For determining Total phenolic content, FC method can be used as following:

1 ml of sample+ 5ml of 10% (v/v) FC reagent + 4ml of 7.5% Sodium carbonate--60 min incubation at room temperature---measure at 765nm, zero on reagent blank.

Construct a calibration curve using Gallic acid as standard. 

Get the absorbance values (Ab) and divide them by a (where 'a' is the value that you get from the standard curve). This (Ab/a) will give you the ug GAE/ml of the extract. Then calculate the Ab/a X dilution factor of your extract. Further, divide this value with 1000 to get mg GAE/ml of the extract.

The results can be expressed as mg of Gallic Acid Equivalents (GAE) per ml of extract (mg GAE/ml) or mg GAE/g of the dry weight of the sample.

Hope that helps.

Regards,

Deep Jyoti Bhuyan

For determining Total phenolic content, FC method can be used as following:

1 ml of sample+ 5ml of 10% (v/v) FC reagent + 4ml of 7.5% Sodium carbonate--60 min incubation at room temperature---measure at 765nm, zero on reagent blank.

Construct a calibration curve using Gallic acid as standard. 

Get the absorbance values (Ab) and divide them by a (where 'a' is the value that you get from the standard curve). This (Ab/a) will give you the ug GAE/ml of the extract. Then calculate the Ab/a X dilution factor of your extract. Further, divide this value with 1000 to get mg GAE/ml of the extract.

The results can be expressed as mg of Gallic Acid Equivalents (GAE) per ml of extract (mg GAE/ml) or mg GAE/g of the dry weight of the sample.

Hope that helps.

Regards,

Deep Jyoti Bhuyan

Estimation of total phenolic content (TPC)

 Phenols, referring to compounds containing a hydroxylated benzene ring, are an important component in most green leafy vegetables.  The analysis of these compounds is a rather complex task due to the number of different phenolic components. Total phenol content in a sample can be measured by Folin-Ciocalteu method.  

Folin-Ciocalteu method involves measuring the number of -OH groups present in a sample, assuming that light absorption increases when more -OH groups are present. The Folin-Ciocalteu assay depends on an oxidation-reduction reaction in which Folin-Ciocalteu reagent, a mixture of molybdenum (Mo) and tungsten (W) salts, is added to a sample containing phenols.  The Mo (VI) and W (VI) are reduced in the presence of the oxidizable phenol groups resulting in a colour change from yellow to blue.

The intensity of the blue colour can then be determined using absorption spectroscopy at 740 nm (Gupta and Prakash, 2009). The intensity of light absorption at wavelength is proportional to the concentration of phenols (AOAC, 1995).

 Reagents

 (i)    Folin Ciocalteau Phenol Reagent (10 fold dilution)

Folin–Ciocalteau reagent (10 ml) was diluted with 90 ml of distilled water in a 100 ml volumetric flask.

     (ii) 4:6 water/methanol

Distilled water (40 ml) was mixed with 60 ml of methanol in a 100 ml volumetric flask.

 I think this may be also another way to estimate the TPC.

Hope this is also help you

Regards

Umaramani.

     (iii) 2% Sodium carbonate

Na2CO3 (2 g) was dissolved in 100 ml of distilled water and boiled. Then it was filtered through Whatman No.41 filter paper and kept in a refrigerator.

(iv) Preparation of tannic acid

Dry tannic acid (0.5 g) was dissolved in 10 ml methanol. The volume was made to 100 ml with 4:6 water/methanol as the solvent. This tannic acid stock solution was kept in a refrigerator until use.

 Different volumes of tannic acid stock solution (0, 1, 2, 3, 5 and 10 ml) were made to 100 ml with distilled water to prepare different phenol concentrations (0, 50, 100, 150, 250 and 500 mg/l respectively) to prepare the calibration curve.

 Sample preparation

Samples of GLVs (1 g of each), in 30 ml of methanol were ground using a mortar and pestle. After recovery of the finely ground mixture, 15 ml methanol was used to wash the mortar and pestle and then pooled with the first mixture. The mixture was double filtered at room temperature. Supernatants were filtered using Whatman No.42 filter paper and the volume was made up to 50 ml with methanol. The extracts were prepared in triplicate and all analysis was carried out in triplicates.

 Assay

TPC was determined by adopting the procedures of Gupta and Prakash (2009), Karakaya (2004) and Matthaus (2002). But slight modification was done for convenience and availability of analytical instruments (Motor and pestle was used instead of blender and double filtration was performed to get clear supernatant instead of using centrifuge).

A sample of methanolic extract (0.2 ml) was mixed with 1 ml of Folin–Ciocalteau reagent (tenfold dilution) and 0.8 ml of 2% Na2CO3. The volume was made to 10 ml with 4:6 water/methanol as the solvent. This was allowed to stand for 30 minutes and absorbance read at 740 nm using a specrotphotometer (Hach DR 2500). Concentration was calculated using tannic acid as standard and the results were expressed as mg tannic acid /100 g wet weight (WW).

Tannic acid, in varying concentrations (0, 50, 100, 150, 250 and 500 mg/l), was used to prepare a standard curve.  This curve is used to relate the absorbance of the unknown samples to Tannic acid equivalents (TAE).  

Please go through this publication

Article Evaluation of radical scavenging potential and total phenol ...

In fact, FC method is widely used to determine phenolic compounds contente. However, it has several interfering compounds (virtually, any compound able to reduce Mo-VI), often conducting to overestimating results. If this is not a problem for you, just follow any of the suggested publications. However, if you need more accurate results, you should perform a chromatographic analysis coupled to mass spectrometry, for instance HPLC-DAD/MS.

For your guidance:

http://ac.els-cdn.com/S0308814614006906/1-s2.0-S0308814614006906-main.pdf?_tid=a61f540a-0518-11e5-8ad5-00000aacb362&acdnat=1432804093_1c2f62d7a26e48dda0b19a0f0054e792

The following paper will help you

Check this publication:

use the equation C (GAE) = c x V/M (where c = concentration determined from standard curve (mg/ml), V = volume used during the assay (mL), and M = mass of the extract used during the assay (g) and will give GAE in mg/g extract.

"The total phenolic concentration in  extracts was determined according to the Folin-Ciocalteu method using gallic acid as the standard."

Please see this article

http://faculty.missouri.edu/~glaserr/3700s14/Antioxidant_ORAC_et_al_jf803011r.pdf

 Also see this research and follow the same steps