International Journal of Physical Science, Volume 6, Number 1, 2011

Printed in Nigeria. All Right Reserved ISSN: 2006 -1064 Copyright©2QUDuncanScienceCompany

EXTRACTION OF INDIGO DYE (POWDERED, FORM) FROM THE LEAF OF INDIGOFERA

TINCTORIA

Olusola Adeyanju 1, Emmanuel S. E. and Akomolafe S. F.2

'Dept. of Chemistry, University of Jos, Jos, Plateau State, Nigeria
*Dept. of Biochemical and Chemical Sci., University of Science and Tech. Ifaki-Ekiti, Ekiti State, Nigeria

ABSTRACT
Many plants have been identified as potentially rich in natural dye content and some of
them have been used for natural dyeing for quite some time. Normally natural dyes are
extracted from roots, stems, leaves, flowers and fruit of various plant. In living Indigo
plant, form, indigo exist as a colourless substance comprised of sugars and indoxyl.
Indigo plants are harvested and soaked or compostled, and through bacteriological
action (fermentation) which was replaced by the used of an alkali solution (sodium
hydroxide solution of different concentration 2.0m,2,5m, 3.0m and 4.0m) and acidified
hydrogen peroxide the airing, combined with oxygen (oxidation) to form indigo, the
concentrated blue pigment (powdered form). Highest yield of indigo dye (powdered
form) was obtained from using 2.0 sodium hydroxide solution.

Keywords: Indigo Plant, Extraction, Indigofera tinctoria, Dye.

INTRODUCTION
Dye is simply defined as an organic chemical compound which has selective ability to impact colour evenly on
a fairly permanent basis by selectively retaining same of the wave length of the light falling on the
surface without causing any unbearasble damage to the material.(l11
Over the years, there had being a lot problem with natural dyes. These short coming are, poor fastness to
light and washing, inability of the dyes to be obtained in powdered form. Dependence on natural
fermentation and atmospheric oxidation for vat solution, dependence on local ash for reduction, and
prolonged time for the preparation of vat solution are mostly associated with indigo dye which is the oldest,
most abundant and important natural dye in existence, mostly used for jeans dyeing. These set back has head to
the shif6ting of emphasis from natural dye to synthetic dyes, which are believed to make up with most of the
afore mentioned shortcomings. But the cost of producing or synthesizing these dyes make it impossible to look
completely away from the ancient way of obtaining dyestuff.
It was later discovered that a dye consist of two colour producing substance(s). the chromophore (electron
acceptor) and a part to regulate the solubility and dyeing properties the auxochrome (electron donor).
Without both parts, the materials is simply a colourless substance.|4)
Liebeman et all (lD) showed that reducing (hydrogenation) agents almost invariably destroyed the colour of
organic compounds and so concluded that colour was associated with unsaturation. Today colour is still
attributed, among other things, to unsaturation.
The significance of these study is to create awareness in the mind of dyers in of the availability of raw
material for dyeing purposes in Nigeria and that it can be developed or processed to their taste with cost. To
further broaden the research base and scientific progress in Nigeria in such a way as to save the country a lot of
money in foreign exchange and prevent economic depletion owing to importation and improve the
commercial production of this natural dyestuff. This will provide job opportunities for our citizens.
Furthermore, this production method will make a lot of improvement on the usual local me4thod as this
method gives room for the final product to be in refined form thereby easing storage problem and reducing the
time required for the preparation of vat solution, eliminating the process of fermentation of the leaves before
extraction. The need for the addition of local ash for reduction is eliminated so also the problem of inadequate
oxidation.
This research work only covers the extraction, precipitation and concentration of natural dye from the leaves of
the specie of indigo plant dndigot'era tmctona). It does not cover the extraction from other species.

INDIGO PLANT
Indigo has long been regarded as one of the most valuable and imported of all colouring matters. It was in
India and Egypt long before the Christian era. It was introduced into Europe in the sixteen century. The
generic name is derived from the Latin word indicum, indicating that the plants came from India itself. They are
herbs or smell shrubs with pinnate leaves. When these plants are the source of indigotine the dyestuff is
obtained from the leaves.

