Free radical scavenging activity of different solvent extract....

1. “Free radical scavenging activity of different solvent extract
(Water extract, Ethanol : water extract & Methanol extract) of
Cordyceps militaris fruiting body
Anamika kori
(M.Sc Biochemistry)
Rani durgawati University

3. Cordyceps fungi ( Medicinal mushroom)
• Cordyceps , the name given to fungi on insect, it is always has been one of the rarest and most
mysterious mushroom.
• Cordyceps mushroom used as a medicine in China and Tibet.
• Cordyceps , is an ascomycetes mushroom that grows only on the dead subterranean larva of
some species of moths, known as Himalayan ghost moths .
• Cordyceps is a entmopathogenic fungus .
• Cordyceps militaris and Cordyceps sinensis ,are two very beneficial Cordyceps species .
Cordyceps militaris Cordyceps sinensis

4. Cordyceps militaris
• Cordyceps militaris (Winter worm and summer grass) is one of the most valued traditional Chinese
medicines for fighting cancer, increasing longevity, and improving immunity. In recent years number of
studies had been carried out to cure dreadful non-communicable diseases like cancer, HIV, diabetes
etc. with the use of herbal or natural medicine.
• The host of Cordyceps militaris is Lepidopteran pupa and the shade of its fruiting is yellow or orange .
• Cordyceps militaris Produce a diversity of bioactive compounds such as nucleoside, Cordycepin and its
derivatives, ergosterol, polysaccharides, glycoproteins . Cordycepin is a most important bioactive
compound.
• Cordyceps militaris have various health beneficial properties –
1. Anti-inflammatory properties 6. Reduce physical fatigue
2. Antioxidant properties 7. Helps to treat chronic kidney disease
3. Improves liver functions 8. Beneficial Oxygen uptake in the body
4. Boost physical performance 9. Blood pressure regulation
5. Help manage blood glucose 10. Anti—Aging and antitumor properties

5. ANTIOXIDANTS
• Antioxidant is a molecule capable of inhibiting the oxidation of other molecules.
• Oxidation reactions can from free radicals and these start chain reactions that damage cells.
• Antioxidants terminate these chain reactions by removing free radical intermediates and
inhibit other oxidation reactions.
• Antioxidant are help to neutralize free radicals in our bodies, and this is thought to boost
overall health.
Free radicals are waste substances produced by cells as the body processes food reacts to the
environment . If the body cannot process and remove free radicals efficiently, oxidative
stress can result. This can harm cells and body function. free radicals are also known as
reactive oxygen species (ROS).

6. .
Factor that increase the production of free radicals in the body:-
• Internal- inflammation, mental stress.
• External- pollution, UV exposure, cigarette smoke, drug.
NEED OF ANTIOXIDANTS (antioxidant stop oxidative stress)
Oxygen
Cell
Energy
(ATP)
Free radical as a
by product
Damage
cells
Damage
tissues
Diseases
and
disorders
DPPH DPPH (2,2-diphenyl-1picrylhydrazyl) it is a dark colored crystalline powder
composed of stable free-radical molecules. DPPH has major applications in laboratory
research: A monitor of chemical reactions involving radicals, most notably it is a
common antioxidant assay

7. ANTIOXIDANT PROPERTY OF CORDYCEPS MILITARIS
• Cordyceps militaris have antioxidant property, its prevent or reverse oxidative
damage to a target molecule and can be synthesized in vivo or taken from the diet
and they are also prevent many chronic disease including cancer, diabetes or
neurodegenerative disorders.
• C. militaris is a natural antioxidant and their use in the production of new generation
of food has increased, food have rich antioxidant Reactive Oxygen species(ROS) with
systemic health diseases.
• Cordyceps mushroom are promising in this respect as their fruiting bodies are readily
available and rich in natural antioxidant.
C. militaris dry fruiting bodies used as antioxidant
.

8. Different extraction solvent used for CM antioxidant
activity (Method)

9. Screening of Anti-oxidant activity of the Cordyceps militaris fruiting body of
different extraction sample by - spectrophotometric DPPH radical scavenging
activity method :
•

