discoloration is most likely caused by iron and/or manganese and not tannins,
but if the intensity of the colour remains intact, it is most likely caused by
tannins (Franzmann et al. 2001).
3. Ferric chloride test: In two millilitres (2 mL) of the hot aqueous solution of
tannin extract, few drops of 10% Ferric chloride solution is added. The
occurrence of blackish blue colour showed the presence of gallotannins and a
green-blackish colour indicated presence of catechol tannins (Auwal et al.
2014) and also the occurrence of greenish brown colour indicates the presence
of condensed tannins (Pizzolato 1977).
4. Alkaline reagent test: To 1 ml of the plant extract, a few drops of 10% Sodium
hydroxide (NaOH) solution was added. An instant appearance of yellow to red
precipitate indicates the presence of tannins (Bharudin et al. 2013).
5. Phenazone test: In a 5 ml aqueous extract of tannin, sodium acid phosphate (0.5
gm.) is added and the mixture is heated, cooled and filtered. A solution of 2%
phenazone is added to the filtrate. A bulky coloured precipitate is formed
(Alotaibi 2019).
6. Match stick test (Catechin test): Match stick test is specially done for detection
of catechins. The principle of this qualitative test based upon simple reaction of
catechins, which can produce phloroglucinol in the presence of acid. A match
stick (which contains lignin) is dipped in aqueous extract of tannin (catechins),
dried and moistened with concentrated hydrochloric acid and is warmed in the
flame. Catechins in the presence of HCl produces phloroglucinol which stains
the lignin (present in match stick) produces pink or red colour (Thiruchenduran
et al. 2017).
7. Gambir-Fluorescein test: It is often performed to identify the pale catechu. The
alcoholic extract of pale catechu is mixed with sodium hydroxide solution, and
petroleum ether. After the mixing, the whole mixture is shaken and kept aside
for few minutes. The petroleum ether layer show green fluorescence due to the
presence of fluorescein present in pale catechu. Black catechu gives negative
result in this test (Ray et al. 2006).
8. Bromine water test: In an aqueous extract of condensed tannins, the addition of
acetic acid followed by bromine water gives a result of buff coloured precipitate. Hydrolysable tannin doesn’t response to this test (El Sissi and El
Sherbeiny 1967).
9. Lead sub-acetate test: In the ethanolic extract of tannin, addition of acetic acid
and lead sub-acetate solution results in gelatinous cream coloured precipitate
(Ukoha et al. 2011).
10. Chlorogenic acid test: When an extract of chlorogenic acid containing sample is
treated with aqueous ammonia. A green colour is formed on exposure to air
(Ashok and Upadhyaya 2012).
11. Vanillin-hydrochloric acid test: This test is performed by taking alcoholic
extract of sample, vanillin alcohol (vanillin 1 gm, alcohol 10 ml) and concentrated hydrochloric acid in a ratio of 1:10:10. The presence of tannin is
confirmed by a pink or red colour formation due to phloroglucinol (Rangra
et al. 2019).
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S. Mal and D. Pal
but if the intensity of the colour remains intact, it is most likely caused by
tannins (Franzmann et al. 2001).
3. Ferric chloride test: In two millilitres (2 mL) of the hot aqueous solution of
tannin extract, few drops of 10% Ferric chloride solution is added. The
occurrence of blackish blue colour showed the presence of gallotannins and a
green-blackish colour indicated presence of catechol tannins (Auwal et al.
2014) and also the occurrence of greenish brown colour indicates the presence
of condensed tannins (Pizzolato 1977).
4. Alkaline reagent test: To 1 ml of the plant extract, a few drops of 10% Sodium
hydroxide (NaOH) solution was added. An instant appearance of yellow to red
precipitate indicates the presence of tannins (Bharudin et al. 2013).
5. Phenazone test: In a 5 ml aqueous extract of tannin, sodium acid phosphate (0.5
gm.) is added and the mixture is heated, cooled and filtered. A solution of 2%
phenazone is added to the filtrate. A bulky coloured precipitate is formed
(Alotaibi 2019).
6. Match stick test (Catechin test): Match stick test is specially done for detection
of catechins. The principle of this qualitative test based upon simple reaction of
catechins, which can produce phloroglucinol in the presence of acid. A match
stick (which contains lignin) is dipped in aqueous extract of tannin (catechins),
dried and moistened with concentrated hydrochloric acid and is warmed in the
flame. Catechins in the presence of HCl produces phloroglucinol which stains
the lignin (present in match stick) produces pink or red colour (Thiruchenduran
et al. 2017).
7. Gambir-Fluorescein test: It is often performed to identify the pale catechu. The
alcoholic extract of pale catechu is mixed with sodium hydroxide solution, and
petroleum ether. After the mixing, the whole mixture is shaken and kept aside
for few minutes. The petroleum ether layer show green fluorescence due to the
presence of fluorescein present in pale catechu. Black catechu gives negative
result in this test (Ray et al. 2006).
8. Bromine water test: In an aqueous extract of condensed tannins, the addition of
acetic acid followed by bromine water gives a result of buff coloured precipitate. Hydrolysable tannin doesn’t response to this test (El Sissi and El
Sherbeiny 1967).
9. Lead sub-acetate test: In the ethanolic extract of tannin, addition of acetic acid
and lead sub-acetate solution results in gelatinous cream coloured precipitate
(Ukoha et al. 2011).
10. Chlorogenic acid test: When an extract of chlorogenic acid containing sample is
treated with aqueous ammonia. A green colour is formed on exposure to air
(Ashok and Upadhyaya 2012).
11. Vanillin-hydrochloric acid test: This test is performed by taking alcoholic
extract of sample, vanillin alcohol (vanillin 1 gm, alcohol 10 ml) and concentrated hydrochloric acid in a ratio of 1:10:10. The presence of tannin is
confirmed by a pink or red colour formation due to phloroglucinol (Rangra
et al. 2019).
734
S. Mal and D. Pal