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Comparison of Acidic and Enzymatic Pectin Extraction from Passion Fruit

Peels and Its Gel Properties

Article  in  Journal of Food Process Engineering · June 2016

DOI: 10.1111/jfpe.12243

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Journal of Food Process Engineering ISSN 1745-4530

COMPARISON OF ACIDIC AND ENZYMATIC PECTIN

EXTRACTION FROM PASSION FRUIT PEELS AND ITS
GEL PROPERTIES
S.Q. LIEW, N.L. CHIN1, Y.A. YUSOF and K. SOWNDHARARAJAN
Department of Process and Food Engineering, Faculty of Engineering, Universiti Putra Malaysia, Serdang, Selangor 43400, Malaysia

1
Corresponding author. ABSTRACT
TEL.: +60389466353;
FAX: +60389464440; The influences of extractor concentration, extraction temperature and time on the
EMAIL: chinnl@upm.edu.my yield of pectin and degree of esterification (DE) were investigated by the acidic
and enzymatic extraction methods. Citric acid and Celluclast were selected as
Received for Publication January 19, 2015
pectin extractors for environmentally friendly reasons. The peels of yellow passion
Accepted for Publication April 29, 2015
fruit using the acidic and enzymatic extraction methods gave pectin yield of 7.16
doi:10.1111/jfpe.12243 and 7.12%, and DE of 71.02 and 85.45% in the optimized condition of extraction
time of 102 min with citric acid concentration of 0.19% (w/w) at 75C and
Celluclast concentration of 1.67% (w/w) at 61.11C, respectively. The enzymatic
extraction method has greater capability of producing high methoxyl pectin. The
morphological features of fruit peel powder and the extracted pectin examined by
scanning electron microscopy suggested that the nanostructure of wet passion
fruit pectin was dependable on the type of extraction process. The formed pectin
gel also has pseudoplastic liquid behavior and its viscosity was greatly affected by
sugar.

PRACTICAL APPLICATIONS
Pectin has been intensively used as natural gelling agent and stabilizer to alter
rheological properties in food ingredients by most food processing industries to
achieve desired textural quality. Pectin could be recovery from fruit wastes. The
conversion of passion fruit peel into pectin offers great scope for utilization. Citric
acid and enzymatic extraction methods are effectively used for pectin extraction
which may be of interest by pectin industry and consumer with these eco-friendly
processing technology with no using harmful chemicals. Furthermore, scientific
work of this study such as the optimized condition, morphological features of
extracted pectin and pectin gel formation contributes valuable information on
pectin, which could be beneficial for pectin industry improving the process quality
of pectin as well as process profitability.

ing countries of Brazil, Indonesia, Columbia and Ecuador

INTRODUCTION
The yellow passion fruit (Passiflora edulis f. flavicarpa
Degener) is a popular tropical fruit primarily due to its
unique pleasant combination of sweet and sour flavors. Its
juicy pulp is rich in vitamin C (16.5 to 19.1 μg/mL; De
Oliveira et al. 2012), total carotenoids (0.036 to 0.631 mg/g
DW; De Oliveira et al. 2013), and also antioxidant proper-
ties (Janzatti et al. 2012). Because of these high nutritional
values, an annual worldwide production of passion fruits
has achieved 951,000 metric tonnes from the major produc-
(González and González 2010). The dietary and albedo-
fiber powders obtained from the co-products of yellow
passion fruit registered a higher level of total phenolic and
flavonoid contents with strong antioxidant and antibacterial
properties (López-Vargas et al. 2013). The albedo-fiber was
used to improve nutritional value of pork burgers, particu-
larly increasing the fiber content level (López-Vargas et al.
2014). Additionally, pectin has been extracted from the
passion fruit peels in a recent effort of waste recoveries
(Pinheiro et al. 2008; Seixas et al. 2014).

