ARTICLE IN PRESS

Food Hydrocolloids 23 (2009) 434– 440

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Food Hydrocolloids
journal homepage: www.elsevier.com/locate/foodhyd

Enzymatic hydrolysis of granular native and mildly heat-treated tapioca

and sweet potato starches at sub-gelatinization temperature
Y.N. Shariffa a, A.A. Karim a,, A. Fazilah a, I.S.M. Zaidul b
a
Food Biopolymer Science Research Group, School of Industrial Technology, Universiti Sains Malaysia, 11800 Minden, Penang, Malaysia
b
Department of Food Science, Faculty of Food Science and Technology, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor DE, Malaysia

a r t i c l e in f o a b s t r a c t

Article history: The effect of mild heat treatment (below gelatinization temperature) towards the susceptibility of
Received 21 December 2007 granular starch to enzymatic hydrolysis was investigated. Tapioca and sweet potato starches were
Accepted 12 March 2008 subjected to enzymatic hydrolysis with a mixture of fungal a-amylase and glucoamylase at 35 1C for
24 h. Starches were hydrolyzed in native (granular) state and after heat treatment below gelatinization
Keywords: temperature (60 1C for 30 min). The dextrose equivalent (DE) value of heat-treated starch increased
Starch significantly compared to native starch, i.e., 36–50% and 27–34% for tapioca and sweet potato starch,
Granule respectively. Scanning electron microscopy examination showed that enzymatic erosion occurred
Enzyme hydrolysis
mainly at the surface of starch granules. Hydrolyzed heat-treated starch exhibited rougher surface and
a-Amylase
porous granules compared to native starch. X-ray analysis suggested that enzymatic erosion
Glucoamylase
Reducing sugar preferentially occurred in amorphous areas of the granules. The amylose content, swelling power and
solubility showed insignificant increase for both starches. Evidently, heating treatment below
gelatinization temperature was effective in enhancing the degree of hydrolysis of granular starch.
& 2008 Elsevier Ltd. All rights reserved.

1. Introduction Industrial production of starch hydrolysis products such as

Starch is a mixture of two polysaccharides, the linear molecule
of amylose, which consists of polymers of glucose, and amylo-
pectin, a highly branched molecule. Starch can be sub-divided into
cereal, legumes, palm and tuber or root starches. Tapioca (Manihot
esculenta Crantz) and sweet potato (Ipomea batatas Lam) starches
are examples of starch derived from roots or tubers. Granules of
tuber and root starches are oval, although round, spherical,
polygonal and irregular shapes also exist (Lideboom, Chang, &
Tyler, 2004). Starch susceptibility to enzyme attack is influenced
by several factors such as amylose and amylopectin content (Holm
& Bjorck, 1988; Ring, Gee, Whittam, Orford, & Johnson, 1988),
particle size, crystalline structure and the presence of enzyme
inhibitors. Among these factors, granular structure is believed
to be the most important (Zhang & Oates, 1999). Among non-
cereal starches, tapioca starch has relatively higher enzyme
susceptibility than other starches such as potato and sweet
potato (Rickard, Asaoka, & Blanshard, 1991; Zhang & Oates, 1999).
According to Hizukuri et al. (1988) and Kainuma (1988), sweet
potato starch shows stronger resistance to a-amylase and
glucoamylase attack.
 Corresponding author. Tel.: +604 6533888x2221; fax: +604 6573678.
E-mail address: akarim@usm.my (A.A. Karim).
glucose syrup is typically based on acid or enzyme hydrolysis on
gelatinized starch. Conventional processes for production of
glucose syrups or bioethanol basically involved liquefaction and
saccharification process of starch. Liquefaction and saccharifica-
tion require the starch granules to be extensively gelatinized at
high temperature. This is an energy-intensive process requiring
the addition of heat energy to starch granule slurries until the
gelatinization temperature of the starch is exceeded. The whole
process requires a high-energy input, thus increasing the produc-
tion cost of starch-based products. In view of energy costs,
effective utilization of natural resources and viscosity pro-
blems, direct hydrolysis of starch below gelatinization tempera-
ture is desirable.
Typically, enzyme hydrolysis of granular starch yields low
degree of conversion to fermentable sugars. Although starch
macromolecules can be hydrolyzed in a granular state, however,
attempts to hydrolyze native granules invariably result in a slowly
and often poorly hydrolyzed product (Oates, 1997). Generally, the
action of a- and b-amylases on native starch granules is not very
effective because granules are very resistant to amylolytic
digestion and a long hydrolysis period is required for degrading
the starch (Sarikaya, Higasa, Adachi, & Mikami, 2000). With new
technological advances in recent years, however, new generation
of enzymes such as a-amylase from Aspergillus kawachi and
glucoamylase from Aspergillus niger have been discovered. These

