Tannins

By

Dr. Ahmed Metwaly

Objectives:

1. Definition

2. Classification

3. General characters

4. Distribution and physiological role

5. Tests for identification

6. Quantitative determination

7. Medicinal and biological properties

8. Economic importance

Definition;

• They are high molecular weight phenolic compounds

capable of precipitation of animal proteins in hides and

converting them into leather. “Tanning Industry”

.

They are condensation products of Gallic acid or

Flavan-3-ol as well as their glycosides.

Gallic acid Flavan-3-ol

True Tannins

(High molecular weight compounds)

Hydrolysable Tannins

(Pyrogallol)

Condensed Tannins

(Catechol)

Pseudotannins

Low molecular weight compounds

e.g. Gallic acid, Flavan-3,4-diol

Gallitannins Ellagitannins

Hexahydroxydiphenic acidGallic acid

Flavan-3-ol

Complex

tannins

True tannins

These are complex phenolic compounds of high

molecular weights ranging from about 1000 to 5000.

They display the general properties of tannins and

precipitated by gelatin in 1% solution.

True tannins can be sub-classified into hydrolysable

tannins, condensed tannins and complex tannins.

Hydrolysable tannins (pyrogallol tannins)

These may be hydrolyzed by acids or enzymes such as

tannase.

They are formed from several molecules of phenolic

acids such as gallic and hexahydroxydiphenic acids

which are united by ester linkages to a central sugar

(mainly glucose) molecule.

Like gallic acid their solutions turn blue with iron salts.

They were formerly known as pyrogallol tannins, because

on dry distillation gallic acid and similar components are

converted into pyrogallol.

Two principal types of hydrolysable tannins are

gallitannins and ellagitannins which are, respectively,

composed of gallic acid and hexahydroxydiphenic

acid units.

Tannaseis an enzyme that catalyzes the chemical reaction

digallate + H2O 2 gallate

This enzyme belongs to the family of hydrolases, specifically

those acting on carboxylic ester bonds. The systematic name

of this enzyme class is tannin acylhydrolase. Other names in

common use include tannase S, and tannin acetylhydrolase.

Tannase is a key enzyme in the degradation of gallotannins.

It is present in a diverse group of microorganisms,

including rumen bacteria

The difference between Ellagitannins and gallotannins

that; galloyl groups are linked through C-C bonds in ellagitannins,

whereas the galloyl groups in gallotannins are linked by depside

bonds.

A depside is a type of polyphenolic compound composed of two or

more monocyclic aromatic units linked by an ester bond.

Ellagic acid (the depside of Hexahydroxydiphenic acid) can arise

by lactonization during chemical hydrolysis of the tannin; thus, the

term ellagitannin is a misnomer.

C-glucosidic ellagitannins are common in a number of families

including the Myrtaceae, Hamamelidaceae, Punicaceae and

Rosaceae.

Ellagic acidHexahydroxydiphenic acid

Tannic acid 1,2,3,4,6-Pentagalloyl glucose

Grandinin

Casuarictin

raspberry ellagitannin

Condensed tannins

(Catechol) (proanthocyanidins)

Unlike hydrolysable tannins, these are not readily

hydrolyzable to simpler molecules and they do not contain

a sugar moiety.

They are related to flavonoid pigments and have

polymeric flavan-3-ol structures (usually di- and trimers).

• On dry distillation they yield catechol and these

tannins are therefore sometimes called catechol

tannins.

• Like catechol itself, their solutions turn to green with

ferric chloride.

Catechins which also occur with the tannins and flavan-

3,4-diols (leucoanthocyanidins) are intermediates in the

biosynthesis of the polymeric molecules.

Stereochemical variations add to the variety of possible

structures.

On treatment with acids or enzymes condensed tannins

are converted into red insoluble compounds known as

phlobaphenes.

Phlobaphenes give the characteristic red color to many

drugs such as red cinchona bark, which contain these

phlobatannins and their decomposition products.

Catechin Epicatechin

Schematic representation of a

condensed tannin molecule.

