food and bioproducts processing 9 0 ( 2 0 1 2 ) 615623

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Food and Bioproducts Processing

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Extraction of anthocyanins from red cabbage and

purication using adsorption
J. Chandrasekhar, M.C. Madhusudhan, K.S.M.S. Raghavarao
Department of Food Engineering, Central Food Technological Research Institute, Council of Scientic and Industrial Research, Mysore
570020, India

a b s t r a c t
There is an increasing interest in anthocyanins, as natural food colorants, in food products and also in pharmaceutical products due to their antioxidative potential. The present study deals with extraction and purication of
anthocyanins from red cabbage. Conventional extraction methods of anthocyanin from plant material are nonselective and yield pigment solutions with large amounts of byproducts such as sugars, organic acids and proteins.
Some of these impurities may accelerate anthocyanin degradation. Different extracting media were used and the
mixture of 50% (v/v) ethanol and acidied water resulted in maximum anthocyanin content (390.6 mg/L). In order to
obtain anthocyanins in a puried form, adsorption was carried out with six different adsorbents. Among these, nonionic acrylic ester adsorbent, namely Amberlite XAD-7HP, showed the highest adsorption capacity (0.84 mg/mL of
resin) and desorption ratio (92.85%). Adsorption results were found to be correlated best using the Langmuir isotherm
equation especially at low temperature. The resulting anthocyanin solution after purication was free from sugars,
which are the major cause for degradation of anthocyanin. No browning was observed and chroma increased by 27%
compared to crude anthocyanin.
2012 The Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.
Keywords: Anthocyanin; Extraction; Adsorption; Purication; Red cabbage; Natural color

1.

Introduction

Edible pigments are important additives, which can increase

the acceptability of food products. The safety of synthetic pigments has been questioned, leading to a reduction in the
number of permitted food colorants (Giusti and Wrolstad,
2003). As a result, interest in natural colorants has increased
considerably, mainly because of the apparent lack of toxicity and eco-friendliness (Chethana et al., 2007). Natural
colors such as betalains from beetroot, carotenoids from
carrot, and anthocyanins from grape extract are some examples, which were evaluated in various food systems and
relatively good stability was observed (La Bell, 1993; Sapers,
1994). Anthocyanins are glycosylated polyhydroxy and polymethoxy derivatives of 2-phenylbenzopyrylium (avylium)
salts accounting for the colors in several fruits, vegetables
and owers (Marianne et al., 2001). They are water soluble,
which facilitates their incorporation into aqueous food systems. Besides their color attributes, anthocyanins have been

reported to be benecial to health as potent antioxidants and

to improve visual acuity (Liu et al., 2004). Anthocyanins have
a range of biological activities that may produce health benets; examples range from inhibition of DNA damage in cancer
cells in vitro (Hou, 2003), induction of insulin production in
isolated pancreatic cells (Jayaprakasam et al., 2005), reduction in inammatory responses (Tall et al., 2004) to protection
against age-related decline in brain function (Lau et al., 2006).
The presence of anthocyanins is universally associated with
attractive colorful fruits around the world, such as grapes,
strawberries, raspberries, pomegranates, mangoes, gs, red
cabbage and sweet potato (Liu et al., 2004; Francis, 1989). Many
natural sources were suggested, but the major problem with
all anthocyanins is their instability during processing and storage (Monica and Ronald, 1996).
Red cabbage (Brassica oleracea L.) is one of the sources of
anthocyanins for coloration of food since its anthocyanins are
unique, exhibiting color over a very broad pH range. In contrast, anthocyanins from other sources such as grape skin,

Corresponding author. Tel.: +91 821 2513910; fax: +91 821 2517233.
E-mail address: fed@cftri.res.in (K.S.M.S. Raghavarao).
Received 27 September 2011; Received in revised form 4 July 2012; Accepted 21 July 2012
0960-3085/$ see front matter 2012 The Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.fbp.2012.07.004