Journal of Food Process Engineering •• (2015) ••–•• © 2015 Wiley Periodicals, Inc. 1
ACIDIC AND ENZYMATIC PECTIN EXTRACTION FROM FRUIT PEELS
Besides being used as an edible film (Espitia et al. 2014),
food packaging (Bierhalz et al. 2012; Penhasi and Meidan
2014) and fruit-filling (Agudelo et al. 2014), pectin has been
recently exploited for different biomedical applications
including drug delivery, gene delivery, wound healing and
tissue engineering due to its simple and cytocompatible
gelling mechanism (Munarin et al. 2012). These wide
variety of applications of pectin in food and drug industries
had led to the exploration of effective methods to obtain
pectin from various sources, such as using subcritical water
from apple pomace and citrus peel (Wang et al. 2014),
microwave-induced heating (Seixas et al. 2014) and high
hydrostatic pressure (Guo et al. 2014) from the peels of
pomelo, besides the conventional method of acid extraction
(Pinheiro et al. 2008). However, recent awareness on envi-
ronmental impact has led to the emergence of enzymatic
extraction method to obtain pectin. Pectin was extracted
using several enzymes from a variety of sources such as the
cellulase and α-amylase for the pumpkin (Cui and Chang
2014), cellulase for the butternut (Fissore et al. 2013),
Viscozyme L for Yuza (Lim et al. 2012) and apple pomace
(Min et al. 2011), Celluclast and alcalase for the rapeseed
cake (Jeong et al. 2014) and α-amylase and neutrase for the
banana peel (Qiu et al. 2010). Celluclast 1.5 L is commonly
used to promote the release of pectin substances in pectin
extraction. This enzyme acts as catalyst to breakdown cellu-
lose in chicory cell wall and turns them into glucose,
cellobiose and higher glucose polymers (Panouille et al.,
2006).
The aim of the present study was to compare pectin
extraction from the conventional acid extraction method
with the enzymatic method for the yellow passion fruit peel.
The optimum condition for pectin extraction was studied
by varying three extraction conditions, the extractor con-
centration, extraction temperature and time on pectin yield
and degree of esterification (DE). The extracted pectin was
classified by its type, and formed into pectin gels where its
viscosity was measured.
MATERIAL AND METHODS
Preparation of Raw Materials
Yellow passion fruits (Passiflora edulis f. flavicarpa Degener)
at the same stage of ripeness, i.e., with similar peel colors
were collected from the Multi-Rich Pitaya Orchard,
Selangor (Malaysia) during the months of January to April
2012. Firstly, the fruit was washed twice with distilled water
and then the flesh was separated from the fruit peels. The
peels were dried in a convectional oven (UM 500, Memmert
GmbH, Schwabach, Germany) at 55C until a constant
weight was achieved. The dried peels were then milled into
500 μm powder using an electronic miller (DFT-150,
2 Journal of Food Process Engineering •• (2015) ••–•• © 2015 Wiley Periodicals, Inc.
S.Q. LIEW ET AL.
Dickson, Zhejiang, China). The fruit peel powder was
packed in a polyethylene bag and stored at −15C in a freezer
(ACF15F, Acson, Kuala Lumpur, Malaysia). Citric acid
(Systerm, Shah Alam, Malaysia) and Celluclast 1.5 L
(ρ = 1.22 g/mL; Novozymes, Copenhagen, Denmark) were
as the acidic and enzymatic extractor by varying its concen-
trations. All analytical chemicals used were of analytical
grade and supplied by Syarikat Saintifik Jaya Sdn Bhd.
Selangor, Malaysia.
Acidic and Enzymatic Extraction Procedures
Pectin was extracted under different concentration, time
and temperature by the acidic and enzymatic extraction
methods. A total of 10 g of fruit peel powder, measured on
an analytical balance (B204-S, MK II, Mettler Toledo,
Greifensee, Switzerland) was blended with distilled water.
The acidic extraction was carried out at different acid solute
concentrations of 0.05, 0.14 and 0.21% (w/w) using 20, 65
and 110 mL of 0.1 N citric solution which contained 0.14,
0.46 and 0.77 g of citric acid, respectively, mixed with
250 mL distilled water at total weight of mixture 269.96,
314.87, 359.78 g while the enzymatic extraction was at con-
centration of 0.49, 1.68 and 2.84% (w/w) using 1.0, 3.5 and
6.0 mL of Celluclast (ρ = 1.22 g/mL; 700 EGU/g). The
mixture was then stirred using a stirrer until the fruit peel
powder was evenly moistened by distilled water in a homog-
enous form. The pectin extraction procedure was continued
by treating the samples for 30, 75 and 120 min at 35, 60,
85C in a shaking water bath (Lab Companion 37L, Jeio
Tech, Seoul, Korea). The mixture solution was kept at room
temperature for 24 h for protein precipitation.
The precipitated pectin was recovered by using a refriger-
ated centrifuge (Mikro 22R, Hettich, Tuttlingen, Germany)
at 6,000 rpm for 10 min. Water bath heat-treated samples
were then filtered and double volume of 95% ethanol (1:2
v/v) was added to allow more pectin precipitation. The
samples were stored in dark condition at room temperature
of 25C for 24 h to allow pectin flotation which was later
separated by filtration and subsequently washed twice using
70% ethanol. Acetone was then added in a drop-wise
manner until the top liquid phase completely cleared to
remove unwanted color of pectin (Pinheiro et al. 2008). The
resulting pectin substance was dried in a conventional oven
(UM500, Memmert GmbH, Schwabach, Germany) at 65C
until a constant weight was reached. The percent yield of
pectin from the fruit peel was determined as a gram of
product obtained per 10 g of fruit peel powder following
Eq. (1):
Product obtained ( g )
Pectin yield (%) = × 100% (1)
10 ( g ) Fruit peel powder
S.Q. LIEW ET AL. ACIDIC AND ENZYMATIC PECTIN EXTRACTION FROM FRUIT PEELS