Condensed tannins can be linear (with

4→8 bounds) or branched (with 4→6

bounds)

Complex tannins

These represent a group of tannins that are

biosynthesized from both hydrolysable tannin (mostly

a C-glucoside ellagitannin) (flavono-ellagitannin)and

condensed tannin.

Acutissimin A

Pseudotannins

Pseudotannins are polyphenolic compounds of lower molecular

weight than true tannins.

They do not respond to the goldbeater's skin test and may,

under certain conditions, e.g. in concentrated solutions, give

precipitates with gelatin and be partly retained by hide powder.

Examples are gallic acid, catechins and chlorogenic acid.

Chlorogenic acidCatechinGallic acid

Distribution

Type of tannin Plant name and organ

Hydrolysable tannins

Gallitannins

Ellagitannins

Rhubarb rhizome, clove buds, red rose petals, galls, hamamelis leaves and

chestnut bark.

Pomegranate rind and bark, eucalyptus leaves, myrobolans and oak bark.

Condensed tannins Barks: e.g. cinnamon, hamamelis and cinchona.

Roots and rhizomes: e.g. krameria and male fern.

Flowers: e.g. lime and hawthorn.

Seeds: e.g. cocoa, kola and areca.

Leaves: e.g. hamamelis, hawthorn and tea, especially green tea.

Extracts and dried juices: catechu and Indian kino.

Complex tannins Members of Fam. Combretaceae, Myrtaceae and Fagaceae.

Pseudotannins

Gallic acid

Catechins

Chlorogenic acid

Ipecacuanhic acid

Galls, rhubarb and most drugs that contain gallitannins.

Catechu, cocoa and most drugs containing condensed tannins.

Coffee particularly when unroasted.

Ipecacuanha.

Major sources of commercial tannins, for use in leather industry, are

obtained from chestnut bark trees.

Pharmaceutical tannin is prepared from oak galls.

Chestnut bark

Castalagin

Oak galls

Tannic acid

Physiological role

They exert an inhibitory effect on many enzymes due to protein

precipitation and, hence, may contribute a protective function in barks

and heartwoods.

• They play an important role in the mechanism of hydrogen transfer in

plant cells, probably due to their high affinity for oxygen.

• Certain drugs contain only one type of tannin. Others may contain

more than one type e.g. tea, hamamelis leaves and bark contain both

hydrolysable and condensed tannins.

Physical properties

Tannins are non-crystallizable compounds.

They are soluble in water forming colloidal solutions

with acidic reaction and sharp astringent taste.

They are soluble in dilute alkalis, alcohol, glycerol and

acetone, but only sparingly soluble in other organic

solvents.

Their solutions precipitate heavy metals, alkaloids,

glycosides and protein (e.g. gelatin).

Tests for identification

• Goldbeater's skin test (Skin tanning test):

Goldbeater's skin is a membrane prepared from the intestine of the ox

and behaves similar to an untanned hide.

Soak a small piece of goldbeater's skin in 25% HCl; rinse with distilled

water and place in the solution to be tested for 5 min. Wash with distilled

water and transfer to a 15 % solution of ferrous sulphate.

A brown or black color on the skin denotes the presence of true

tannins.

• Gelatin test:

Solutions of true tannins (about 0.5-1%) precipitate a 1% solution of

gelatin containing 10% sodium chloride.

Gallic acid and other pseudotannins may react in concentrated

solutions.

• Test with phenazone:

Add 0.5 g of sodium acid phosphate to about 5 ml of an aqueous extract

of the drug; warm, cool and filter. To the filtrate add 2 % phenazone

solution.

All tannins are precipitated, the precipitate being bulky and often

colored.

• Test with ferric chloride:

Add 5 % ferric chloride solution drop by drop to 2-3 ml of the extract and

observe the color produced.

Hydrolysable tannins (gallitannins and ellagitannins) give bluish-black color

or precipitate and condensed tannins brownish-green ones.

If the test is carried on an extract that contains both types of tannins, a blue

color is produced which changes to olive-green as more ferric chloride is

added.