137
 Extraction of Indigo Dye (Powdered, Form) from the Leaf of Indigofera Tinctoria

The indigo plant is a leguminous, annual, normally busy, erect, usually less than 75cm in height, much
branched with well developed roots and producing numbers of small pods containing round, usually yellow or
green seeds (l ,

THE ROOT SYSTEM

The root system is extensive, with a tap-root which may exceed 1.5m in length rise to many lateral branched in
the 0,03 horizon. Root development of indigo plant in the field indicate three separate phases
corresponding to a specific period of vegetative development.
j. Extensive of the tap-root and shallow horizontally laterals accompany vegetative tap growth.
ii. Root development to 75cm related to flowering and pod formation.
iii. Extensive and deep penetration of lateral roots occurs during seed maturation. Soil temperature affect the
rate of growth and range of 27-32)°c appears to be optimum for quick initial growth. (11) THE STEM
The stern normally round, usually less than 75cm in height , the internodes becoming woody with age.
Plants are naturally much branched, although modern cultivars usually have less than six branches and there are
determinate and indeterminate types.

THE LEAVES
The leaves are alternate, slightly variable in shape, normally trifoliate, the three ovate or laneolate leaflet
borne on a long petiole with shades of brown colour.
Indigo plants leaves have characteristic calvin cycles photosynthesis, and there is initially little variation
between leaf photosynthesis rates until main flowering and pod filling, then new, upper leaves have rates
much higher than previous leaves, although the extent of this difference and the general rate of
photosynthesis is also directly related to the cultivars and environment.(ll)

THE FLOWERS
The flowers are born on short racemes originating in leaf axile in florescence bearing up to twenty, purple or
unite flowers, which are typically leguminous in shape. Self pollination is the rule, pollen being shed just
before or when flowers open and normal out crossing is estimated at 0.5-1.0% cool or cold periods. 15-200c
adversely affect flower development, but have little effect on yield once the seed has been formed.

CULTIVATION
The principal species cultivated for the manufacture of dyes are the indigofera sitmatrana,. indigofera arrela.
indigofera aniL Indigofera tmctoria and others.
The indigo seed is sown about the end of February or beginning of March, it germinate in the course of four to
five days and at the middle of June. The plant can obtained a height between 3 and 5 feet and has a stem of
about a quarter of an inch in diameter as the plant grows. The colouring principle gradually increases until it
reaches a maximum about the end of August.
When the paste of indigo is agitated with alkali sodium hydrosulphite (Na2S2Od), a reducing agen( in large vats,
the insolution indigo reduced to the soluble "leuco compounds, indigo white", C-OH which then reacts with
alkali toform the water soluble enolate C-ONa. Cotton and other textile fibres haved affinity for this solution,
which is dark yellow-green or at times orange brown. When exposed to the atmosphere or treated with an
oxidant such as hydrogen peroxide (H 2O2) in acid solution, the leuco compound is oxidized to the original
insoluble vat-dye coloured dark blue. C=C.ONa + H2Q-*. C-C.OH + NaOH

CHEMISTRY OF INDIGO
The clouring matter in the indigo (indigofera linctoria) exists naturally as glycoside known as indicant
(C14H1206N).

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 International Journal of Physical Science, Volume 6, Number 1, 2011

C-O-CH-(CHOH)3.CHCH2OH

Indican is a combination of glucose and indoxyl. When this glucoside is decomposed either by enzymic
action during fermentation or by dilute acides, it decomposes by hydrolysis to give indoxyl and glucose.
Hydrolysis of indicant is carried out by crushing the plants in water when the enzymes B-glycosidase
catalyses the hydrolysis of the glocuses(6)