carboxy group was calculated from the volume of 0.1 N

Determination of DE
The DE is defined as a ratio of esterified galacturonic acid
groups to the galacturonic acid groups present (Van Buren
1991). The DE of pectin was determined by titrimetric
method of Food Chemical Codex with a slight modifica-
tion. 0.2 g of dried pectin sample was moistened with
ethanol and dissolved in 20 mL distilled water. After the
sample was completely dissolved, three drops of phenol-
phthalein were added, the sample was titrated with 0.1 N
sodium hydroxide and the result was recorded as initial
titration volume once slight pink color appeared. The
number of free carboxy group was calculated from the
volume of 0.1 N sodium hydroxide solution spent for
the initial titration. Then, 10 mL of 0.1 N sodium hydroxide
was added to neutralize the polygalacturonic acid. The
sample was plugged with a stopper and shaken vigorously,
then allowed to stand at room temperature for 2 h to
de-esterify pectin. 10 mL of 0.1 N hydrochloric acid was
added to the neutralized sodium hydroxide and the sample
was shaken until the pink color disappeared. Three drops of
phenolphthalein were added into the sample for further
titration with 0.1 N sodium hydroxide. The volume of
titration was recorded as the final titration volume, once a
slight pink color appeared. The number of the esterified
sodium hydroxide solution spent for the final titration fol-
lowing equation (2):
⎛ ⎞
⎜ Final titration volume (mL ) ⎟
DE (%) = ⎜ ⎟ × 100% (2)
⎜ Initial titration volume (mL ) ⎟
⎝ + Final titration volume (mL ) ⎠
Experimental Design and Statistical Analysis
A response surface methodology (Design Expert 8, Stat-Ease,
Inc., Minneapolis, U.S.A.) was used to determine the
optimum conditions for pectin extraction from passion fruit
peel following the central composite design. The three inde-
pendent variables used were the acid or enzyme concentra-
tion, extraction time and temperature. The complete design
consisted of 20 runs including seven axial experiments
(levels ± α), seven factorial experiments (levels ±1) and six
replicates in a center point (levels 0; Table 1). The response
functions measured were pectin yield and DE of extracted
pectin. All measurements were made in triplicate. A third-
order polynomial model equation (Eq. 3) was defined and
fitted for each response from an experimental data.

TABLE 1. CENTERED COMPOSITE DESIGN WITH AVERAGE RESPONSES VALUES FOR EACH RUN OF ACIDIC AND ENZYMATIC EXTRACTION FROM
YELLOW PASSION FRUIT PEEL