10. Absorbance of the antioxidant activity of water extraction sample of cordyceps militaris fruiting
body after 30min, 1hrs, 1:30 hrs, 2nd days, 3rd days of incubation-:
Absorbance of the antioxidant activity in 517nm
S No. Different conc.
of (µl) of
sample
DPPH Absorbance at
517nm
30min. 1 hrs 1:30 hrs 2nd days 3rd days
1 0.5 µl 3 ml 0.056 0.023 0.053 0.198 0.226
2 01 µl 3 ml 0.082 0.032 0.116 0.119 0.136
3 02 µl 3 ml 0.092 0.072 0.080 0.102 0.075
4 03 µl 3 ml 0.090 0.052 0.093 0.108 0.053
5 04 µl 3 ml 0.365 0.289 0.253 0.126 0.066
6 05 µl 3 ml 0.145 0.028 0.054 0.147 0.069
7 06 µl 3 ml 0.062 0.037 0.061 0.142 0.046
8 07 µl 3 ml 0.081 0.056 0.302 0.162 0.055
9 08 µl 3 ml 0.131 0.026 0.265 0.220 0.082
10 09 µl 3 ml 0.149 0.018 0.071 0.049 0.036
11 10 µl 3 ml 0.048 0.040 0.025 0.216 0.244
12 PC 3 ml 0.065 0.057 0.051 0.081 0.088
13 NC 3 ml 0.816 0.894 0.976 1.190 1.141
Free radical scavenging activity (%) of water solvent
extract
S No. Different conc.
(µl) of sample
DPPH Absorbance at 517nm
30min. 1 hrs 1:30 hrs 2nd days 3rd days
1 0.5 µl 3 ml 93.137 97.427 94.569 83.361 80.192
2 01 µl 3 ml 89.950 96.420 88.114 90.000 88.080
3 02 µl 3 ml 88.725 91.946 91.803 91.428 92.426
4 03 µl 3 ml 88.970 94.183 90.471 90.924 95.354
5 04 µl 3 ml 55.269 67.673 74.077 89.411 94.215
6 05 µl 3 ml 82.230 96.868 94.467 87.647 93.952
7 06 µl 3 ml 92.401 95.861 93.75 88.067 95.968
8 07 µl 3 ml 90.073 93.736 69.057 86.386 95.179
9 08 µl 3 ml 83.946 97.091 72.848 81.512 92.813
10 09 µl 3 ml 81.740 97.986 92.725 95.882 96.844
11 10 µl 3 ml 94.117 95.525 97.438 81.848 78.615
12 PC 3 ml 92.034 93.624 94.774 93.193 92.287
13 NC 3 ml 000 000 000 000 000

11. Absorbance of the antioxidant activity of ethanol and water (50:50) extraction sample of
Cordyceps militaris fruiting body after 30min, 1hrs, 1:30 hrs, 2nd days, 3rd days of incubation-:
Absorbance of antioxidant activity in 517nm
S No. Different conc.
ogf (µl) of
sample
DPPH Absorbance at 517nm
30min. 1 hrs 1:30 hrs 2nd days 3rd days
1 0.5 µl 3 ml 0.186 0.162 0.160 0.327 0.317
2 01 µl 3 ml 0.223 0.234 0.244 0.395 0.397
3 02 µl 3 ml 0.234 0.222 0.200 0.091 0.126
4 03 µl 3 ml 0.306 0.209 0.316 0.148 0.168
5 04 µl 3 ml 0.255 0.202 0.254 0.105 0.156
6 05 µl 3 ml 0.282 0.231 0.239 0.166 0.155
7 06 µl 3 ml 0.249 0.259 0.250 0.103 0.111
8 07 µl 3 ml 0.349 0.273 0.330 0.128 0.121
9 08 µl 3 ml 0.392 0.238 0.398 0.075 0.130
10 09 µl 3 ml 0.362 0.200 0.360 0.068 0.049
11 10 µl 3 ml 0.411 0.190 0.400 0.120 0.133
12 PC 3 ml 0.065 0.057 0.051 0.081 0.088
13 NC 3 ml 0.816 0.894 0.976 1.190 1.164
Free radical scavenging activity (%) of ethanol : water extract
S No. Different conc.
ogf (µl) of
sample
DPPH Absorbance at 517nm
30min. 1 hrs 1:30 hrs 2nd days 3rd days
1 0.5 µl 3 ml 77.205 81.879 83.606 72.521 72.766
2 01 µl 3 ml 72.671 73.825 75.00 66.806 65.893
3 02 µl 3 ml 71.323 75.167 79.508 92.352 89.175
4 03 µl 3 ml 62.50 76.621 67.622 87.563 85.567
5 04 µl 3 ml 68.75 77.404 73.975 91.176 86.597
6 05 µl 3 ml 65.441 74.161 75.512 86.050 86.683
7 06 µl 3 ml 69.485 71.364 74.385 91.344 90.463
8 07 µl 3 ml 57.230 69.463 66.188 89.243 89.604
9 08 µl 3 ml 51.960 73.378 59.221 93.697 88.831
10 09 µl 3 ml 55.637 77.628 63.114 94.285 95.790
11 10 µl 3 ml 49.632 78.747 59.016 89.915 88.573
12 PC 3 ml 92.034 93.624 94.774 93.193 92.439
13 NC 3 ml 000 000 000 000 000