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food and bioproducts processing 9 0 ( 2 0 1 2 ) 615623

black currant and elderberry, which possess a reasonable

degree of color only at pH < 4 (Marianne et al., 2001). The colors of anthocyanins from red cabbage vary from red at low
pH to blue and green at high pH (Mazza and Miniati, 2003) and
their use is therefore not limited to acidic foodstuffs but can be
extended to neutral products as well. Hence they have potential to provide a natural alternative to synthetic colorants.
Red cabbage anthocyanins are currently used to color various
beverages, candies, dry mixed concentrates, chewing gums,
yoghurts, and sauces (Dorota and Janusz, 2007). Conventional
extraction methods of anthocyanins from plant material are
non-selective and yield pigment solutions with large amounts
of byproducts such as sugars, sugar alcohols, organic acids,
amino acids and proteins. Some of these impurities such as
sugars accelerate anthocyanin degradation during storage or
cause problems in further processing steps such as spray drying (Liu et al., 2004).
Efcient extraction methods should maximize anthocyanin recovery with minimal amount of adjuncts and
minimal degradation or alteration of its natural state. Several methods employing solvents such as ethanol, methanol
and acetone for the extraction of anthocyanin are efcient.
However, some of these solvents such as methanol/acetone
could be toxic to human health (Patil et al., 2009). Adsorption is an effective method for purication of anthocyanin
pigments in a single step, while achieving the elimination of
polar, nonphenolic impurities. Adsorption has been used for
purication of secondary metabolites, including hesperidin
(Scordino et al., 2003), genistein and apigenin (Liu et al.,
2010), rutin and quercetin (Zhao et al., 2011). Different adsorbents for the purication of anthocyanins were reported from
cultivars of mulberry (Liu et al., 2004), Arona melanocarpa
var Nero (Kraemer-Schafhalter et al., 1998) and red cabbage
(Lopes Toni et al., 2007). The pigments of red cabbage consist
primarily of cyanidin 3-sophoroside-5-glucoside and cyanidin 3-sophoroside 5-glucoside acylated with sinapic, ferulic,
p-coumaric and malonic acids (Sapers et al., 1981). These
anthocyanins are almost entirely acylated, have high pKa
values and high value of tinctorial strength (Francis, 1989).
Because of these special characteristics, which give more stability to anthocyanins, red cabbage was selected as source in
the present study. Thus the extraction of anthocyanins from
red cabbage followed by purication by adsorption employing
six different adsorbents was carried out.

2.

Materials and methods

2.1.

Materials

Adsorbents namely, Amberlite XAD7HP, Amberlite XAD4,

Amberlite IRC 80, Amberlite IRC 120 and Dowex 50WX8 (hydrogen form) were procured from Sigma Aldrich, St. Louis, USA.
Silica gel (60120 mesh) was obtained from SD ne-chem Ltd.,
Mumbai, India. Red cabbage was purchased from local market.
All the chemicals used were of analytical grade.

2.2.

Methods

2.2.1.

Extraction of anthocyanin

Red cabbage leaves were cut into small pieces. Extraction was
carried by taking 25 g of these leaves and 50 mL of extraction media while providing thorough contact in a mixing
unit (Singer, India FP-450). The solidliquid (leaves to extraction media) ratio of 1:2 and the contact time of 1 min were

maintained for all the extractions. Different types of solvents

namely, water, acidied water, mixtures of ethanol with water
and acidied water (at different volume ratios), methanol,
acidied methanol, acetone and 70% aqueous acetone were
used for extraction. The anthocyanin extract obtained in each
extraction was ltered through a muslin cloth to remove
coarse particles. In order to know the efciency of the extraction medium, the number of extractions was restricted to
only one. In other words, after the rst extraction the lter
cake obtained was discarded. The individual ltrates obtained
for different extraction media were centrifuged separately
at 6000 rpm for about 15 min to remove the ne suspended
particles. A UVvis spectrophotometer (Double beam spectrophotometer, Shimadzu, Japan, Model UV-160A) was used
for the analysis. The anthocyanins concentration in all the
extracts was estimated using pH differential method, employing the following equation (Flueki and Francis, 1986) and the
results were compared.
Anthocyanin pigment concentration (mg/mL)
=