Actual and coded level of variables Experimental responses

Concentration % (wt/wt) Acidic extraction method Enzymatic extraction method

Acid Enzyme Extraction Extraction Pectin yield DE Pectin Pectin yield DE Pectin
Run mixture mixture time (min) temperature (C) (%) (%) type (%) (%) type
1 0.05 (−α) 0.49 (−α) 75 (0) 60 (0) 7.51 ± 0.12 33.33 LM 6.21 ± 0.08 61.29 HM
2 0.09 (−1) 0.97 (−1) 48 (−1) 45 (−1) 5.91 ± 0.11 78.26 HM 3.54 ± 0.17 52.63 HM
3 0.09 (−1) 0.97 (−1) 102 (1) 45 (−1) 3.76 ± 0.06 72.34 HM 4.13 ± 0.12 85.71 HM
4 0.09 (−1) 0.97 (−1) 102 (1) 75 (1) 6.62 ± 0.06 78.57 HM 6.55 ± 0.06 74.19 HM
5 0.09 (−1) 0.97 (−1) 48 (−1) 75 (1) 5.55 ± 0.08 59.18 HM 4.43 ± 0.02 68.42 HM
6a 0.14 (0) 1.68 (0) 75 (0) 60 (0) 4.25 ± 0.05 77.78 HM 6.70 ± 0.09 86.96 HM
7a 0.14 (0) 1.68 (0) 75 (0) 60 (0) 5.27 ± 0.05 72.00 HM 5.69 ± 0.13 81.82 HM
8a 0.14 (0) 1.68 (0) 75 (0) 60 (0) 3.91 ± 0.02 71.11 HM 5.36 ± 0.04 80.49 HM
9a 0.14 (0) 1.68 (0) 75 (0) 60 (0) 4.04 ± 0.04 70.91 HM 5.73 ± 0.07 83.67 HM
10a 0.14 (0) 1.68 (0) 75 (0) 60 (0) 4.43 ± 0.07 73.33 HM 4.97 ± 0.04 78.00 HM
11 0.14 (0) 1.68 (0) 75 (0) 35 (−α) 3.68 ± 0.03 77.08 HM 5.09 ± 0.02 61.54 HM
12a 0.14 (0) 1.68 (0) 75 (0) 60 (0) 4.18 ± 0.03 72.50 HM 5.55 ± 0.04 82.05 HM
13 0.14 (0) 1.68 (0) 75 (0) 85 (α) 5.76 ± 0.04 78.43 HM 2.59 ± 0.02 73.53 HM
14 0.14 (0) 1.68 (0) 120 (α) 60 (0) 7.71 ± 0.13 60.38 HM 9.17 ± 0.21 80.00 HM
15 0.14 (0) 1.68 (0) 30 (−α) 60 (0) 4.02 ± 0.06 68.75 HM 5.70 ± 0.06 85.00 HM
16 0.19 (1) 2.38 (1) 48 (−1) 45 (−1) 3.84 ± 0.03 67.21 HM 6.42 ± 0.09 77.78 HM
17 0.19 (1) 2.38 (1) 48 (−1) 75 (1) 6.12 ± 0.11 78.38 HM 4.11 ± 0.16 80.00 HM
18 0.19 (1) 2.38 (1) 102 (1) 75 (1) 6.58 ± 0.12 67.69 HM 6.87 ± 0.04 82.50 HM
19 0.19 (1) 2.38 (1) 102 (1) 45 (−1) 5.05 ± 0.12 62.50 HM 6.37 ± 0.02 80.00 HM
20 0.21 (α) 2.84 (α) 75 (0) 60 (0) 4.62 ± 0.08 58.33 HM 3.63 ± 0.01 69.57 HM
a
Central point.