C14HI206N + H20 C8H7ON + C6H12O6

Indican water indoxyl glucose

139
Atmospheric oxidation(oxygen) converts indoxyl to dark blue indigo, which is also known as indigotin

CH + O:

o
2H2O
 c
i.e 2 8 H2 oN + o 2 C6 H10 o2N2 + 2H2 o
Natural indigo contains a red isomer of indigo known as indirttbin and other impurities in varying properties.
Extract
of Indigo Dye (Powdered, Form) from the Leaf of Indigofera Tinctoria
 N-H

It has been shown by X-ray crystallography that indigo normally exits in the 'trans' form, but both 'cis' and 'trails' have
been isolated; the 'cis' rapidly reverts to the 'trans' form during storage. Many attempts have been made to explain the
deep colour of indigo which appears inconsistent with the absence of a long conjugated chain but phenomenon has
been explained by regarding indigo as resonance hybrid of its canonical structures .|1!i

CH+O2

MATRIALS AND METHODS

Two stages involved in the production of dye from indogofera tintoria; the preparatory stage and the actual
production stage.

THE PREPARATORY STAGE

The pretreatment stage forms the initial stage of sample preparation. Fresh indigofera linctoria plants were
obtained from the local indigo fanners at Diko village in Niger state, the leaves were plucked from the
plants, washed, pre-dried in the sun and then weighed. The weighted sample was then dried in the oven at
60oc for six hours; then it was ground using mortar and pestle. Sieve analysis was used, not to classify the
powder into size, but to remove stone and different foreign bodies (particles) present in the crushed leaves
for the experiment.
s*

METHODOLOGY
30g of dried, crushed sample of indigofera tinctorial leaves were added to the beakers containing 250ml of
different concentration of sodium hydroxide solution (2.0m, 2.5m, 3.0m and 4.0m NaOH), to compare the
percentage yield with concentration of sodium hydroxide solution. The mixture was then stirred for about five
minutes for maximum contact and allowed to stand for maximum extraction. The mixture was then filtered
with filter paper to separate the residue from the filtrate the residue collected was washed with sodium
hydroxide solution for complete extraction. 28% glacial acetic acid solution in 100cm3 of hydrogen peroxide
(H;O2) was then added to each of the filtrate drop wise to participate the dyestuff from the filtrates. The
precipitates formed were then collected in different crucibles and dried in an oven at a temperature of 110°c
for concentration. The resulting dried indigo-pastes (dyestuff) were ground into powder and stored in a container.
 International Journal of Physical Science, Volume 6, Number 1,2011

FLOWCHART OF THE PROCESS

Fresh indigofera tinctoria leaves.

Drier coven

Crusher

Separator

Extractor

Separator

Drier

Table 1: result of dryingjtnaiysis

Mass of Wet Leaves (g) Time taken (Minutes)
480 464.22 458.13 449.97 449.76 0
60 120 180 240
Extraction of Indigo Bye (Powdered, Form) from the Leaf of Indigofcra Tinctoria
Table 2: Result of Extraction using different concentrations of sodium hydroxide
solution.
Con. Of NaOH 2.00m 2.50m 3.00m 3.50m 4.00m
(mol/dm3) 1 2 1 2 1 2 1 2 1 2
Av Av Av. Av. Av

Vol. of NaOH 250 250 250 250 250 250 250 250 250 250 250 250 250 250 250
(ML)
Vol. of H2O2 60 60 60 60 60 60 60 60 60 60 60 60 60 60 60
Mass of 30 30 30 30 30 30 30 30 30 30 30 30 30 30 30
Sample (g)
Mass of 23.06 23.13 23.66 23.66 23.85 23.65 23.75 24.75 24.47 24.55 24.90 24.8 3 24.87 24.85
Raffinate (R) 23.10
Extractable 6.94 6.87 6.90 6.34 6.34 6.34 6.15 6.35 6.25 5.53 5.38 5.10 5.17 5.13 5.15
Matter (g)
% Extractable 23.13 22.90 21.13 21.13 21.13 20.50 21.17 20.83 18.43 17.93 17.00 23.2 3 17.10 20.16
Matter 23.0
Yield {g) 4.30 4.29 4.30 3.72 3.69 3.71 3.64 3.58 3.61 3.51 3.53 3.25 3.52 3.26 3.3.26
M = C-D
% M = C-D/C X 100/1
Where
C = mass of wet sample (g) = 480g
D =mass of dry sample(g)= 449.76g
M = moisture content = 480-49.76
= 30.24g %
M = 30.24 x 100 =6,3%
480