A Mw DF
L

(1)

where A = (A530 A700 )pH 1.0 (A530 A700 )pH 4.5 , Mw is the
molecular weight of anthocyanin (449.2 g/mol), DF is the dilution factor, is the extinction coefcient (26,900 L/cm mol) and
L is the path length (1 cm). Considering that red cabbage anthocyanins are all derived from cyanidin glycoside, quantitative
data were expressed as cyanidin-3-glycoside (Hrazdina et al.,
1977).

2.2.2.

Adsorption and desorption experiments

The adsorption process of red cabbage anthocyanins extract

was carried out in the following manner. 2 mL of each adsorbent (activated) was contacted with 30 mL of centrifuged
extract of red cabbage in a 50 mL conical ask while agitating on a vibratory shaker. The solutions after adsorption were
analyzed for anthocyanins. The following equation was used
to quantify the adsorption capacity (qe mg/mL of adsorbent)
at equilibrium.
qe =

(C0 Ce )Vi
Va

(2)

where, C0 and Ce are the initial and equilibrium concentrations

of anthocyanins in the solution (mg/mL), respectively. Vi and
Va are the volumes (mL) of initial sample solution adsorbent,
respectively.
Desorption study was performed as follows: after reaching
adsorption equilibrium, the resins were washed by deionized water and then desorbed with 30 mL of ethanol solution.
The eluent samples were analyzed by spectrophotometer. The
following equations were used to quantify the desorption
capacity (qd mg/mL of resin) at equilibrium and ratio of desorption (D).
qd =

D=

Cd Vd
Va
Cd Vd
(C0 Ce ) Vi

(3)

(4)

where Cd is the concentration of the solute in the desorption

solution (mg/mL) and Vd is volume of the eluent (mL).

food and bioproducts processing 9 0 ( 2 0 1 2 ) 615623

2.2.3.

Equilibrium studies and adsorption isotherms

Activation of adsorbent was performed by overnight treatment with 2 bed volumes (BV) of distilled ethanol followed by
rinsing with 5 BV of distilled water. All the adsorption experiments were carried out in a batch mode at room temperature
(25 1 C). 2 mL of each adsorbent (activated) was contacted
with 15 mL of centrifuged extract of red cabbage in a 50 mL
conical ask while agitating on a vibratory shaker. Aliquots
of 100 L were drawn from the solution at regular intervals of
2 min, up to 20 min, in order to study the equilibrium of the
adsorption process. Analysis was carried out and the amount
of anthocyanin adsorbed was calculated by the difference.
In order to obtain equilibrium adsorption isotherms of the
adsorbent selected (Amberlite XAD7HP), experiments were
conducted by contacting 20 mL of sample solutions at different
concentrations with 2 mL of resin, and then shaking for 1 h at
25 2 C and 30 1 C, respectively. The initial and equilibrium
concentrations at different temperatures were determined
by spectophotometric method. The Langmuir and Freundlich
equations are frequently used to describe adsorption behavior between sorbent and adsorbed material (Langmuir, 1918; Al
Duri, 1996). The Langmuir isotherm is best known for monolayer adsorption and the equation is given by
qe =

qm Ce
KL + Ce

(5)

where KL is the adsorption equilibrium constant and qm is the

empirical constant.
Another widely used model for nonideal adsorptions is Freundlich isotherm, an empirical equation and can be expressed
as
1/n

qe = KF Ce

where KF is the Freundlich constant and 1/n is an empirical

constant. qe and Ce are the same as described above.