Journal of Food Process Engineering •• (2015) ••–•• © 2015 Wiley Periodicals, Inc. 3
ACIDIC AND ENZYMATIC PECTIN EXTRACTION FROM FRUIT PEELS
3 3 3 12
Y = β0 + ∑ βi Xi + ∑ βii Xi2 + ∑ βij Xi X j (3)
i =1 i =1 i =1
i< j
Y indicates the estimated response, β0, βi, βii and βij are
constant coefficients. X1, X2 and X3 are the uncoded values
of the independent variables of extractor concentration,
extraction time and temperature, respectively.
Pectin Morphology by Scanning Electron
Microscope (SEM)
The SEM (S-3400N, Hitachi, Tokyo, Japan) images were
taken to elucidate the morphological changes of optimally
extracted pectin samples by acidic and enzymatic extraction
methods. Wet pectin sample was the crude pectin and dry
pectin sample was the crude pectin dried using conventional
oven at 65C until a constant weight was achieved. Pectin
samples were mounted on aluminum studs and coated with
gold in a vacuum before being observed using a SEM.
Gel Formation and Viscosity
The gel formation of the pectin was tested using four condi-
tions, i.e., addition of water as control, and using solutions
of calcium chloride, sugar or mixture of sugar and calcium
chloride (Herbstreith and Fox Corporate Group 2005). 0.2 g
of dry pectin was dissolved in 20 mL of distilled water. The
solutions’ pH was kept around 3 by adding 0.1 N citric acid.
15 g sugar, 1.5 g calcium chloride and 15 g sugar plus 1.5 g
calcium chloride were added, respectively. The mixture was
heated and stirred constantly until bubbles formed around
the edge before boiling for a minute.
Viscosity of the passion fruit pectin gel was investigated
using a controlled-stress rheometer (ARG2, TA Instru-
ments, New Castle, U.S.A.) with a 60 mm diameter cone
plate at 0.3 mm gap. The gel samples prepared under four
conditions from the two extraction methods were com-
pared. Samples for the rheological tests were subjected to a
flow test by linearly increasing rates from 0 to 100 s-1 at 25C
over a duration of 2 min with 6-s intervals. From the 20
data collected, the average viscosity, η was calculated. The
stress, σ versus strain rate, γ curve was fitted with the
Power law model, σ = kγ n to obtain flow behavior index, n.
RESULTS AND DISCUSSION
Effects of Concentration, Extraction Time
and Temperature on Pectin Yield
The extracted pectin yields obtained using acid and enzyme
extraction methods are presented in Table 1. The acid
extracted pectin yield ranged from 3.68 to 7.71%, while the
4 Journal of Food Process Engineering •• (2015) ••–•• © 2015 Wiley Periodicals, Inc.
S.Q. LIEW ET AL.
10
7.51
Pecn yield (%)
8
6.21
5.66
6 4.62
4.73
3.63
4
2
0
0.05, 0.49 0.14, 1.68 0.21, 2.84
Concentraon(%-W/W)
Acidic extracon Enzymac extracon
FIG. 1. EFFECT OF CONCENTRATION ON PECTIN YIELD FROM
PASSION FRUIT PEEL AT CENTER POINT 75 MIN AND 60C
enzyme extracted pectin yield ranged from 2.59 to 9.17%.
The acid extracted pectin DE ranged from 33.33 to 78.57%,
while the enzyme extracted DE ranged from 52.63 to
86.96%. The enzymatic extraction method gave the highest
pectin yield of 9.17% with the DE of 86.96% compared
with the acidic extraction method.
Figure 1 shows the effect of extractor concentration on
pectin yield. Within the levels studied for the enzymatic
extraction method, pectin extraction yield decreased signifi-
cantly as concentration of enzymatic solution increased,
which suggests that a minimal concentration of enzymatic
solution is preferable for pectin extraction. Above concen-
tration of 1.68% (w/w), pectin yield decreased more rapidly,
which is probably caused by the effect of supersaturated or
over substrate exposure. At supersaturated or over substrate
exposure of enzyme concentration, excess enzyme hardly
played a role and led to the decrease of pectin yield.
Donaghy and McKay’s (1994) results were consistent with
this finding as he also found lower enzyme concentrations
from 0.6 to 3.0 U increased pectin yield. Similarly, with
acidic extraction method, pectin yield also decreased with
acid concentration increase from 0.05 to 0.21% (w/w). This
observation was similar to those obtained by Kulkarni and
Vijayanand (2010) and Kalapathy and Proctor (2001).
However, Yapo et al. (2007) and Levigne et al. (2002) found
otherwise. They found that pectin yields increased with
increase of sulfuric and nitric acids concentration. This con-
flicting trend is most probably because of the mineral acid
used for pectin extraction.
Figure 2 shows the effect of time on pectin yield where
the highest pectin yield was obtained at the longest extrac-
tion time of 120 min for both the enzymatic and acidic
extraction methods. For the enzymatic extraction, enzyme
takes time to interact with the cellulose and galactan side
chain of pectin. As such, a longer time is needed for the
Celluclast 1.5 L to digest the cellulose in plant wall more
completely for more pectin to be extracted out. For the
S.Q. LIEW ET AL. ACIDIC AND ENZYMATIC PECTIN EXTRACTION FROM FRUIT PEELS

12 acidic extraction, carbohydrate polymer needs heat to

9.17
soften its structure for pectin extraction and longer extrac-
10
7.71 tion time allows more reaction opportunity.
Pecn yield (%)

8 The highest pectin yield was obtained at 60C for the

6
5.70 enzymatic extraction method while for the acidic extraction
4.77 5.15
4.02 method was 85C (Fig. 3). The Celluclast enzymes are pro-
4 teins that are thermally sensitive thus requiring a lower tem-
2
perature than the acid method where higher temperature is
useful in increasing activation energy and reaction rate (Yu
0 and Sun 2013). A high extraction temperature can cause
30 min 75 min 120 min
enzyme to loose its function or cause enzyme inactivation.
Acidic extracon Enzymac extracon It is clear that heat is good for pectin yield as temperature
FIG. 2. EFFECT OF EXTRACTION TIME ON PECTIN YIELD FROM
increase from 35 to 85C gave significant pectin increase.
PASSION FRUIT PEEL AT CENTER POINT CONCENTRATION OF 0.14% Higher temperature accelerates acidic hydrolysis of pectin
FOR (ACIDIC) AND 1.68% FOR (ENZYMATIC) W/W AND 60C sugar side chains, which further increase the pectin yields
(Chan and Choo 2013).
The DE of more than 50% can be considered as high
methoxyl pectin. In general, pectin extracted from passion
12
fruit contains high methoxyl group, which is similar to the
10 result found by Pinheiro et al. (2008) and Kliemann et al.
(2009). Specifically, pectin obtained by enzymatic extraction
Pecn yield (%)