DISCUSSION OF RESULT
It was observed that when l0ml of acidified hydrogen peroxide solution is added to the
beaker containing the filtrates, different response to the addition was noticed in the beaker
containing 2.0m sodium hydroxide-extract mixture, dark foamy substance were noticed in
the beakers containing 2.0m and 2.5m sodium hydroxide extract mixture. This colour
charges shows that all the dyestuff have been precipitated from the filtrate of 2.0m sodium
hydroxide solution, that the precipitate dyestuffs from 2.0m to 2.5m sodium hydroxide-
extract mixture is almost completed and, those in 3.0m to 4.0m sodium hydroxide-extract
mixture are not.near completion.
Subsequence addition of the acidified hydroxide peroxide solution did not alter the quantity of
precipitate in any the beakers containing 2.0m, and 2.5m sodium hydroxide-extract mixture; but
there were increase in the formation of foamy substance in the other beakers. Again, this shows
that 3 .Om of sodium hydrogen solution only extract little quantity of dyestuff from the sample.
It took 40ml acidified hydrogen peroxide solution to precipitate the dyestuff from the 2.0m and
2.5m sodium hydroxide-extract mixture.
The hydrogen peroxide was acidified because the reduced form of a dye does not lend
itself to easy oxidatiation in an alkali solution; but the precipitator is not too acidic
otherwise; a greenish precipitate containing much of the -enol form of indigo will be obtained.
Table 2 shows that although equal volumes of sodium hydroxide
Solution and mass of sample were used, the percentage of extractable matter and yield obtained
differs; both decreases with increase in concentration of sodium hydroxide 1.27% extractable
matter for water through 23% for 2.0m sodium hydroxide solution to 17. 10% for 4.0m
sodium hydroxide solution; which is in agreement with the 27-30% extractable matter in the
literature.(1''
Table 1 shows the drying of the fresh leaves. The leaves loose moisture rapidily at first but later
gradually with time. The 6.3 moisture content is still within the range of moisture content in other
green leaves.'"' The pH of the dyestuff was found to be 5.5 indicating that the dye is acidic; but
the sodium hydroxide extract mixture was found to be slightly neutral (6.5 to 7.5).

CONCLUSION
The aims of this research work were achieved as the product was obtained in the proposed powdered
form. The pH of the dyestuff obtained is 5.5. the mixture of sodium hydroxide-extract was also found to
be between 6.5 and 7.5.The volume of acidified hydrogen peroxide solution needed to precipitate the
product (dyestuff) from the sodium hydroxide -extract mixture is proportional to the concentration of
sodium hydroxide solution used;
 International Journal of Physical Science, Volume 6, Number 1, 2011

that is, the higher the concentration of sodium solution used, the more the volume of the precipitating agent
needed.
The optimum yield obtained is 4.30g and it is from 2.0m sodium hydroxide solutions; hence for economical
yield concentration between 2.0m and 2.5 sodium hydroxide solution should be employed.
The colour of die dye obtained is deep blue from the colour tinctometer.

RECOMMENDATIONS
Since the quantity of synthetic dyes from the limited number of synthetic dye manufacturing companies in
Nigeria are not meeting up with dye demand of the populace, the development of dye manufacturing from
local plants should be encouraged as it will provide the opportunity to use indigenous raw material hence
provide job opportunity for the country's citizens.
Further research should be conducted in scare for more yields. Other precipitating agents like acidified
potassium tetraoxomanganate VII, acidified ferric chloride solution should be used and die yields obtained
from this research work.
For more, acidified hydrogen peroxide solution should be prepared and sold to local dyes for safety because
of their non-technicality.

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J.M. Tedder,(1972). Basic organic Chemistry. Part4, (Natural products) John Wifley and sons; New York. J. W.

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