Investigation on the chemical constituents of the marine-derived fungus strain Aspergillus brunneoviolaceus MF180246

Abstract Aspergillus have been proven to be excellent resources for new natural products. During our systematic biodiversifying new compounds from marine derived fungi, one novel compound, asperbrunneo acid (1), along with seven bistetrahydroxanthone analogues, secalonic acid D (2), secalonic acid F (3), secalonic acid F1 (4), secalonic acid H (5), penicillixanthone A (6), chrysoxanthone C (7), and asperdichrome (8), one ketodivinyllactonic steroid, herbarulide (9), as well as one tyrosine-derived compound, aspergillusol A (10), were isolated from the marine-derived fungus Aspergillus brunneoviolaceus MF180246. These structures were elucidated by HRMS, 1 D and 2 D NMR analysis. Compound 1 possessed the first reported new carbon skeleton natural product. Compounds 1, 4, 5, 6, 7 and 8 showed antibacterial activity against Staphylococcus aureus with minimum inhibitory concentration values of 200, 25, 50, 6.25, 50, and 25 µg/ml, respectively. Graphic Abstract


Introduction
Fungi of the genus Aspergillus have been proved to be excellent resources for the discovering new natural chemical entries (Zhang et al. 2018;Lima et al. 2019;Zhang et al. 2019).With the deep investigation on the secondary metabolites from marine-derived fungi, a large number of novel compounds with variety bioactivities were obtained from fungi of the genus Aspergillus (Zhao et al. 2016;Blunt et al. 2018;Carroll et al. 2019).During our ongoing investigation on secondary metabolites from marine-derived fungi, a fungus strain MF180246 was isolated from the mud sample collected from the root of mangrove forest of Xinglin Bay, Xiamen, China, which identified as Aspergillus brunneoviolaceus.Further scaled-up fermentation and chemical studies on this fungus resulted in the identification of one new compound, asperbrunneo acid (1), together with nine known compounds, including secalonic acid D (2) (Ma et al. 2011), secalonic acid F (3) (Andersen et al. 1977;Yamazaki et al. 2016), secalonic acid F1 (4) (Rotinsulu et al. 2017), secalonic acid H (5) (Chen et al. 2017), penicillixanthone A (6) (Jiang et al. 2017), chrysoxanthone C (7) (Zhen et al. 2018), and asperdichrome (8) (Yamazaki et al. 2016), one ketodivinyllactonic steroid, herbarulide (9) (Krohn et al. 1999), one tyrosine-derived compound, aspergillusol A (10) (Ingavat et al. 2009).To the best of our knowledge, this is the first report on chemical investigation on Aspergillus brunneoviolaceus.In this study, we described the isolation, structure elucidation and antibacterial activity of these compounds.

Fungal material
Strain MF180246 was isolated from a mud sample collected from the mangrove forest in Xinglin Bay, Xiamen, China, and grown on a potato dextrose agar plate at 28 C.This strain was identified as Aspergillus brunneoviolaceu based on gene sequence analysis of b-tubulin (BT) using conventional primer pair of BT2A (5 0 -GGTAACCAAATCGGTGCTGCTTTC-3 0 ) and BT2B (5 0 -ACCCTCAGTGTAGTGACCCTTGGC-3 0 ).

Fermentation and extraction
HS5-2 was incubated at 28 C for 7 d on a potato dextrose agar.HS5-2 was cut about 1 cm 2 with a sterilized knife and placed into teen 1 L conical flasks, each containing solid medium consisting of rice (200 g) and distilled water (150 mL), and the flasks were incubated stationary for 38 d at 28 C. The cultures were extracted three times by EtOAc:MeOH (80:20), and the combined extracts were reduced to dryness in vacuo and the residue was partitioned between EtOAc and H 2 O. Then the EtOAc layer was dried in vacuo to yield a dark residue (14.38 g).

Antimicrobial assays
The antimicrobial assays were performed according to the Antimicrobial Susceptibility Testing Standards outlined by the Clinical and Laboratory Standards Institute document M07-A7 (CLSI) and our previous report (Xu et al. 2021).
12 and H-13 to C-2 revealed the position of keto of C-1 and double bond carbon of C-2.The HMBC correlations from H 3 -18 to C-6, C-9, C-10 and C-11 indicated the linkage from C-10 to C-6, C-9, and C-11.The acrylic acid moiety and connection between C-7 and C-15 were confirmed by the HMBC crossing peaks from H 2 -16 to C-7, C-15 and C-17.The HMBC correlations from H 2 -5 to C-3 and from H 2 -4 to C-2 suggested the attachment between C-2 and C-3.Thus, the planer structure of compound 1 was established.The ROESY correlation from H-12 (3.01, m) toH-13 (2.31, m)   and from H 3 -18 (1.00, s) to H 3 -19 (0.85, d, J ¼ 6.0 Hz), and the absence from H-12 (3.01, m) and H 3 -19 (0.85, d, J ¼ 6.0 Hz) indicated C-18 and C-19 were the same side of the ring system and H-12 on the other side.The ROESY correlation from H 3 -18 (1.00, s) to H-7 (2.78, m) revealed the trans-conformation between C-18 and the acrylic acid moiety.H-6 and acrylic acid moiety were assigned to cis on the basses of ROESY correlation from H-6 (2.06, m) toH-16b (5.62, s).The ROESY correlations between H-4 and H-12, between H-12 and H-13 revealed the relative configurations of C-12 and C-13.Finally, the relative configurations were determined as show in Figure1.3.Experimental section3.1.General experimental procedures Optical rotations ([a] D ) were measured on Anton Paar MCP 200 Modular Circular Polarimeter (Austria) in a 100 Â 2 mm cell at 22 C. 1 H and 13 C NMR, along with