The answer to your question is 'NO'.

Typical phenolics that possess antioxidant activity are known to be mainly phenolic acids and flavonoids. Phenolic acids are a major class of phenolic compounds, widely occurring in the plant kingdom especially in fruits and vegetables. Because of the diversity and complexity of the natural mixtures of phenolic compounds in hundreds of herb extracts, it is rather difficult to characterize every compound and elucidate its structure, but it is not difficult to identify major groups and important aglycones of phenolic compounds. Many medical herbs and spices have been studied and to some extent their phenolic chemistry is known. The contents of phenolic compounds, measured by HPLC after enzymatic hydrolysis, were found to be different from those values measured using the Folin–Ciocalteu method. The amount of polyphenols was also dependent on the extraction method. The samples for HPLC when subjected to enzymatic hydrolysis which, in contrast to methanol extraction, resulted in specific disruption of linkages and deglycosylation of phenolic compounds. Many authors report that aglycones exhibit higher antioxidant activity than do glycosides.

It is obvious that the total phenolic content measured by the Folin–Ciocalteu procedure does not give a full picture of the quality or quantity of the phenolic constituents in the extracts.

Pl. see the following papers for more details:

Katsube, T., Tabata, H., Ohta, Y., Yamasaki, Y., Anuurad, E., Shiwaku, K., et al. (2004). Screening for antioxidant activity in edible plant products: Comparison of low-density lipoprotein oxidation assay, DPPH radical scavenging assay, and Folin–Ciocalteu assay. Journal of the Agricultural and Food Chemistry, 52, 2391–2396.

Wu, X., Beecher, G. R., Holden, J. M., Haytowitz, D. B., Gebhardt, S. E., & Prior, R. L. (2004). Lipophilic and hydrophilic antioxidant capacities of common foods in the United States. Journal of the Agricultural and Food Chemistry, 52, 4026–4037.

Regards

Sometimes it is true but not all the time, as flvanoids may not be phenolics all the time.

What does the meaning of "Total Phenol"? Does it refer to all of the compounds(Secondary metabolites) which are produced by Shikimic acid pathway or Malonic acid pathway and contain at least 1 phenol moiety(Flavonods, Tannins, Isoflavones, etc)? Or it refers only to Phenolic acids?

I am in agreement with Neeraj Sethiya. They are also found as alkaloids and glycosides. So not necessary that the equation is equal in terms of concentration, it may vary depending on the utilization of the molecule in the complex phytochemical factory of the plant.

Dear Dr Nahata,

let me emphasis on the word "Total". Can we exactly assess all of the secondary metabolites with Phenolic moieties by Folin- Ciocalteu reagent method? Does this method assesses all of such compounds( I mean Flavonoids, Isofavons, Anthocyanins, Tannins, etc) or only Phenolic acids(free or conjugated)? Thx

The answer to your question is 'NO'.

Typical phenolics that possess antioxidant activity are known to be mainly phenolic acids and flavonoids. Phenolic acids are a major class of phenolic compounds, widely occurring in the plant kingdom especially in fruits and vegetables. Because of the diversity and complexity of the natural mixtures of phenolic compounds in hundreds of herb extracts, it is rather difficult to characterize every compound and elucidate its structure, but it is not difficult to identify major groups and important aglycones of phenolic compounds. Many medical herbs and spices have been studied and to some extent their phenolic chemistry is known. The contents of phenolic compounds, measured by HPLC after enzymatic hydrolysis, were found to be different from those values measured using the Folin–Ciocalteu method. The amount of polyphenols was also dependent on the extraction method. The samples for HPLC when subjected to enzymatic hydrolysis which, in contrast to methanol extraction, resulted in specific disruption of linkages and deglycosylation of phenolic compounds. Many authors report that aglycones exhibit higher antioxidant activity than do glycosides.

It is obvious that the total phenolic content measured by the Folin–Ciocalteu procedure does not give a full picture of the quality or quantity of the phenolic constituents in the extracts.

Pl. see the following papers for more details:

Katsube, T., Tabata, H., Ohta, Y., Yamasaki, Y., Anuurad, E., Shiwaku, K., et al. (2004). Screening for antioxidant activity in edible plant products: Comparison of low-density lipoprotein oxidation assay, DPPH radical scavenging assay, and Folin–Ciocalteu assay. Journal of the Agricultural and Food Chemistry, 52, 2391–2396.