8
exhibited higher DE of 86.96% than acidic extraction
5.87 5.76
6 5.09 4.99 having DE of 78.57%. Similar result was obtained by Min
3.68 et al. (2011) and Lim et al. (2012). The HM pectin gives the
4
2.59 advantages in making high sugar products like jelly and jam
2 because its pectin molecule allows additional links of sugar
molecule to form a gel network (Herbstreith and Fox
0
35°C 60°C 85°C
Corporate Group 2005).
Acidic extracon Enzymac Extracon
Model Selection and Verification of the
FIG. 3. EFFECT OF EXTRACTION TEMPERATURE ON PECTIN YIELD Pectin Yield and DE
FROM PASSION FRUIT PEEL AT CENTER POINT CONCENTRATION OF
0.14% FOR (ACIDIC) AND 1.68% FOR (ENZYMATIC) W/W AND Table 2 lists all regression coefficients (C) and significant
75 MIN levels (p) of the independent variables of concentration

TABLE 2. REGRESSION COEFFICIENTS AND PROBABILITY VALUES FOR EACH RESPONSE

Pectin yield DE
Acid extraction Enzymatic extraction Acid extraction Enzymatic extraction
Term C P C P C P C P
Constant 17.0888 0.0000 −3.51381 0.0000 109.1900 0.0000 −100.1545 0.0000
X1 −0.1773 0.1784 2.66672 0.8468 1.3235 0.3518 26.6114 0.0436**
X2 −0.1630 0.0828* −0.17831 0.0185** 0.1650 0.6074 0.7814 0.1585
X3 −0.0851 0.0193** 0.34588 0.5178 −2.9197 0.8519 3.1270 0.2350
X1.X2 0.0005 0.3337 −0.00001 0.9992 −0.0050 0.2398 −0.1073 0.0820*
X1.X3 0.0004 0.6386 −0.02898 0.1256 0.0092 0.2347 0.0025 0.9802
X2.X3 0.0008 0.3833 0.00136 0.1874 0.0061 0.4222 −0.0085 0.1568
X1.X1 0.0008 0.0480** −0.12687 0.3492 −0.0109 0.0047*** −2.2995 0.0114**
X2.X2 0.0007 0.0761* 0.00085 0.0591* −0.0016 0.5977 0.0013 0.5747
X3.X3 0.0004 0.7281 −0.00300 0.0426** 0.0158 0.1357 −0.0196 0.0249**

*P < 0.1,
**P < 0.05,
***P < 0.005, denoting different significant effect.
X1, concentration; X2, time; X3, temperature.

Journal of Food Process Engineering •• (2015) ••–•• © 2015 Wiley Periodicals, Inc. 5
ACIDIC AND ENZYMATIC PECTIN EXTRACTION FROM FRUIT PEELS S.Q. LIEW ET AL.

10 10
9
a R2=0.705 9 b R2=0.691
Predicted response

Predicted response
8 8
7 7
6 6
5 5
4 4
3 3
2 2
2 4 6 8 10 2 4 6 8 10
Observed response Observed response

90 90
c d
80 80
Predicted response

Predicted response

70 70

60 60

50 50
FIG. 4. CORRELATION OF PREDICTED
40 R2=0.715 40 R2=0.760 RESPONSES VERSUS OBSERVED RESPONSES,
(A) ACIDIC EXTRACTED PECTIN YIELD, (B)
30 30 ENZYMATIC EXTRACTED PECTIN YIELD, (C)
30 50 70 90 30 50 70 90 ACIDIC EXTRACTED PECTIN DE AND (D)
Observed response Observed response ENZYMATIC EXTRACTED PECTIN DE