Japan). The Dubois method (Dubois et al., 1956) was used for
the estimation of total sugars present in the extract. The sugars have absorption maxima at 480 nm. Dextrose was used as
a standard for the determination of sugars. Parameters such
as browning and degradation index describing the quality of
anthocyanins were estimated (Wrolstad et al., 2005) as
Browning = absorbance at 420 nm of bisulfate treated sample
Degradation index = ratio between monomeric and total
anthocyanin concentration

2.2.7.

2.2.8.

HPLC analysis of anthocyanins

The HPLC analysis of anthocyanins was performed as

described by Liu et al. (2004). The pigment solutions were ltered through a 0.45 m syringe-driven lter unit (Millipore
Corporation, Bedford, MA). The HPLC system consisted of two
LC-10AD pumps, SPDM10A diode array detector, CTO-10AS
column oven, DGV-12A degasser, SIL-10AD auto injector, and
SCL-10A system controller (Shimadzu, Japan) equipped with
Luna (3 m C18(2), 4.6 mm 100 mm, Phenomenex, CA) column at 25 C. The following solvents in water with a ow rate
of 1 mL/min were used: A (1.5% phosphoric acid) and B (1.5%
phosphoric acid, 20% acetic acid and 25% acetonitrile). The
elution prole, a linear gradient elution with 2585% solvent B
in solvent A, was for 85 min. The chromatograms were monitored at 530 nm and recorded and the relative concentrations
of individual pigments were calculated from the peak areas.

Results and discussion

Elution experiments

To determine the optimum ethanol concentration for elution of the adsorbed anthocyanins, 2 mL of Amberlite XAD7HP
adsorbent saturated with anthocyanins was contacted with
different proportions of 25 mL of acidied (1% HCl, v/v)
aqueous ethanol (water:ethanol) while agitating in different
conical asks using a vibratory shaker. The contents of anthocyanins in the eluates were measured spectrophotometrically
as described earlier (Section 2.2.1).

2.2.5.

Color analysis

The color characteristics (Hunter CIE L*, a*, b*) of anthocyanin

before and after purication were measured using a colorimeter (Lab Scan XE, Hunter Lab, Virginia). The sample was placed
in 1 cm path length optical glass cell and CIE L*, a*, b* values
were measured in the transmission mode, using Illuminant C
and 2 observer angle.

3.
2.2.4.

617

Dynamic break through curve

In order to obtain the break-through point dynamic adsorption

and desorption experiments were carried out in glass columns
(2 cm diameter and 11 cm length) with 50 mL of resin. Anthocyanin solution passed through the glass column at the ow
rate of 2 mL/min and the concentration of anthocyanins was
analyzed. After reaching adsorptive saturation, the adsorbent
in the column was washed thoroughly using deionized water
and then eluted using ethanol (100%, v/v) with a ow rate of
2 mL/min.

The results of extraction of anthocyanin employing different

solvents followed by purication using adsorption are discussed in the following sections.

3.1.

Extraction of anthocyanin with acidied water

In order to examine the effect of HCl concentration on the

degree of extraction of anthocyanin, concentration of HCl was
varied from 0% to 4% (v/v) in water and the results are presented in Fig. 1. It can be observed from gure that the degree
of extraction of anthocyanin was higher in the case of acidied
water when compared to water and the degree of extraction
increased with an increase in HCl concentration. The stability of anthocyanin increases with a decrease in pH (Kirca
et al., 2007). Although the degree of anthocyanin extraction
increased with an increase in concentration of HCl, lower concentration of HCl (1%, v/v) was selected in the subsequent
extractions, because higher concentration of HCl is not preferable for the food applications (Patil et al., 2009).

2.2.6. Determination of pH, soluble solids and sugar

concentration

3.2.