Wu, X., Beecher, G. R., Holden, J. M., Haytowitz, D. B., Gebhardt, S. E., & Prior, R. L. (2004). Lipophilic and hydrophilic antioxidant capacities of common foods in the United States. Journal of the Agricultural and Food Chemistry, 52, 4026–4037.

Regards

Dear Pakzad

The phenolics are all those compounds which contains phenolic group not -hydroxy. It may be mono-phenol, di-phenol, tri-phenol or polyphenol. These phenolics can be estimated by Folin- Ciocalteu reagent assay. This method gives a perspective about phenolic content in the sample. Though as Dr Nahata explained all phenolics could not be estimated by single extraction or by any single method due to the complexity of compounds.

While flavanoids are those compounds that have basic carbon skelton of C6-C3-C6.

Majority of flavanoids are phenolics that means they contains atleast single phenolic gp.

So theoretically it is true that during quantification of phenolics and flavanoids the value of phenolics should be more than flavanoids. But practically there are other situations such as which reference standard you are using for determination of phenolics and flavanoids and the occurrence of other compounds in the sample, extraction solvent etc.

Folin-Ciocalteu (F-C) reagent is not only reactive to phenolic compounds. Non-phenolic compounds too react with F-C reagent, for e.g., Fe(2+), Mn(2+), SO3(2-) (Everett et al., J. Agric. Food Chem. 2010). Further, nitrogen-containing compounds such as hydroxylamine, hydrazine, aromatic amines are also known to respond to F-C reagent (Ikawa et al., J. Agric. Food Chem., 2003). In fact, the F-C reagent was first developed to measure tyrosine and tryptophan, where tryptophan is a non-phenolic compound (J. Biol. Chem. 1927). So, to me, it is surprising to note from your post that determination of flavonoids (by AlCl3) using quercetin as standard yielding higher value than total-polyphenol estimation by F-C assay. Yes, flavonoids constitute a subclass of polyphenols.

Please refer Sim

The term phenol or phenolics stand for a group encompassing different set of compounds including phenolic acids, phenolic aldehydes, acetophenones, tyrosine derivatives, phenylacetic acids, hydroxycinnamic acids, coumarins, isocoumarins phenylpropenes, chromones, naphthoquinones, xanthonoids, stilbenoids, anthraquinones, chalconoids, flavonoids, isoflavonoids, neoflavonoids, lignans, neolignans, biflavonoids, lignins, catechol, melanins, flavolans (condensed tannins), polyphenolic proteins, other polyphenols, etc widely distributed in the plant kingdom. The TPC measured by the Folin–Ciocalteu procedure and presented as GAE equivalent does not give a complete account (neither qualitative nor quantitative) of the phenolic constituents in the plant extracts. Since flavonoids constitute a part of the total phenolics, their quantity cannot be greater than the total phenolics, if estimation of both TPC and flavonoids in a given plant extract is done appropriately and accurately.

Dear Sir

You`re right, the largest natural phenols are the flavonoids, but there is a common mistake in some articles. The noted method is not an appropriate procedure for estimation of “Total phenolic compounds”.  

Approximately, phenolic acids are belong to the simplest forms of phenolic compounds, which are classified to three categories. As one of the phenolic acids categories, Trihydroxybenzoic acids can be evaluated through Folin-Ciocalteu reagent and Gallic acid. 

According to my knowledge, even polyphenols are higher than phenolic acids in the most plant species.

What percent does Coumarins comprise among total phenolics in Herbs?

Dear all,

Greetings!

"HAVE A NICE WEEK"

I am Dr. Ravichandran, I have isolated two compounds from plants through column. These two compounds were obtained in Chloroform fraction. Both gives positive results for flavonoids tests in preliminary phytochemical screening and in spraying reagents for favonoids. But both the compounds are not given results for  Folin- Ciocalteu reagent tests. Please help clarify there is any chances are there these two compounds are to be  flavonoids.

Dear Dr. Ravichandran

What kind of preliminary phytochemical screening tests have you done?

What ever the argument may be Total flavonoids can not be more than Total phenols in any food sample. I have done many assays with Folin- Ciocalteu reagent which over estimates becoz of other reducing agents in foods .However total flavonoids can not be more than total phenolics.