(X1), extraction time (X2) and temperature (X3) on pectin
yield and DE. For acidic extraction, the extraction tempera-
ture (P < 0.05) gave most significant effect to pectin yield
followed by extraction time (P < 0.1) while DE was most
affected by two-level interaction effects of acid concentra-
tion (P < 0.005). For the enzymatic extraction, pectin yield
was highly affected by extraction time (P < 0.05) and the DE
was affected by enzyme concentration (P < 0.05). The
pectin yield and DE for passion fruit peels are well
described in the quadratic regression models as given in
Eqs. (7)–(10) with R2 of 0.70, 0.72, 0.69 and 0.76:
Yay = 17.0888 − 0.1773 X1 − 0.1630 X 2 − 0.0851X 3
+ 0.0005 X1 X 2 + 0.0004 X1 X 3 + 0.0008 X 2 X 3
+ 0.0008 X12 + 0.0007 X 22 − 0.0004 X 32
Yey = −3.51381 + 2.66672 X1 − 0.17831X 2 + 0.34588 X 3
− 0.00001X1 X 2 − 0.02898 X1 X 3 + 0.00136 X 2 X 3 (8)
− 0.12687 X12 − 0.00085 X 22 − 0.00300 X 32
Yad = 109.1900 + 1.3235 X1 + 0.1650 X 2 − 2.9197 X 3
− 0.0050 X1 X 2 + 0.0092 X1 X 3 + 0.0061X 2 X 3
− 0.0109 X12 − 0.0016 X 22 + 0.0158 X 32
Yed = −100.1545 + 26.6114 X1 + 0.7814 X 2 + 3.1270 X 3
− 0.1073 X1 X 2 + 0.0025 X1 X 3 − 0.0085 X 2 X 3 (10)
− 2.2995 X1 + 0.0013 X 2 − 0.0196 X 3
2 2 2
Where, Yay and Yey are the yields and Y2p and Y2d are the
DE of the acidic and enzymatic extracted pectins, respec-
tively. The correlations between the observed and predicted
responses are presented in (Fig. 4). The optimum condi-
tions suggested for acidic extraction were citric acid concen-
tration of 0.19% (w/w), 102 min at 75C, which gave 7.16%
pectin yield and 71.02% of DE. For the enzymatic extrac-
tion, the optimum conditions suggested by the software
were Celluclast concentration of 1.67% (w/w), 102 min at
61.11C, which gave 7.12% pectin yield and 85.45% of DE.
The experimental values of pectin yield and DE at optimum
conditions obtained for acidic extraction were 7.52 and
73.47%, respectively, and 7.74 and 83.68% for enzymatic.
(7) The percentage error calculated were 5.03 and 8.71% for
pectin yield while for DE were 3.45 and 2.07%, for the
acidic and enzymatic extraction, respectively. The percent-
age error determined was lower than 10% suggesting
adequacy of the response surface models (McLaughlin and
Magee 1998) and good fit.
Figure 5a–c shows the surface plots of pectin yields as a
function of acidic concentration, time and temperature
(9) where extraction temperature has the most significant
effect, followed by time, and acid concentration had least
effect on pectin yield. Increase in the extraction temperature
promoted pectin yield increase from 4.8 to 7.2%. Time
increase promoted pectin yield increase from 3.0 to 6.8%.
This is in general agreement with the studies of Kulkarni

6 Journal of Food Process Engineering •• (2015) ••–•• © 2015 Wiley Periodicals, Inc.
S.Q. LIEW ET AL. ACIDIC AND ENZYMATIC PECTIN EXTRACTION FROM FRUIT PEELS

FIG. 5. EFFECT ON PECTIN YIELD (A) TIME AND CITRIC ACID CONCENTRATION, (B) TEMPERATURE AND CITRIC ACID CONCENTRATION, (C)
TEMPERATURE AND TIME FOR ACIDIC EXTRACTION, (D) TIME AND CELLUCLAST CONCENTRATION, (E) TEMPERATURE AND CELLUCLAST
CONCENTRATION, (F) TEMPERATURE AND TIME FOR ENZYMATIC EXTRACTION; EFFECTS ON DE (G) TIME AND CITRIC ACID CONCENTRATION, (H)
TEMPERATURE AND CITRIC ACID CONCENTRATION, (I) TEMPERATURE AND TIME FOR ACIDIC EXTRACTION, (J) TIME AND CELLUCLAST
CONCENTRATION, (K) TEMPERATURE AND CELLUCLAST CONCENTRATION, (L) TEMPERATURE AND TIME FOR ENZYMATIC EXTRACTION

