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Article Evaluation of Antioxidant and Cytotoxic Activity of Bioactiv...

1 -  t/c * 100

Where, 

T= sample reading, c= control reading 

I recommend that the SC 50 value calculations.). In the presence of antioxidants, the purple colour of DPPH decays, and the change of absorbance is monitored at 517 nm against reagent and sample blank. Briefly, 0.75 mL of a 0.1 mM DPPH in methanol was mixed with equal volume of methanolic honey extracts, and incubated in dark for 50 min, and absorbance was measured at 517 nm in the presence of different concentration of samples (0.1-10 mg/mL). Radical scavenging activity of DPPH was expressed as SC50, which represents the concentration of the extract (mg/mL) required to inhibit 50% of the free radical-scavenging activity; higher SC50 value indicates lower radical scavenging activity.

you can search my manuscript.. 

 All anwers ok.

Determination of antioxidant efficiency

a) Free radical scavenging capacity

The free radical scavenging capacity of various solvent extracts of guava seeds was measured by using 1, 1- diphenyl-2-picrilhydrazyl (DPPH) assay (Juntachote and Berghofer 2005). Aliquot (100 μL) of extract was mixed with 5 ml of 6×10–3 M methanolic solution of DPPH radical. The mixture was shaken vigorously and left to stand for 30 min in the dark. The absorbance was then measured at 517 nm

against a blank. The control was prepared, as above, without any extract and methanol was used for the base line correction. The radical-scavenging activity was expressed as percentage of inhibition and calculated using the following formula:

% Radical scavenging activity  

% Radical scavenging activity

=(  ½ðAbs control−Abs sampleÞ/ Abs control x 100:

Where, Abs control is the absorbance of DPPH radical +

methanol; Abs sample is the absorbance of DPPH radical +

sample extract/standard.

References.

Juntachote T, Berghofer E (2005) Antioxidative properties and stability of

ethanolic extracts of Holy basil and Galangal. Food Chem 92:193–

202

El Anany, A.M. (2014) .  Nutritional composition, antinutritional factors, bioactive compounds and antioxidant activity of guava seeds (Psidium Myrtaceae) as affected by roasting processes . Journal of Food Science and Technology

ISSN 0022-1155 Volume 52 Number 4

J Food Sci Technol (2015) 52:2175-2183

DOI 10.1007/s13197-013-1242-

I agree with Oktay Yildiz to express it as EC50 values: to do so you should incubate  methanolic solutions with  different concentration of the antioxidant and the same concentration of DPPH for each one in the darkness for a certain time, i.e. 30 minutes (exactly for each sample), then measure the absorbance at 515-517 nm. The value  of DPPH, expressed as %Remaining DPPH will be calculated as:

%DPPH remaining = Absorbance sample/ Absorbance control  x 100

The control will be a methanolic solution with just DPPH

Then you will represent the values of %DPPH remaining versus sample concentration of your samples, so you can get the value of EC50 (Effective concentration) which is the concentration of antioxidant able to destroy 50% of the initial DPPH. The values are usually given as micromolar concentration.

For more information you can check Prior, R.L.; Wu, X.; Schaich, K. J. Agric. Food Chem. 2005, 53, 4290-4302.

I don't understand why all these answers are different. Does nobody use a standard like ascorbic acid or different dilutions of DPPH to make a standard curve?

%of  RADICAL SCAVENGING ACTIVITY = (Ab of control-Ab of sample)/Ab of controlx100

Calculate % if RSA for all concentrations used for the assay and draw a graph % RSC Vs sample concentration.

Using the equation of the graph find the sample concentration which gives 50% RSA (IC 50)

Express it as the DPPH radical scavenging activity of the sample.

Measurement of antioxidant activity by DPPH

Extraction.

Ten g (10 g) of sample were homogenized with100 ml methanol for 1 min and centrifuged at 10 000 rpm for 15 min at 4 ◦C. The clear supernatant was transferred to a glass bottle and measured immediately for total antioxidant activity using DPPH assay. 100 μL of sample were added to 5 ml DPPH solution and the absorbance of DPPH reagent was determined at 515 nm after 30 min of incubation(15). The inhibition percentage of the absorbance was calculated as follows:

Inhibition % = (Abst0 - Abst30 min)/ Abst0 ×100

Abst0 min was the absorbance of DPPH at time 0. Abst30 min was the absorbance of DPPH after 30 min of incubation.

Hi,

I use standard curve, but still wondering how to express the results (only Total fraction, not methanolic). Does anyone can explain me how to do one of the options (mM/Kg, mmol/l.)? 

Hi Mr. Osman,

you can use RSA % with this formula

%RSA= ((Blanko Absorbances)-(sample absorbances))/Blanko Absorbances x100%

thanks all of you.

DPPH scavenging effect %= AD-AS/ AD *100

Where AD is absorbance value at 517 nm of the methanolic solution of DPPH and AS is the absorbance value at 517 for the sample

Thanks

Hi everyone!

About IC50, is it true that higher IC50 means worse antioxidant activity?

I mean, I prepare coprecipitate of betacarotene and PVP. My coprecipitates have a IC50 lower than BC alone. Is it possible? 

Alessia Di Capua, YES. it is all about how much antioxydant (in the extract) is needed to destroy 50% DPPH. When the IC50 is high, the antioxydant effect is low, because the extract is poor in antioxydant, this is why we need to increase the concentration of the extract. In contrast, when the concentration of the extract needed to destroy 50% of DPPH is low, it means that the extract is rich in antioxydant (no need for high concentration of extract)

Hope that helped you

% scavanging activity = Absorbance of control - absorbance of test sample/Absorbance of control X 100

DPPH radical scavenging activity (%) =[(Ax-Ao)/Ao] *100

where Ax and Ao are respectively the absorbance of extract with DPPH solution and the absorbance of DPPH solution without extract (control) at 517 nm.

DPPH clearance (%) = 1- (Ap-Ac)/Amax-A0) * 100

when A0 represents the absorbance value of the zero removal group; Amax represents the absorbance value of the blank group; Ap represents the absorbance value of the sample group; Ac represents the absorbance value of the control group.

Dear All,

I calculated my DPPH scavening activity based on that formula and my results were negative (even for the standard solutions based on asorbic asid).

I prepared blank samples as follow:

4 replications as 0.05 ml of H20 and 1.95 ml of methanol

4 replications as 0.05 ml of H20 and 1.95 ml of DPPS

(Because I foud as well different problems with that).

Anyway, for both of the trial my scavening activity in all cases were negative.

What could be the problem for that? Thank you.

aa. hi .i have fruit extract ... how to prepare different aliquots of concentrations for DPPH assay

It depends on the concentrations you're working on. You can prepare a stock solution (e.g 100 mg of crude extract in 2 mL of methanol which gives you a stock conc. of 50 mg/mL). Then, from your stock prepare your working concentrations using two-fold serial dilution.