0268-005X/$ - see front matter & 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.foodhyd.2008.03.009
 ARTICLE IN PRESS
Y.N. Shariffa et al. / Food Hydrocolloids 23 (2009) 434–440
enzymes work synergistically to hydrolyze granular starches that
can directly hydrolyze the raw starch in a single step at moderate
temperature well below the gelatinization temperature. These
enzymes have the advantages of exo-activity of glucoamylase,
which is able to drill sharp and deep pinholes, as well as the endo-
activity of a-amylase that enables widening of the pinholes. This
combination enhances the release of fermentable glucose con-
tinuously from granular starches. Franco, Preto, Ciacco, and
Tavares (1988) reported that more hydrolysis can be obtained by
hydrolyzing the starch with a-amylase together with glucoamy-
lase. Further modification can be made in order to increase the
efficiency of native starch hydrolysis. Oates (1997) suggested that
better hydrolysis of native starch can be achieved by increasing
the incubation temperature approximately to 60 1C. If the degree
of conversion of native starch could be increased further, it would
be very useful in the industrial process of fermentable sugars and
bioethanol.
The present study was designed to study the effect of mild heat
treatment (below gelatinization temperature) towards the sus-
ceptibility of granular starch to enzymatic hydrolysis.
2. Materials and methods
2.1. Materials
Tapioca and sweet potato starch were obtained from SIM
Company Sdn. Bhd. (Penang, Malaysia).
2.2. Enzyme
The commercial enzyme, STARGEN 001TM, was a product of
Genencor Internationals, B.V. (Genencor International, Palo Alto,
CA). This enzyme contains A. kawachi a-amylase expressed in
Trichoderma reesei and a glucoamylase from A. niger. The specific
gravity of the enzyme is 1.10–1.15 g/ml, optimum pH is 4.0–4.5,
recommended temperature is 20–40 1C and the minimum activity
is 456 GSHU/g (GSHU is defined as granular starch hydrolyzing
units). The enzyme’s activity was determined by reaction at 37 1C
with soluble starch (1%) that was buffered with sodium acetate
(pH 4.4). Aliquots were taken after 10 min for determining the
amount of D-glucose released. The glucose was determined by
using the dinitrosalicylic acid method (Miller, 1959). The enzyme
activity was 3736 units/g starch.
2.3. Determination of pasting temperature
The gelatinization temperature of starches was determined in
triplicate by using a Rapid ViscoTM Analyzer (Model RVA Series 4,
Newport Scientific Pvt. Ltd., Warriewood, Australia) and a
Differential Scanning Colorimeter, DSC-Q100 (TA Instruments,
Lukens Drive, New Castle, USA), equipped with a refrigerated
cooling system (RCS). For the determination by a rapid visco
analyzer (RVA), 2 g of starch sample (corrected to 14% moisture
basis) and 25 ml of distilled water were combined and stirred in
the aluminum RVA sample canister. Temperature was held at
50 1C for 1 min and then raised to 95 1C in 3.75 min, held for
2.5 min, cooled to 50 1C in 3.75 min and held for 5 min. The paddle
speed was set at 960 rpm for the first 10 s to evenly disperse the
starch slurry and reduced to 160 rpm throughout the entire
experiment. The units of viscosity were expressed as RVU.
For the determination by a differential scanning colorimeter
(DSC), 2 mg of starch sample was loaded into an aluminum
pan and distilled water was added to achieve a starch–water
suspension containing 75% water. Samples were heated from
435
30 to 130 1C at a heating rate of 10 1C/min under an oxygen-free N2
flow rate of 50 ml/min.
2.4. Mild heat treatment
The starch slurry (25% w/v) in 400 ml sodium acetate buffer
was incubated in a water bath at 60 1C for 30 min with continuous
stirring using an overhead stirrer. The temperature of the slurry
was brought down to 35 1C by immersing the starch slurry in the
water before being subjected to hydrolysis.
2.5. Starch hydrolysis
The starch slurry (25% w/v) was prepared in 400 ml sodium
acetate buffer. The enzyme (3736 units/g starch) was added
(1% w/v) into the slurry. The samples were incubated in an
incubator shaker (JEIO Tech, SI-600R, Seoul, Korea) at 35 1C with
the speed of 150 rpm. After 24 h, the hydrolysis was stopped by
adjusting the slurry to pH 1.5–1.6 with 2 M HCl and left for 2 h.
Then, the pH of starch suspensions was adjusted back to pH 5–6
by washing the starch with distilled water. The starch residues
were dried at 40 1C for 2 days.
2.6. Dextrose equivalent (DE)
The reducing sugar value was measured using the dinitrosa-
licylic acid method (Miller, 1959) to determine its dextrose
equivalent (DE). A small portion of aliquot was withdrawn from
each batch of starch slurry at various time intervals up to 24 h
hydrolysis time. The absorbance was measured at 504 nm by
using a UV/visible spectrophotometer (UV-160A, SHIMADZU,
Kyoto, Japan). Glucose was used as a standard. Each analysis
was performed in duplicate. DE was calculated as follows:
g reducing sugar expressed as glucose
DE ¼  100%
g dry solid weight
2.7. Scanning electron microscopy
The microstructure of a starch granule was viewed with a field
emission scanning electron microscope (FESEM Leo Supra 50VP,
Carl-Ziess SMT, Oberkochem, Germany). The starch granules were
stuck on aluminum specimen stubs with double-sided adhesive
tape and sputtered with a 20–30 nm gold layer using a sputter
coater [Polaron (Fisons) SC515, VG Microtech, Sussex, UK].
2.8. Swelling and solubility
Swelling power and solubility of starch were determined in
triplicate by adopting the method of Schoch (1964).
2.9. Amylose content
The amylose content of each sample and raw starch was
determined in triplicate according to the procedure described by
McGrance, Cornell, and Rix (1998) with minor modification. Pure
potato amylose and amylopectin (Sigma Chemical Company,
Steinheim) were used as the standards. The results were exp-
ressed on a dry basis. Starch (0.1 g, dry basis) was accurately
weighed and dissolved by heating in dimethyl sulfoxide for 15 min
on a hot plate at 85 1C while stirring continuously with a magnetic
