound 2 in mixture of CDCl 3 and CD 3 OD (600 MHz). (H 3-20/H 3-24) (H-10/H-14) 36

37 Figure S30. 1 H NMR spectrum of 2 in DMSO (600 MHz). 37

38 Figure S C NMR spectrum of 2 in DMSO (600 MHz). 38

39 Figure S32. DEPT-90 spectrum of compound 2 in mixture of DMSO (150 MHz). 39

40 Figure S33. DEPT-135 spectrum of compound 2 in mixture of DMSO (150 MHz). 40

41 Figure S34. HSQC spectrum of 2 in DMSO (600 MHz). 41

42 Figure S35. 1 H- 1 H COSY spectrum of 2 in DMSO (600 MHz). 42

43 Figure S36. 1 H- 1 H COSY spectrum of 2 in DMSO (600 MHz). 43

44 Figure S37. NOESY spectrum of 2 in DMSO (600 MHz). 44

45 % Figure S38. HRESI TOF MS spectrum of compound 1. H Dec :11:17 H Waters SYNAPT G2-Si 1512A (0.226) 1: TOF MS ES e m/z Mass Calc. Mass mda PPM DBE Formula C 27 H 41 O 5 Limits: (1) Charge: -1; (2) Tolerance: 5.0 ppm; (3) Mass Mode: Monoisotopic; (4) Electron State: Even electron only; (5) Elements Used: 12 C(0~30); 1 H(0~60); 14 N(0~2); 16 O(0~10); 45

46 % Figure S39. HRESI TOF MS spectrum of compound 2. H Dec-2015 H Waters SYNAPT 1512A (0.166) 1: TOF MS ES e m/z Mass Calc. Mass mda PPM DBE Formula C25 H39 O4 Limits: (1) Charge: -1; (2) Tolerance: 5.0 ppm; (3) Mass Mode: Monoisotopic; (4) Electron State: Even electron only; (5) Elements Used: 12 C(0~30); 1 H(0~60); 14 N(0~2); 16 O(0~10); 46

Group 13 BN dehydrocoupling reagents, similar to transition metal catalysts but with unique reactivity. Part A: NMR Studies

Group 13 BN dehydrocoupling reagents, similar to transition metal catalysts but with unique reactivity. Part A: NMR Studies Part A: NMR Studies ESI 1 11 B NMR spectrum of the 2:1 reaction of i Pr 2 NHBH 3 with Al(NMe 2 ) 3 in d 6 -benzene 24 h later 11 B NMR ESI 2 11 B NMR spectrum of the reaction of t BuNH 2 BH 3 with Al(NMe