Background
Materials and methods
Chemicals
Extraction methods
Synergy testing
Phytochemical analyses
Minerals estimation
HPLC analysis for identification and quantification of phenolics
Determination of the antioxidant activity
The TAC assay
Radical scavenging activity
Ferric reducing power
Anti-lipid peroxidation assay
Cytotoxicity and anti-inflammatory assays
Cytotoxicity assay
Anti-inflammatory assay
TNF-α, NO, and lipid peroxidation assays
Quantitative real-time polymerase chain reaction (qRT-PCR) of some inflammatory mediators
Cellular antioxidant indices
Data analysis
Results and discussion
Phytochemical and minerals contents
Constituents | Ganodermalucidum extract (GLE) | Chlorella vulgaris extract (CVE) |
---|---|---|
Phytochemicals (mg/g extract) | ||
Total phenolics | 4.44 ± 0.08
a
| 5.18 ± 0.58
a
|
Flavonoids | 0.14 ± 0.00
b
| 3.46 ± 0.21
a
|
Tannins | 1.24 ± 0.00
a
| 1.02 ± 0.01
a
|
Triterpenoids | 23.88 ± 0.00
a
| 5.45 ± 0.25
b
|
SPs | 3.66 ± 0.03
a
| 1.04 ± 0.05
b
|
Minerals (μg/g extract) | ||
Cu | 6.4 ± 0.00
b
| 30.08 ± 0.01
a
|
Se | 606.7 ± 0.00
a
| 276 ± 0.00
b
|
Zn | 16.3 ± 0.00
b
| 4300 ± 0.01
a
|
Fe | 1100 ± 0.00
b
| 4180 ± 0.00
a
|
Ca | 00.00 | 20,000 ± 0.00 |
Mg | 00.00 | 4260 ± 0.01 |
Antioxidant activities of single and combined extracts
In vitro antioxidant models | |||||||||||
TAC (mg Asc eq/ g extract) | Effect | NO scavenging activity (IC50, μg/mL) | Effect | DPPH scavenging activity (IC50, μg/mL) | Effect | Reducing power (IC50, μg/mL) | Effect | Superoxide scavenging activity (IC50, μg/mL) | Effect | ||
GLE-CVE |
OV
|
301.00 ± 1.00
a
| Sy |
46.12 ± 2.92
a
| Sy |
29.48 ± 1.45
a
| Sy |
103.47 ± 2.04
a
| An |
30.01 ± 3.45
a
| Sy |
EV | 38.83 ± 2.16
b
| 1111.02 ± 128.75
b
| 169.29 ± 9.36
b
| 71.98 ± 1.99
b
| 660.73 ± 14.92
b
| ||||||
Hydroxyl radical scavenging activity (IC50, μg/mL) | Effect | Anti-Lipid peroxidation activity (IC50, μg/mL) | Effect | ||||||||
OV
|
131.6 ± 5.42
a
| Sy |
54.15 ± 2.45
a
| Ad | |||||||
EV | 213.18 ± 2.13
b
| 51.49 ± 0.96
a
| |||||||||
Inflammatory mediators | |||||||||||
NO (nmol/mL) | Effect | iNOS (Expression) | Effect | COX-2 (Expression) | Effect | NF-кB (Expression) | Effect | TNF-α (mmol/mL) | Effect | ||
GLE-CVE |
OV
|
39.26 ± 4.37
a
| Sy |
0.146 ± 0.006
a
| Sy |
0.056 ± 0.001
a
| Sy |
0.49 ± 0.015
a
| Sy |
242.88 ± 18.5
a
| Sy |
EV | 52.08 ± 1.08
b
| 1.29 ± 0.29
b
| 0.18 ± 0.002
b
| 0.85 ± 0.038
b
| 422.04 ± 2.65
b
| ||||||
Oxidative stress parameters | |||||||||||
Lipid peroxidation (nmoL/mL) | Effect | GSH (nmoL/mL) | Effect | Superoxide dismutase (SOD, IU/mg protein) | Effect | Glutathione peroxidase (GPX, IU/mg protein) | Effect | Glutathione reductase (GR, IU/mg protein) | Effect | ||
GLE-CVE |
OV
|
7.29 ± 0.84
a
| Sy |
0.59 ± 0.01
a
| Sy |
0.66 ± 0.0002
a
| Sy |
0.07 ± 0.001
a
| Sy |
3.25 ± 0.06
a
| Ad |
EV | 59.58 ± 1.62
b
| 0.48 ± 0.0002
b
| 0.63 ± 0.002
b
| 0.06 ± 0.0001
b
| 3.19 ± 0.02
a
|