Journal of Food Process Engineering •• (2015) ••–•• © 2015 Wiley Periodicals, Inc. 7
ACIDIC AND ENZYMATIC PECTIN EXTRACTION FROM FRUIT PEELS
FIG. 6. SCANNING ELECTRON MICROGRAPH OF PASSION FRUIT PEEL
POWDER
and Vijayanand (2010), Panchev et al. (1994), Gan and
Latiff (2011), who reported that by increasing either tem-
perature or time of extraction, yield of pectin is maximized.
Kim et al. (2004) also found that temperature was the main
parameter influencing the pectin recovery from mandarin,
using temperatures between 30 and 90C. The surface plots
for the enzymatic extraction (Fig. 5d–f) shows that extrac-
tion time has the most significant effect where pectin yield
increased quite proportionally from 4 to 7.2% with the
increase of time from 48 to 102 min.
The effects of acid or enzyme concentration, time and
temperature on the DE (Fig. 5g–l) indicate that concentra-
tion has the most significant effect on DE. This in agree-
ment with the studies of Pinheiro et al. (2008), Kulkarni
and Vijayanand (2010) who reported that concentration
have most significant effect on DE of pectins from passion
fruit peel.
Structure of Peel and Extracted Pectin
Figure 6 shows the SEM image of passion fruit peel powder,
which is flakey in shape, split and compact. Figure 7a shows
that the wet acidic extracted pectin has nanostructure,
which was smooth and compact with little wrinkle on the
surface. Figure 7b shows that the structure of wet enzymatic
extracted pectin was unique in shape, smooth and has more
wrinkle. Figure 7c and d are dry form pectin structure
extracted by acidic and enzymatic extraction, respectively.
With the effect of microwave heating, small bubbles grow
on the surface of pectin. Comparing this to Fig. 7c with d,
the amount of bubble on the enzymatic extracted pectin
surface was lesser.
8 Journal of Food Process Engineering •• (2015) ••–•• © 2015 Wiley Periodicals, Inc.
S.Q. LIEW ET AL.
Viscosity Analysis of Pectin Gel
The gel formed from the optimally extracted pectin was
governed by its DE. With DE values greater than 50%, the
pectin samples are defined as HM pectin where there are
two decisive factors that initiate gel formation, i.e., sugar
and low pH condition (Urias-Orona et al. 2010). Figure 8
shows the viscosity of pectin gel formed under the different
extraction methods and different gelation combinations.
Gel formed by the combinations of water, calcium chloride
and sugar (WCS) registered the highest average viscosity
values of 1.462 Pa·s and 1.183 Pa·s, followed by gel formed
by the combinations of “WS”, “WC” and “W”. Among the
materials added, sugar gave the most influence on the vis-
cosity of pectin gel. This could be better explained by the
dehydrating effect of sugar on the pectin molecules where
sugar facilitates the approach of the polymer chains and
enables cross linkage of the hydrogen bridges, thus increas-
ing gel viscosity (Herbstreith and Fox Corporate Group
2005). The results also suggested that calcium chloride
effects on HM pectin gel viscosity were less. This work is in
agreement with Herbstreith and Fox Corporate Group
(2005) and Urias-Orona et al. (2010) who found that the
calcium chloride has more significant effect on the LM
pectin (Urias-Orona et al. 2010). As the flow behavior
indices from the Power law model fitting obtained were less
than 1 (Table 3), the HM pectin gel is classified as non-
Newtonian, pseudoplastic liquid.
CONCLUSIONS
The acidic and enzymatic extraction methods gave the
pectin yields of 7.71 and 9.17% with DE of 78.57 and
86.96%. The extraction time was the most significant
extraction factor for the pectin extracted through the enzy-
matic method while extraction temperature was the most
important extraction factor for the acidic extracted pectin
from passion fruit peels in terms of its yield. The extraction
concentration had highly significant effects on the pectin
DE. The DE of extracted pectin across the varied extraction
factors were quite consistent at about 68.90 and 76.26%,
which is classified as the HM pectin. The SEM images of
passion fruit pectin provided structural information of wet
passion fruit pectin, which is unique in shape following dif-
ferences in extraction process. The pectin gel produced
exhibited a pseudoplastic behavior.
ACKNOWLEDGMENT
The authors gratefully acknowledge the Graduate Research
Fellowship (GRF) Scheme sponsored by Universiti Putra
Malaysia.
 S.Q. LIEW ET AL. ACIDIC AND ENZYMATIC PECTIN EXTRACTION FROM FRUIT PEELS

FIG. 7. SCANNING ELECTRON MICROGRAPH OF (A) ACIDIC EXTRACTED WET PECTIN, (B) ENZYME EXTRACTED WET PECTIN, (C) ACIDIC
EXTRACTED DRY PECTIN, (D) ENZYME EXTRACTED DRY PECTIN

TABLE 3. AVERAGE VISCOSITY OF PECTIN GELS

1.6 1.462
Average viscosity, Flow behavior
Average viscosity, m (Pa·s)

1.183 Method μ (Pa·s) index, n

1.2 1.073
1.048 Acidic W 0.082 0.745
0.839 WC 0.481 0.748
0.8
WS 1.073 0.902
0.481 WCS 1.462 0.988
0.4 Enzymatic W 0.107 0.697
0.107 WC 0.839 0.723
0.082
0 WS 1.048 0.845
W WC WS WCS WCS 1.183 0.879
Acidic Enzymac

FIG. 8. COMPARING THE VISCOSITY OF PECTIN GEL PRODUCED BY REFERENCES

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SOLUTION, WS (IV) WATER, SUGAR AND CALCIUM CHLORIDE
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