stirrer bar. After the solution had dissolved, it was diluted to 25 ml
in a volumetric flask with deionized water. An aliquot (1 ml) of
this solution was diluted with 50 ml of deionized water. Five
 ARTICLE IN PRESS
436 Y.N. Shariffa et al. / Food Hydrocolloids 23 (2009) 434–440
milliliters iodine (0.0025 mol/l) in potassium iodide (0.0065 mol/l)
was added with mixing and the absorbance of this solution in a
1 cm path length glass cell read at 600 nm using a UV/visible
spectrophotometer (UV-160A, SHIMADZU, Kyoto, Japan). Samples
were left for 15 min after the addition of iodine before taking the
readings on the spectrophotometer.
2.10. X-ray diffraction
Crystallinity patterns of starch granules were examined by the
X-ray diffraction method as described by Lauro, Forssell, Suortti,
Hulleman, and Poutanen (1999). The dried starches were condi-
tioned overnight at 100% relative humidity (RH) at room
temperature. The starches were scanned by an X-ray diffract-
ometer (Diffractometer D5000, Siemens, Karlsruhe, Germany).
Diffractograms were recorded in the reflection mode in the
angular range 4–401 (2y). The Cu Ka-radiation (l 1.5406 Å),
generated at 40 kV and 30 mA, was made monochromatic using
a 15 mm Ni foil. Scattered radiation was detected using a
proportional detector.
2.11. Statistical analysis
Duncan’s least significant test was used to compare means at
the 5% significance level. Simple Pearson’s correlation and
regression analysis were evaluated using SPSS 12.0 software
(SPSS, Inc., Chicago, IL, USA).
3. Results and discussion
The gelatinization temperature of tapioca and sweet potato
starches was determined by using a RVA and DSC. Results
obtained showed that the gelatinization temperature of both
starches are higher than 60 1C (given in Table 1); therefore, the
temperature of 60 1C has been chosen to treat the starch for mild
heat treatment because it is below the gelatinization temperature
of those starches. We hypothesized that the mild heat treatment
would allow the starch granules to swell more and open up the
small pores or crevices on the granule surface, which would
facilitate the access of enzyme into the starch granules. Therefore
the mild heat treatment was expected to increase the degree of
hydrolysis of the starches. This approach was based on the
observation by Haska and Ohta (1992) who reported that
treatment of sago starch by heating to below gelatinization
temperature at lower pH resulted in an increase in the ability of
enzyme to digest sago starch granules.
3.1. Degree of hydrolysis
As shown in Fig. 1, after 24 h of hydrolysis at 35 1C, heat-treated
tapioca showed the highest DE (50%) followed by native tapioca
(36%), heat-treated sweet potato (34%) and native sweet potato
(27%). Evidently, treating the starch with mild heat before enzyme
Table 1
Gelatinization temperature of raw tapioca and sweet potato starches determined
using rapid visco analyzer (RVA) and differential scanning calorimeter (DSC)
Sample Gelatinization temperature (1C)
RVA DSC
Tapioca 70.4270.25 64.2470.22
Sweet potato 74.1870.32 66.2870.24
Fig. 1. Hydrolysis profiles of native and heat-treated tapioca and sweet potato
starches at sub-gelatinization temperature (35 1C) for 24 h. The error bar
represents71 standard deviation (n ¼ 3).
hydrolysis enhanced the degree of hydrolysis of starch signifi-
cantly (po0.05). Holding the starch at 60 1C for 30 min could
cause partially irreversible swelling of the amorphous region in
the granules, consequently allowing greater access for the enzyme
to act on the starch. It is also possible that during this time, the
slight swelling of the granule caused expansion of the naturally
present pinholes and internal cavities in starch granules, allowing
the enzyme to penetrate easily into the granules. According to
Juszczak, Fortuna, and Krok (2003), pores present on starch
surfaces could become centers of enzymatic attack. The large
increase in hydrolysis of heat-treated starch could also be
attributed to the effect of heat on the weaker areas on the starch
granule (e.g. truncated or damaged granules shown by arrows in
Fig. 2a), allowing the enzyme to degrade the starch granules more
extensively. In addition, it has been found that heat–moisture
treatment (heating starch at a relative humidity of 100%) is
effective in enhancing the adsorption of a-amylase (Kurakake,
Tachibana, Masaki, & Komaki, 1996).
3.2. Scanning electron microscopy
The susceptibility of starch granules can be classified by the
intensity and the manner by which the granules are eroded and
corroded (Gallant et al., 1982). The modes of attack on hydrolyzed
native and hydrolyzed heat-treated tapioca and sweet potato
starch were observed by using SEM). From SEM micrographs
(Fig. 2), the enzymatic erosion occurred mainly at the surface of
starch granules. Qualitatively, rough surface and eroded granules
were observed in hydrolyzed heat-treated starch (Fig. 2d and h) as
compared to hydrolyzed native starch (Fig. 2b and f), which
displayed rough surfaces with limited erosion and fewer holes for
both tapioca and sweet potato starches. Heating of starch in water
causes disruption of hydrogen bond between the polymer chain,
thereby weakening the granule, so that the enzyme can penetrate
and degrade the starch easily. This resulted in rougher and more
eroded surfaces of heat-treated hydrolyzed starch compared to
hydrolyzed native starch. At higher magnification of SEM, the
granules of hydrolyzed heat-treated tapioca and sweet potato
starches (Fig. 3c and d) appear to be composed of a substantial
number of small pores as compared to hydrolyzed native starch
(Fig. 3a and b). From this result, it is suggested that the mild heat
treatment caused the starch to swell, resulting in the enlargement
of the pinholes on the surface of the starch granules. This permits
the enzyme to penetrate into the granules more extensively and
forms pores and channels in the starch granules during hydrolysis.
This suggestion is also supported by Kurakake et al. (1996), who
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Y.N. Shariffa et al. / Food Hydrocolloids 23 (2009) 434–440 437