• Test with bromine water:

Add bromine water to an aqueous extract of the drug.

- Condensed tannins gives a buff-colored precipitate

- Hydrolysable tannins do not form any precipitate.

• Test for catechin:

Catechins on heating with acids form phloroglucinol and they can be

detected by a modification of the well-known test for lignin.

Dip a matchstick in the plant extract containing catechin, dry, moisten with

concentrated HCl and warm near a flame a pink or red is produced.

• Test for chlorogenic acid:

Add aqueous ammonia to an extract containing chlorogenic acid, expose to

air a green color gradually develops.

Quantitative determination

1. Hide powder method:

This is the most widely used on commercial scale.

A known amount of a reference sample of hide powder is added to the

drug extract.

The difference in the dry weight of the extract before and after treatment

with hide powder is a measure for the tannin content.

1. Copper acetate method:

Tannin is precipitated quantitatively with copper acetate solution.

The precipitate of copper tannate is collected on an ashless filter paper,

washed with water till free from copper ions and ignited.

The residue of copper oxide obtained is completely oxidized by adding a

few drops of nitric acid.

Tannin content is calculated on the basis that 1 part of copper oxide is

equivalent to 1.305 parts of tannins.

• Agglutination method

This depends on that a suspension of red blood cells is agglutinated

with tannins, but haemolysis does not occur (c.f. saponins). The

erythrocytes are agglutinated (stick together) due to formation of a sticky

membrane on their surface in contact with tannins.

The quantity of red blood cells precipitated is proportional to the

amount of "astringent" tannin present.

The end point is reached when red blood cells are completely

precipitated i.e. the solution is colorless on shaking. Reaction is stopped

when the filtrate gives no precipitate with ferric chloride (this indicates

absence of tannins).

The astringency value of the extract is compared with that of tannic acid

used as reference standard, recorded as percentage tannic acid and

referred to as "tannin number".

All the solutions used should be isotonic.

Catechin is inactive.

• Iodometric determination

Tannins and pseudotannins quantitatively consume iodine from alkaline

medium due to their phenolic nature.

Excess iodine is determined by titration, after acidification, with standard

sodium thiosulphate solution using starch as indicator.

True tannins can be removed from the extract by precipitation with gelatin.

This can permit the determination of each group of constituents alone.

Medicinal and biological properties

Tannin-containing drugs precipitate proteins and have been traditionally

used as styptics (stop hemorrhage) and internally for the protection of

inflamed surfaces of mouth and throat.

They play an important role in the treatment of burns. They form a mild

antiseptic protective layer on the surface of the injured skin below which

regeneration of new tissue takes place.

They act as anti-diarrheals, although not recommended in this respect as

they usually delay elimination of bacterial toxins from the body.

Tannins have been employed as antidote in poisoning by heavy metals,

alkaloids and certain glycosides due to their precipitation as

tannates.

Recently tannins as most polyphenols were proved to have a potent

antioxidant effect.

Studies on the antitumor effect of tannins proved that a strong

activity is obtained with ellagitannins having galloyl groups at the O-

2 and O-3 positions of the glucose core(s), as in the

tellimagrandins.

Certain tannins were proved to have anti-HIV activities

Tellimagrandin II (Anti herpes virus)

Economic importance

Tannins are used in leather industry to transform raw animal skin to

leather due to their ability to cross-link with proteins. To be effective for

tannage, the polyphenol molecule must be neither too large as it will

be unable to enter the interstices between the collagen fibrils of the

animal skin nor too small as it will be unable to cross-link between the

protein molecules of adjacent fibrils at several points.

Tannins are used in the manufacture of inks based on the dark colored

complexes they form with iron salts.

Tannins are used in food industries to improve acceptance of many

beverages and foods when a degree of astringency is required.

Summary:

1. Definition

2. Classification

3. General characters

4. Distribution and physiological role

5. Tests for identification

6. Quantitative determination

7. Medicinal and biological properties

8. Economic importance