The measurement of pH of anthocyanin extracts before and

after adsorption was carried out using a pH meter (Control
Dynamics, India, APX 175). Soluble solid content, expressed
as refractive index, was measured with refractometer (ERMA,

In order to know the effect of ethanol concentration on the

degree of extraction of anthocyanin, the concentration of
ethanol in water was varied from 20% to 80% (v/v) and the
results are presented in Fig. 2. It can be observed from gure

Extraction of anthocyanin with ethanol

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food and bioproducts processing 9 0 ( 2 0 1 2 ) 615623

500

350

311
258

450

280

275

200
150
100
50

300

354
321

280

250
200
150

50

1%

water

2%

3%

4%

0
water

HCl Concentration (% v/v)

Extraction of anthocyanin in combination

In order to explore the synergistic effect of HCl and ethanol,

extraction was carried out with mixture of ethanol and acidied water. The concentration of HCl in water was maintained
at 1% (v/v) and the ethanol concentration in water was varied
from 20% to 80% (v/v). The results are presented in Fig. 3. It can
be observed that the degree of extraction increased with an
increase in ethanol concentration in acidied water up to 50%
(v/v) and decreased with further increase. Maximum extraction of anthocyanin was observed as 387.6 mg/L at 50% (v/v)

450
381
365

365
342

350

40%

50%

60%

80%

Fig. 3 Effect of acidied aqueous ethanol concentration on

anthocyanin extraction.

that the degree of extraction increased with an increase

in concentration of ethanol in water up to 50% (v/v) and
decreased with further increase. The decrease in anthocyanin content above 50% (v/v) ethanol concentration could
be mainly due to the non-extraction of hydrophilic anthocyanins as the concentration of water in the extraction media
decreased with an increase of ethanol content. A maximum
anthocyanin content of 381.1 mg/L at 50% ethanol in water
was observed. It was reported that the presence of small
amounts of water is required for the extraction of hydrophilic
anthocyanins (Patil et al., 2009), which supports the above
observation. Hence, it can be inferred that pure ethanol is not
preferable as an extracting solvent.

400

20%

Acidified alcohol concentration (v/v)

Fig. 1 Effect of HCl concentration on anthocyanin

extraction.

341

308
Anthocyanins (mg/L)

328

350

345

100

3.3.

387

400

230

250

Anthocyanin (mg/L)

Anthocyanins (mg/L)

300

300

ethanol in acidied water. The synergistic effect of HCl and

ethanol was not signicant. The decrease in degree of extraction above 50% (v/v) ethanol can be explained based on the
similar reasons given in the previous section.

3.4.

Extraction of anthocyanin by other solvents

Extraction was carried out with mixture of other solvents such as methanol, 0.1% HCl in methanol, 1% HCl in
methanol, 3% formic acid in methanol, methanol + formic
acid + water (50% + 5% + 45%), methanol + acetic acid +water
(50% + 1% + 49%), 70% aqueous acetone, acetone and 3% formic
acid in water. The results are presented in Fig. 4. It can
be observed that the degree of extraction was highest in
case of acetone (425.9 mg/L) followed by 1% HCl in methanol
(419.7 mg/L) when compared to water alone.
From the results discussed so far it can be observed that
the degree of extraction was the highest in case of acetone
followed by 1% (v/v) HCl in methanol and then by acidied ethanol (50%, v/v ethanol). As already mentioned, use of
acetone and methanol for food applications was not preferable (Spagna et al., 2003; Patil et al., 2009). Among all these
extraction media, ethanol is the most acceptable one for use
in food industry. The extraction with acidied ethanol (50%,
v/v ethanol) showed comparatively better results than the
remaining solvents. Hence, in further experiments acidied
ethanol (50%, v/v ethanol) was used. The pH values of combinations of the above solvents and acids were in the range of
0.35.1 as indicated in Fig. 4.

250

3.5.

Adsorption and desorption experiments

200
150
100
50
0
water

20%

40%

50%

60%

80%

Alcohol concentration (v/v)

Fig. 2 Effect of ethanol concentration on anthocyanin

extraction.