Fig. 2. SEM micrographs (3000  ) for (a) control native tapioca, (b) hydrolyzed native tapioca, (c) control heat-treated tapioca, (d) hydrolyzed heat-treated tapioca, (e)
control native sweet potato, (f) hydrolyzed native sweet potato, (g) control heat-treated sweet potato and (h) hydrolyzed heat-treated sweet potato starches after hydrolysis
at sub-gelatinization temperature (35 1C) for 24 h (scale bar ¼ 10 mm).

found that the amount of amylase-digestible regions in the
heat–moisture-treated starches was greater than in native
starch. Enzymes cause surface alterations and degrade the
external part of the granule by exo-corrosion and when endo-
corrosion occurs, the internal part of the granule is corroded
through small pores by which enzymes penetrate the granule
(Aggarwal & Dollimore, 1998).
It was observed that hydrolyzed tapioca showed rougher and
eroded granules (Fig. 2b and d) as compared to sweet potato
starch (Fig. 2f and h) for both native and after mild heat treatment.
Larger and more porous surface can also be observed in the
hydrolyzed heat-treated tapioca than hydrolyzed heat-treated
sweet potato starch granules (Fig. 3b and d). This is consistent
with a higher degree of hydrolysis for tapioca starch than that of
sweet potato. This result is also in accordance with a previous
study by Zhang and Oates (1999), who reported that granular
sweet potato starch is less susceptible than granular tapioca
starch to a-amylase and glucoamylase attack. The rough surface of
hydrolyzed tapioca starch might be due to the exo-corrosion
phenomenon of enzyme proceeding all over the surface. Valetu-
die, Colonna, Bouchet, and Gallant (1993) reported that pitting
occurred preferentially on the edges of the granules. Therefore,
the presence of truncatures in tapioca starch (Fig. 2a, shown by
arrows) was the weak points of the granule structure, leading to
better susceptibility. According to Gallant, Bouchet, Buléon, and
Pérez (1992), some tropical tubers have specific susceptible zones
which became pitted due to endo-corrosion. Therefore, the rough
surface of starch granules in tapioca starch might be due to these
specific zones. Juszczak et al. (2003) also reported that depres-
sions and protrusions were observed on the surface of tapioca
starch granules.
3.3. Swelling power and solubility
The swelling power and solubility of control, native and heat-
treated tapioca, and sweet potato starches after 24 h of hydrolysis
are summarized in Table 2. From the results, the swelling power
and solubility of hydrolyzed native starch were higher compared
to heat-treated hydrolyzed starch for both tapioca and sweet
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438 Y.N. Shariffa et al. / Food Hydrocolloids 23 (2009) 434–440