Hi, Can anyone guide me please???

There are many protocols in many papers but I wonder where we can find most reliable and easy protocols? I am looking for measuring Total Phenolic conetent, Total Flavonoid Content and Antioxidant properties of leaves and flowers from a plant from Elaeagnaceae Family.

Many thanks in advance.

"Classification of Flavonoids

Flavonoids are the largest family of polyphenolic compounds; that is why the words ”polyphenols” and “flavonoids” sometimes may be used interchangeably. However, although all flavonoids are polyphenols, polyphenols not necessarily are flavonoids. Plants produce flavonoids as a protection against parasites, oxidative injury and harsh climatic conditions. Flavonoids are further divided in several subclasses: anthocyanins, flavanols, flavanones, flavonols, flavones and isoflavones."

Source: http://www.livestrong.com/article/479645-polyphenols-vs-flavonoids/

Flavonoids are synthesised in one of the secondary metabolic pathways for phenolics, alkaloids and terpenoids (http://www.genome.jp/kegg/pathway/map/map00400.html), i.e. the phenyl propanoid pathway (http://www.genome.jp/kegg-bin/show_pathway?map00940), the flavonoid biosynthetic pathway (http://www.genome.jp/kegg-bin/show_pathway?map00941), and the Flavone and Flavonol biosynthesys (http://www.genome.jp/kegg-bin/show_pathway?map00944). E.g quercetin is a flavonol, and widely seen as an important antioxidant compound in food and red wine.

Flavonoids can be measured by High Performance Liquid Chromatography (HPLC) with rutin as a standard, and there are many protocols published.

See also

Benetis, R., J. Radusiene, et al. (2008). "Development of an RP-HPLC method for the analysis of phenolic compounds in Achillea millefolium L." Journal of Liquid Chromatography & Related Technologies 31(4): 596-610.

Winkel, B. S. J. (2006). The biosynthesis of flavonoids. The science of flavonoids. E. Grotewold, Springer: 71-95

Dear Pery Prev

Find them in one of my articles such as:

Essential Oil Composition and Antioxidant Activity of the Methanol Extracts of Three Phlomis Species from Iran

https://www.researchgate.net/publication/264161901_Essential_Oil_Composition_and_Antioxidant_Activity_of_the_Methanol_Extracts_of_Three_Phlomis_Species_from_Iran?ev=prf_pub

Article Essential Oil Composition and Antioxidant Activity of the Me...

Very interesting duscussion. I am currently working on estimation of free phenolic acid extraction by acidic hydrolysis, folowed by EtOAc extraction. Could the residue from this method (presumably free phenolic acids) be considered as total phenol? As my attempt on extraction using 80% MeOH aq & vacuum evap. did not yield dry residue to reference against GA equivalent.

Still the question remain as such. After all discussion,  I also  think that total phenolics should be always greater than total flavonoids.

To determine total phenolic acids is not the same as "total phenolic compound". If Folin- Ciocalteu reagent and gallic acid will only determine the phenolic acids.

There is no definite relationship between total Phenol & total Flavonoids.

there is no general relationship between total Phenol and total Flavonoids but it is well known that total phenolics must be always greater than total flavonoids.

flavonoids part of polyphenols

polyphenols As (vanillin, catechin, gallic acid...)

flavonoids As (reutin, quercetin)

Very interesting discussion. Can anyone tell the quercetin equivalent of rutin so that results obtained using quercetin and rutin standards can be compared?

Rutin is a glycoside combining the flavonol quercetin and the disaccharide rutinose. Therefore Rutin can be used as a standard to calculate the amount of quercetin glycoside in a plant sample by HPLC.

I guess it all depends on the functional group that the Folin-Ciocalteau reagent actually responds to. It should be recalled that the reagent also measures proteins where it targets tyrosine, an amino acid that has the phenolic structure.

there is no relationship between these groups

I measured flavonoids and phenols for all parts of the eggplant, and I did not find any clear association with the vegetative and flowering growth stage.

Article Chemical Composition and Antioxidant Capacity of Eggplant Pa...

Article POLYPHENOLIC AND FLAVONOIDS CONTENT, HPLC PROFILING AND ANTI...