It may be noted that the adsorption and desorption performances correlate with the properties of the adsorbents and
chemical features to the solute. Adsorbents similar in polarity
to solute exhibited better adsorption ability. Physical features of adsorbent also play an important role in process of
adsorption and desorption. As shown in Fig. 5, the adsorption
and desorption capacities were high in the case of Amberlite
XAD7HP which can be attributed not only to its similar polarity with the anthocyanins (Table 1), but also to its high surface
area.

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food and bioproducts processing 9 0 ( 2 0 1 2 ) 615623

500
450
400

Anthocyanins (mg/L)

426

420
398

393
373

380

374

334

350
301

296

300
250
200
150
100
50
0
1

10

Solvent

1. Water (pH 5.1); 2. Methanol (pH 4); 3. 0.1% HCl in methanol (pH 0.32); 4. 1% HCl in
methanol (pH 0.3); 5. 3% formic acid in methanol (pH 1.2); 6. Methanol + formic acid + water
(25+2.5+22.5) (pH 1.3); 7. Methanol + acetic acid +water (25+0.5+24.5) (pH 1.1); 8. 70%
aqueous acetone (pH 5.08); 9. Acetone (pH 3.65); 10. 3% formic acid in water (pH 1.1)

Fig. 4 Effect of different solvents on anthocyanin extraction.

3.6.

Equilibrium studies and adsorption isotherms

Equilibrium studies were carried out in a batch mode to

determine the conditions for maximum adsorption of anthocyanins on Amebrlite XAD7HP and the results are presented
in Fig. 6. It can be observed from gure that for all the adsorbents, the adsorption capacity for anthocyanins increased
with an increase in contact time before reaching the equilibrium. The equilibrium time (25 2 C) of anthocyanins was

Table 1 Modes of adsorption and properties of different

adsorbents.
Adsorbent

Mode of adsorption

Silica gel
Amberlite IRC 80
Amberlite IR 120
DOWEX 50WX8
Amberlite XAD4

Reversed phase, non polar

Weakly acidic anion exchanger
Weakly acidic cation exchanger
Strongly acidic cation exchanger
Non-ionic acrylic ester resin moderate
polarity
Non-ionic acrylic ester resin moderate
polarity

(a)
Adsorpon capacity (mg/g)

Amberlite XAD7

1.4
1.2

Adsorpon capacity

Desrbed amount (mg)

0.8
0.6
0.4
0.2
0
XAD7

XAD4

Dowex

IRC 120

IRC80

Silica gel

Absorbent
100

Desoron rao (%)

80
60
40
20
0
XAD7

XAD4

Dowex

IRC 120

IRC80

Silica gel

Adsorbent

Fig. 5 (a) Adsorption and desorption capacities of

anthocyanins on different resins and (b) desorption ratio of
adsorbents.

Adsorbed anthocyanin conc. (mg/L)

(b)

found to be about 15 min for all the resins. It also can be

observed that Amberlite XAD7 has shown highest adsorption capacity. Hence, the following experiments were carried
out to obtain the adsorption isotherms. Adsorption capacity reaches its maximum value at equilibrium and hence
beyond this only negligible change in the anthocyanin concentration in the solution was observed. The distribution of
anthocyanins between the liquid and solid phases is a measure of the position of equilibrium in the adsorption process
and is expressed by the Langmuir and Freundlich equations.

200
180
160
140
DOWEX
XAD 4
XAD 7
IRC 80
IRC 120
Silica gel

120
100
80
60
40
20
0
2

10

12

14

16

18

20

Time (min)

Fig. 6 Adsorption equilibrium studies of anthocyanin.

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food and bioproducts processing 9 0 ( 2 0 1 2 ) 615623

(a) 1.4

450

(a)

400

1
0.8
25 C

0.6

30 C

0.4

350

Concentraon (mg/L)

Adsorpon capaciy (mg/g)

1.2

300
250
200
150
100
50

0.2

0.05

0.1

0.15

Equilibrium concentraon (mg/mL)

400
600
Fracon volume (mL)

800

(b) 14000

0.16

12000

0.14

Concentraon (mg/L)

(b)

200

0.2

y = 0.484x + 0.069
R = 0.932

0.12
Ce/qe

25 C

0.1

30 C

10000

Eluon

8000
6000
4000
2000

y = 0.378x + 0.060
R = 0.996

0.08

0
0

50

100

150

200

250

300

Fracon volume (mL)

0.06
0

0.05

0.1

0.15

0.2

Fig. 8 Adsorption of anthocyanin using Amberlite

XAD7HP.