Fig. 3. SEM micrographs (30,000  ) for (a) hydrolyzed native tapioca starch, (b) hydrolyzed heat-treated tapioca, (c) hydrolyzed native sweet potato and (d) hydrolyzed
heat-treated sweet potato starches after hydrolysis at sub-gelatinization temperature (35 1C) for 24 h (scale bar ¼ 1 mm).

Table 2 Table 3
Swelling power and solubility of control, native and heat-treated tapioca and The amylose content of control, native and heat-treated tapioca and sweet potato
sweet potato starches after 24 h of hydrolysis at 35 1C starches after 24 h of hydrolysis at 35 1C

Sample Swelling power (g/g) Solubility (%) Sample Amylose content

Tapioca Tapioca
Control 15.94e71.08 5.04b70.67 Control 19.42a70.09
Native 16.58e70.63 6.56c70.27 Native 21.43c70.35
Control heat-treated 13.88d70.03 1.78a70.62 Control heat-treated 20.50b70.68
Heat-treated 14.61d70.32 4.54b70.41 Heat-treated 21.94cd70.62

Sweet potato Sweet potato

Control 9.89b70.49 4.89b70.38 Control 19.67a70.23
Native 11.50c70.31 6.41c70.40 Native 21.35c70.29
Control heat-treated 8.31a70.32 4.22b70.55 Control heat-treated 21.57cd70.10
Heat-treated 9.48b70.52 4.92b70.59 Heat-treated 22.22d70.09

Values followed by the same letter within the same column are not significantly Values followed by the same letter within the same column are not significantly
different (p40.05). different (p40.05).
 Mean7SD of triplicate samples.

potato starches. The decrease of swelling power of heat-treated
hydrolyzed starch might be due to the amorphous region in the
starch granules that has been extensively degraded. During the
heat treatment, the pinholes on the starch granules expanded and
made the starch granules more susceptible to enzymatic attack.
After hydrolysis, the enzymes had rendered the granules to
become porous and thus weakened the structure of the granules.
Therefore, the granules could not swell to the maximum capacity.
3.4. Amylose content
and a-1,6 linkages by a-amylase and glucoamylase during
hydrolysis. The higher swelling power for hydrolyzed native
starches could be due to the fact that intrinsic amylose (being less
degraded) was able to hold the integrity of the granule. From the
adsorption spectra of the iodine–starch complex, Kitahara,
Tanaka, Suganuma, and Nagahama (1997) observed an increase
in the blue value as the degree of hydrolysis of starch increased.
This suggestion is in accordance with our result where the
amylose content increased after the starch was treated with
enzyme.