12. Absorbance of the antioxidant activity of Methanol extraction sample of Cordyceps militaris
fruiting body after 30min, 1hrs, 1:30 hrs, 2nd days, 3rd days of incubation-
Absorbance of antioxidant activity in 517nm
S No. Different conc.
ogf (µl) of
sample
DPPH Absorbance at 517nm
30 min 1 hrs 1:30 hrs 2nd days 3rd days
1 0.5 µl 3 ml 0.466 0.364 0.419 0.390 0.316
2 01 µl 3 ml 0.524 0.409 0.410 0.383 0.395
3 02 µl 3 ml 0.410 0.293 0.287 0.242 0.241
4 03 µl 3 ml 0.415 0.229 0.286 0.235 0.242
5 04 µl 3 ml 0.411 0.344 0.233 0.228 0.396
6 05 µl 3 ml 0.454 0.268 0.287 0.224 0.248
7 06 µl 3 ml 0.377 0.200 0.210 0.205 0.209
8 07 µl 3 ml 0.467 0.223 0.221 0.208 0.189
9 08 µl 3 ml 0.378 0.333 0.231 0.245 0.200
10 09 µl 3 ml 0.450 0.218 0.227 0.225 0.212
11 10 µl 3 ml 0.464 0.217 0.235 0.186 0.169
12 PC 3 ml 0.065 0.057 0.251 0.088 0.081
13 NC 3 ml 0.816 0.894 0.976 1.190 1.164
Free radical scavenging activity (%) of methanol solvent
extract
S No. Different conc.
of (µl) of
sample
DPPH Absorbance at 517nm
30min. 1 hrs 1:30 hrs 2nd days 3rd days
1 0.5 µl 3 ml 42.892 59.284 57.069 67.226 72.852
2 01 µl 3 ml 35.784 54.250 57.991 67.815 66.065
3 02 µl 3 ml 49.754 67.225 70.594 76.663 79.295
4 03 µl 3 ml 49.142 74.384 70.696 80.252 79.209
5 04 µl 3 ml 49.632 61.521 76.127 80.840 65.976
6 05 µl 3 ml 44.362 70.022 70.594 81.176 78.694
7 06 µl 3 ml 53.799 77.628 78.483 82.773 82.044
8 07 µl 3 ml 42.769 75.055 77.356 82.521 83.762
9 08 µl 3 ml 53.676 62.751 76.331 79.411 82.817
10 09 µl 3 ml 44.852 75.615 76.741 81.092 81.786
11 10 µl 3 ml 43.137 75.727 75.922 84.369 85.486
12 PC 3 ml 92.034 93.624 74.282 92.605 93.041
13 NC 3 ml 000 000 000 000 000

13. Result
Result obtained for DPPH radical scavenging % according to
spectrophotometric analysis by different solvent extract method.
S.NO. Sample extracts Conc.of
sample for
highest
scavenging
%
Highest
scavenging %
Conc.of
sample for
lowest
scavenging
%
Lowest scavenging
%
1. Water extract 09 µl 97.98% 04 µl 55.26 %
2. Ethanol:water
extract
09 µl 95.79 % 08 µl 51.96 %
3. Methanol extract 10 µl 85.48% 07 µl 42.76%

14. Free radical scavenging activity showed by fruiting body of C. militaris by water extract
After 30 min for adding DPPH After 1 hours for adding DPPH
After 1:30 hours for adding DPPH 2nd days For adding DPPH 3rd days for adding DPPH
water solvent extract add DPPH

15. Free radical scavenging activity showed by fruiting body of C. militaris of ethanol :water
extract
ethanol :water solvent extract add DPPH After 30 minute for adding DPPH After 1 hours for adding DPPH
After 1:30 hours for adding DPPH 2nd days For adding DPPH 3rddays For adding DPPH

16. Free radical scavenging activity showed by fruiting body of C. militaris for methanol extract
methanol solvent extract add DPPH After 30 min for adding DPPH After 1hours for adding DPPH
After 1:30 hours for adding DPPH 3rddays For adding DPPH
2nddays For adding DPPH

17. CONCLUSION
• Medicinal Cordyceps are very important to human beings in preserving our health.
• The aim of this study is provide recent insights on the fruiting bodies of different solvent extract their
experminent evidences of high antioxidant activities.
• From this study, it is evident that Cordyceps produce a variety of compound mainly fruiting body for
various disorder .
• Cordyceps militaris fruiting body showed very high free radical scavenging activity.
• The antioxidant properties of water and ethanol extract from fruiting bodies of Cordyceps militaris was
evaluated by using a range of testing systems in the present study and the result suggested that the
extract had direct and potent antioxidant activities.
• The antioxidant activities of the extract from cultivated fruit bodies of Cordyceps militaris in the assays
performed in this study prompt the possible uses of the cultivated C. militaris to meet the needs of the
natural antioxidant sources from human health.
• Where water extract of Cordyceps militaris fruiting body showed broad scavenging percentage (97.98%)
of free radical using 9µl of sample. Ethanol:water extract of fruiting body scavenged (95.79%)with 9µl of
sample. Cordyceps militaris fruiting body of methanol extract also showed scavenging activity but
compare to water and ethanol extract , methanol extract show low scavenging free radical activity their
are showed scavenging percentage (85.48%) with 10µl of sample.
The different solvent extract show water and ethanol is best on the grounds of
safety and effieiency. Hence was chosen as the best solvent in the extraction
experiment .