Ce (mg/L)

Fig. 7 (a) Adsorption isotherms of anthocyanins on

Amberlite XAD7HP and (b) Langmuir plot for adsorption of
anthocyanins.

The Freundlich equation (Eq. (5)) can be rearranged as

log qe = log KF +

Accordingly, equilibrium adsorption isotherms, constructed

at the temperatures of 25 2 C and 30 1 C, were analyzed.
The initial concentrations of extracts of anthocyanins were
0.0525, 0.0693, 0.105, 0.14 and 0.21 mg/mL, respectively. The
results are presented in Fig. 7(a) and (b) and Table 2. It can be
seen in Fig. 7(a) that the adsorption capacity increased with
an increase in initial concentration.
The Langmuir equation (Eq. (5)) can be rearranged to the
linearized form with Ce and Ce /qe as independent variable as
Ce
KL
Ce
=
+
qe
qm
qm
The linear plot of Ce /qe versus Ce (Fig. 7(b)) ensures the
applicability of Langmuir adsorption isotherm. From Table 2,
the values of Langmuir parameters, qm , KL and R2 are 2.6455,
0.1587, 0.996 and 2.066, 0.1425, 0.932 at 25 and 30 , respectively. It can be seen in Fig. 7(a) that at the same initial
concentration, the adsorption capacity is high at low temperature, which indicates the adsorption to be a thermopositive
process (Zhao et al., 2011). Therefore, 25 2 C was used in the
following experiments.

R2

R2

Temperature
( C)

Langmuir
equations

25

qe =

2.6455Ce
0.1587+Ce

0.996

qe = 6.479C0.41
e

0.993

30

qe =

2.066Ce
0.1425+Ce

0.932

qe = 6.0.32C0.41
e

0.99

Freundlich
equations

From Table 2, the values of KF are 6.479 and 6.023 while

the values of 1/n are 0.41 and 0.41. The value of adsorption
intensity (1/n) less than unity indicates a favorable adsorption.
The correlation coefcients of both Langmuir and Freundlich
equations are high, which indicates the favor of adsorption of
anthocyanins.

3.7.

Elution experiments

In devising a potential industrial production process for any

target product, it is necessary to take into consideration
not only the adsorbent, but also the eluent solvent and
its concentration. In order to know the effect of ethanol
concentration on the elution of anthocyanin, studies were
carried out with acidied aqueous ethanol (with 1%, v/v HCl
and ethanol concentration varying from 20% to 100%, v/v)
and the results are presented in Table 3. It can be seen from

Table 3 Effect of ethanol concentration on the elution of

anthocyanin.
Concentration
of ethanol (%)

Table 2 Langmuir and Freundlich parameters for the

adsorption of anthocyanins.

1
log Ce
n

20
40
50
60
80
100

Anthocyanin
eluted (mg/L)
11.02
57.86
79.40
104.45
103.20
112.71

Elution (%)
9.11
47.10
64.63
85.07
84.01
91.74

food and bioproducts processing 9 0 ( 2 0 1 2 ) 615623

621

Fig. 9 HPLC proles of anthocyanin solution.

the table that acidied ethanol of concentrations above 60%

(v/v) could effectively elute anthocyanins from the adsorbent.
Hence, for higher anthocyanin recovery, higher concentration
of ethanol is recommended. Further, it may be noted that
aqueous ethanol with low concentrations will have a higher
boiling point making it more difcult to concentrate than
those with higher concentrations.