Table 3 shows that the amylose content of hydrolyzed heat-
treated starch had increased significantly as compared to hydro-
lyzed native starch for sweet potato starch but insignificantly for
tapioca starch. It should be noted that the value of amylose
content in hydrolyzed heat-treated starch (Table 3) might come
from the intrinsic amylose and the result of degradation of a-1,4
3.5. X-ray diffraction
X-ray diffraction patterns can be used to differentiate between
native starches and it also detects changes in crystallinity brought
about by physical or chemical treatment of granular starch. X-ray
diffraction patterns of hydrolyzed native and hydrolyzed
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Y.N. Shariffa et al. / Food Hydrocolloids 23 (2009) 434–440 439

Intensity, Cps (a)

(b)

(c)

(d)

10 20 30 40 50 60 70
Diffraction angle, 2θ

Fig. 4. X-ray diffraction patterns of (a) control native tapioca, (b) hydrolyzed native tapioca, (c) control heat-treated tapioca and (d) hydrolyzed heat-treated tapioca starch
after hydrolysis at sub-gelatinization temperature (35 1C) for 24 h.

(a)
Intensity, Cps

(b)

(c)

(d)

0 10 20 30 40 50 60 70
Diffraction angle, 2θ

Fig. 5. X-ray diffraction patterns of (a) control native sweet potato, (b) hydrolyzed native sweet potato, (c) control heat-treated sweet potato and (d) hydrolyzed heat-
treated sweet potato starch after hydrolysis at sub-gelatinization temperature (35 1C) for 24 h.

heat-treated tapioca and sweet potato starch are presented in
Figs. 4 and 5, respectively. Both hydrolyzed heat-treated and
hydrolyzed native tapioca and sweet potato starch exhibited
typical A-type patterns with strong peaks at 2y about 151, 171, 181
and 231 for tapioca and 151, 171, 221 and 231 for sweet potato.
Franco et al. (1988) also observed A-type patterns in tapioca
starch that has been treated with a-amylase and amyloglucosidase
at 37 1C. Moorthy (2002) reported that tapioca and sweet potato
starches possesses A-type patterns with three major peaks at
2y ¼ 15.31, 17.11 and 23.51. Even though the starches show similar
X-ray patterns, the hydrolyzed heat-treated starch appeared to
show a more crystalline character. The peaks appeared to be
sharper and well defined as compared to hydrolyzed native starch.
This indicated that the amorphous region of heat-treated starch
has been hydrolyzed extensively compared to native starch.
Sharper X-ray diffraction peaks indicated that the amorphous
parts of the starch granules had been disrupted (Colonna, Buléon,
& Mercier, 1981). This result was also in accordance with a
previous study by Gallant, Derrien, Aumaitre, and Guilbot (1973)
who mentioned that the amylolysis primarily occurs in the
amorphous regions of the starch granules.
4. Conclusion
The amylolytic enzymes were able to hydrolyze granular starch
at a sub-gelatinization temperature (35 1C) with a high DE value.
The degree of hydrolysis of starch was enhanced by pre-treating
the starch with mild heat below its gelatinization temperature
before being subjected to enzyme hydrolysis. In a granular state,
tapioca starch was found to be more susceptible to enzyme attack
compared to sweet potato starch with or without heat treatment.
Acknowledgements
The authors gratefully acknowledge Genencor International,
Palo Alto, CA, for providing the enzyme STARGEN 001 used in this
 ARTICLE IN PRESS
440 Y.N. Shariffa et al. / Food Hydrocolloids 23 (2009) 434–440
study. This work was funded by Universiti Sains Malaysia Short
Term Research Grant (304/PTEKIND/637081). One of the authors
(N.Y. Shariffa) acknowledges the scholarship by the National
Science Fellowship.
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