Fungal secondary metabolites represent a diverse group of bioactive natural products often produced at a restricted part of the life cycle (Keller et al. 2005).
Therefore, the lack of knowledge of this species recently assigned to section
The culture broth was filtered, using Whatman filter paper number 1, to remove the mycelia. The filtered broth was extracted three times with equal volumes of ethyl acetate (EtOAc) and evaporated until dryness using a rotary evaporator at 45°C. The resulted ethyl acetate extract of the filtrate (EAF) was reconstituted in methanol to get a concentrated stock solution of 200 mg/ml, which was used for further analysis.
The Orbitrap mass spectrometer was operated in both positive and negative mode with the use of the following parameter settings: spray voltage, 4 kV; sheath gas (N2 > 95%), 35 arbitrary units; auxiliary gas (N2 > 95%), 10 arbitrary units; capillary temperature, 290°C; S lens RF level, 50; heater temperature, 305°C. Two scan events were carried out, the LC-MS was used in full scan mode at a resolution of 70 000 fwhm (full width at half maximum) in the range m/z 90−1000 without the use of any lock masses. The maximum injection time (MIT) was 100 ms with one micro scan, and the automatic gain control (AGC) target was set to 1e6. The MS/MS was performed in parallel reaction monitoring (PRM) mode, in order to obtain two product ions for each target compound, at a resolution of 35 000 fwhm with collision energy (CE) of 30, the AGC and MIT were set at 2e5 and 200 ms, respectively. The instrument control and data analysis were performed by Thermo Fisher Xcalibur v. 3.0.63 software.
The DPPH radical scavenging activity was determined according to Jakovljević et al. (2014). Different concentrations of EAF were prepared: 12.5, 25, 50, 100, 200, 400 μg/ml. To 1 ml of DPPH solution, 1 ml of each concentration was added. After incubation for 30 min at 37°C, the absorbance was measured at 517 nm against the blank. Ascorbic acid was used as positive control. The percentage of the DPPH scavenging activity was calculated using the formula:
Metabolites of
Peak No. | Putative compound name | Adduction | Measured mass (m/z) | Productions (m/z) | tRa (min) | Refb |
---|---|---|---|---|---|---|
1 | Asperlactone | [M+H] + | 185.08081 | 141.05444, 113.05948 | 1.73 | Vishwanath et al. 2009 |
2 | NI | [M+Na] + | 211.06121 | 195.03455, 133.02816 | 1.78 | - |
3 | NI | [M+H] + | 308.10992 | 280.95587, 145.10104 | 3.38 | - |
4 | NI | [M+H] + | 327.04734 | 309.18594, 191.15396 | 4.28 | - |
5 | Emodin | [M+H]- | 269.04590 | ND | 5.42 | Sulyok et al. 2007; Lehner et al. 2011; Micheluz et al. 2016 |
6 | NI | [M+H]+ | 251.09145 | 233.09526, 204.09319 | 5.49 | - |
7 | Sterigmatocystin | [M+H]+ | 325.07053 | ND | 6.62 | Lehner et al. 2011; Micheluz et al. 2016 |
8 | NI | [M+Na] + | 423.25041 | 405.27911, 239.14821 | 7.27 | - |
9 | Deoxybrevianamide E | [M+H]+ | 352.20352 | ND | 8.04 | Lehner et al. 2011; Micheluz et al. 2016 |
10 | Norsolorinic acid | [M+H]- | 369.09892 | ND | 8.24 | Micheluz et al. 2016 |
a - Retention time; b - Reference; ND - not detected; NI - not identified
Antimicrobial activity and minimum inhibitory concentration (MIC) of ethyl acetate extract of
Zone of inhibition (mm)a | MIC of ethyl acetate extract (mg/ml) | ||
---|---|---|---|
Ethyl acetate extract | Positive controlb | ||
0 | 23.4 ± 0.3 | - | |
10.0 ± 0.3 | 25.0 ± 0.4 | 2.5 | |
0 | 20.8 ± 0.4 | - | |
8.5 ± 0.6 | 21.5 ± 0.3 | 5 | |
SABLc | 12.8 ± 0.3 | 19.8 ± 0.2 | 0.625 |
MRSEd | 14.0 ± 0.2 | 19.2 ± 0.2 | 0.625 |
20.6 ± 0.8 | 22.8 ± 0.4 | 0.325 | |
13.0 ± 0.3 | 25.7 ± 0.5 | 1.25 |
- Mean of three replicates (±) SD
- Chloramphenicol and ketoconazole were used as the positive control for bacteria and yeasts respectively
-
- Methicillin resistant
0- no zone of inhibition
Table II showed also the MIC values of
Antioxidant activities of
Activity | Extract | Ascorbic acida |
---|---|---|
DPPHa | 89.28 ± 0.32 | 91.39 ± 0.39 |
ABTSa | 92.93 ± 0.30 | 93.03 ± 0.45 |
Total phenolic content (mg GAE/g) | 85.76 ± 0.96 | - |
- Percentage of inhibition at a concentration of400 μg/ml. Values are mean of three replicates (±) SD
Fungi are major producers of secondary metabolites with different biological activities and various chemical structures (Abo-Elmagd 2014). Until today, except for
Our study demonstrated that
The ability of
In the antimicrobial assays, our study indicated that
Our study also brings additional data on
In conclusion, UHPLC-MS/MS analysis revealed that