3.8.
Dynamic breakthrough curve on Amberlite
XAD7HP
The dynamic breakthrough curve on Amberlite XAD7HP resin
was obtained based on the volume of eluent liquid and the
concentration of solute therein and is given in Fig. 8(a). It can
be seen from the gure that anthocyanins in solution were
almost completely adsorbed by the resin before the cumulative volume of the fractions reaching 260 mL (breakthrough
point). Above this point, the concentration of anthocyanins in
the solution (coming out of the adsorption column) increased
until it reached a steady plateau when the cumulative volume
of 460 mL is reached (adsorption break-through). Therefore,
volume of 260 mL of sample solution was selected for dynamic
adsorption experiments.

The dynamic desorption curve was obtained based on the

volume of desorption solution and the concentration of solute
therein and is given in Fig. 8(b). It can be seen from the gure
that anthocyanins could be completely desorbed by approximately 150 mL (3 BV) of desorption solution.
The optimum adsorption process parameters for the
enrichment of anthocyanins with Amberlite XAD7HP arrived
at are as follows, for adsorption: concentration of anthocyanins in sample solution, processing volume, ow rate
and temperature are 0.21 mg/mL, 5.3 BV, 2 mL/min and
25 2 C, respectively; and for desorption: eluent volume and
ow rate are 3 BV of ethanol (100%, v/v) and 2 mL/min,
respectively.

3.9.

Evaluation of anthocyanin solution

Along with anthocyanin pigments the red cabbage extract

also contains other components. Hence, in order to examine the nal product (anthocyanin) stability, evaluation of
anthocyanin solution was carried out for total sugars, soluble
solids and pH of the extract before and after adsorption with
Amberlite XAD7H under saturated adsorption conditions. In
addition, the colorant properties browning, degradation index,
color density and Hunter L*, a*, b*, values were evaluated. The

622

food and bioproducts processing 9 0 ( 2 0 1 2 ) 615623

Table 4 Evaluation of stability of anthocyanin.

Components

Before
purication

After
purication

pH
Sugars
Total anthocyanins recovered
Total solids
Browning
Degradation index
Color density
L
a
b
Hue angle
Chroma

2.32
224.2 g/mL
20 mg
11 B
0.055
1.96
10.53
17.81
32.66
9.66
16.47
34.05

1.63
0.07 g/mL
18.56 mg
18.5 B
0
1.20
2.25
25.04
44.81
13.01
16.18
46.67

results are presented in Table 4. It can be observed from the

table that the concentration of sugars (which are the major
cause for product degradation) was observed to decrease from
an initial 224.7 g/mL to 0.07 g/mL after purication. Browning was observed to be nil, degradation index decreased from
1.96 to 1.2 and the chroma value increased from 34.05 (crude)
to 46.67 indicating an increase in the stability of anthocyanin
after purication.

3.10.

HPLC analysis of anthocyanin

The HPLC proles of the puried and crude pigment extracts

were identical, consisting of three peaks (Fig. 9). This suggests
that pigment purication using the adsorbents did not change
the composition of the anthocyanin mixture.

4.

Conclusions

An integrated process for extraction and purication of anthocyanin pigment from red cabbage was developed. Among the
employed extraction media acidied aqueous ethanol (50%,
v/v) showed the best results (anthocyanin content 390.6 mg/L).
Adsorption could be successfully employed for the purication
of anthocyanin. Out of six adsorbents employed, Amberlite XAD7HP showed highest adsorption capacity (0.84 mg/mL
of resin) and elution capacity (92.85% recovery). Acidied
pure ethanol showed the best result for anthocyanin elution. The evaluation of the nal product with respect to
total sugars, chroma, browning and degradation index indicated an increase in the stability of anthocyanin after
adsorption.

Acknowledgments
Authors thank the Director, CFTRI, Mysore, for the infrastructural facilities at the institute. Chandrasekhar J. and
Madhusudhan M.C. gratefully acknowledge CSIR, Government
of India for the research fellowship.

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