An anti-inflammatory isoflavone from soybean inoculated with a marine fungus Aspergillus terreus C23-3

Figures & data

Figure 1. Structures of compounds 1–8.

Table 1. The NMR date for compound 1.

Position	δc^a (in CDCl3)	δH^a (J in Hz) (in CDCl3)	δH^b(J in Hz) (in DMSO-d6)
2	152.0	7.92 (1 H, s)	8.38 (1 H, s)
3	124.8
4	175.7
5	105.2	7.63 (1 H, s)	7.43 (1 H, s)
6	147.9
7	154.4
6-OMe	56.9		3.99 (3 H, s)
7-OMe	56.9		4.00 (3 H, s)
8	100.0	6.88 (1 H, s)	7.22 (1 H, s)
9	152.4
10	118.2
1’	128.7
2’	126.5	7.43 (1 H, d, J = 1.9 Hz)	7.39 (1 H, d, J = 1.8 Hz)
3’	127.4
4’	159.8
5’	104.5	6.84 (1 H, d, J = 8.2 Hz)	6.78 (1 H, d, J = 8.2 Hz)
6’	129.1	7.25 (2 H, dd, J = 8.2, 1.9 Hz)	7.27 (2H, dd, J = 8.2, 1.8 Hz)
7’	31.4	3.21 (2H, m)	3.16 (2H, m)
8’	89.8	4.66 (1 H, t, J = 9.1 Hz)	4.58 (1 H, t, J = 8.9 Hz)
9’	71.9
10’	26.6	1.36 (3 H, s)	1.14 (3 H, s)
11’	24.8	1.24 (3 H, s)	1.15 (3 H, s)
8ʹ-OH		Not observed	4.63 (1 H, s)

^a C NMR at 125 MHz and ¹ HMR at 500 MHz recorded in CDCl₃, assigned by HSQC and HMBC; ^{b 1} HMR recorded at 500 MHz in DMSO-d6.

Figure 2. The ¹H-¹H COSY and key HMBC correlations of compound 1.

Figure 3. The experimental ECD spectrum of compound 1 and the calculated one for its S-isomer.

Figure 4. The HPLC spectra (detected under 254 nm) of compound 1 and the extracts of A. terreus C23-3 culture on soybean medium with 5-azacytosine (the main fermentation), A. terreus C23-3 culture on soybean medium (control), and non-inoculated soybean medium. (1: psoralenone; 2: genistein; 5: biochanin A; 7: butyrolactone I).

Figure 5. Evaluation of cytotoxicity (line) and inhibition of LPS-induced NO production (bars) by compound 1 at 32 μM. 1% ethanol alone served as the negative control, and with DMSO (0.75%) was used as the positive control. (n = 3; **** and ***represent adjusted P values of <0.0001 and 0.0005, respectively). Amount of NO production calculated from a standard curve.

Table 2. Antimicrobial activities of compounds 2–8 (paper disk method, 10 µg/disk, n = 3).

Compounds	Diameters of inhibition zone (mm)
	MRSA	V. parahemolyticus	C. albicans	P. aeruginosa
2	-	11.9 ± 0.4	-	12.0 ± 0.1
3	10.0 ± 0.1	11.0 ± 0.2	13.0 ± 0.1	13.1 ± 0.3
4	10.1 ± 0.1	11.9 ± 0.2	13.8 ± 0.4	-
5	11.7 ± 0.2	13.8 ± 0.1	11.1 ± 0.1	11.0 ± 0.2
6	10.8 ± 0.2	11.9 ± 0.3	12.9 ± 0.1	10.6 ± 0.1
7	14.9 ± 0.2	22.1 ± 0.5	20.8 ± 0.4	23.2 ± 0.3
8	9.2 ± 0.2	9.9 ± 0.4	-	-
Ampicillin	16.0 ± 0.4	23.0 ± 0.4	nt	24.0 ± 1.4
Ketoconazole	nt	nt	20.0 ± 0.0	nt

-: no activity. nt: not tested. 2: genistein; 3: daidzein; 4: 4ʹ-hydroxy-6,7-dimethoxyisoflavone; 5: biochanin A; 6: psoralenol; 7: butyrolactone I; 8: blumenol A.

Table 3. DPPH free radical scavenging, acetylcholinesterase inhibitory, and larvicidal activities of compounds 2–8 (n = 3).

Compounds	AChE inhibition (IC50, µM)	DPPH scavenging (EC50, µM)	Brine shrimp larvicidality (LC50, µM)
2	42.5 ± 3.1	96.7 ± 2.5	13.7 ± 1.4
3	>102.4	>102.4	>102.4
4	>102.4	>102.4	98.5 ± 3.2
5	>102.4	>102.4	11.6 ± 2.1
6	>102.4	>102.4	68.2 ± 1.7
7	>102.4	25.0 ± 2.7	12.6 ± 2.8
8	>102.4	>102.4	89.1 ± 3.5
Donepezil HCl	0.3 ± 0.5	nt	nt
Vitamin C	nt	31.8 ± 1.2	nt
Hg(NO3)2	nt	nt	77.0 ± 2.4

nt: not tested. 2: genistein; 3: daidzein; 4: 4ʹ-hydroxy-6,7-dimethoxyisoflavone; 5: biochanin A; 6: psoralenol; 7: butyrolactone I; 8: blumenol A.