Electronic Journal of Polish Agricultural Universities (EJPAU) founded by all Polish
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2007
Volume 10
Issue 3
Topic:
Food Science and Technology
ELECTRONIC
JOURNAL OF
POLISH
AGRICULTURAL
UNIVERSITIES
Copyright © Wydawnictwo Akademii Rolniczej we Wroclawiu, ISSN 1505-0297
Le Than J. , Błaszczak W. , Lewandowicz G. 2007. DIGESTIBILITY VS STRUCTURE OF
FOOD GRADE MODIFIED STARCHES, EJPAU 10(3), #10.
Available Online: http://www.ejpau.media.pl/volume10/issue3/art-10.html
DIGESTIBILITY VS STRUCTURE OF FOOD GRADE
MODIFIED STARCHES
Joanna Le Than1, Wioletta Błaszczak2, Grażyna Lewandowicz1
1
Department of Biotechnology and Food Microbiology, The August Cieszkowski Agricultural
University of Poznan, Poland
2
Institute of Animal Reproduction and Food Research of Polish Academy of Sciences,
Division of Food Science, Olsztyn, Poland
ABSTRACT
Digestibility of food grade modified starches were examined by the controlled enzymatic hydrolysis with the
mixture of pancreatic alpha-amylase and glucoamylase. The structure and physicochemical properties of the
experimental starch samples were examined by the Brabender rheological method, light microscopy, and X-ray
diffractometry. It was found that investigated food grade modified starches revealed reduced digestibility-up to
about 10%. The extent of decrease of digestibility depends on the degree of substitution of modified starches, but
this correlation is not linear. Granular food grade modified starches reveal unchanged crystal structure, identical
as native starch. The modification of starch changes its physicochemical properties i.e. pasting profile,
gelatinisation temperature and solubility in water, however these changes do not correlate with digestibility.
Changes in conformation of starch macromolecules in solution, caused by incorporation of modifying groups,
hinder enzymatic action of amylases and cause a drop in digestibility of starch.
Key words: modified starches, structure, digestibility.
INTRODUCTION
In diet, starch is primarily considered as a source of energy and it mainly determines its
nutritive importance. However, in many foodstuffs it escapes complete digestion by amylases
of alimentary tract, which implicates their nutritional value. From this point of view, starch
can be classified into three basic groups: rapidly digestible starch (RDS), slowly digestible
starch (SDS), and resistant starch (RS) [7].
The digestibility of native starch depends on its botanical origin and significantly rises after
gelatinisation [9, 15]. The chemical modification of starch may ultimately affect its rate and
extent of digestion in the small intestine [17, 18]. It is especially important when creating so
called functional food designed for groups of people suffering from diabetes or metabolic
syndromes. Up to now it has been stated that the oxidation or dextrinisation of starch, its
substitution with hydroxypropyl, acetyl or octenylsuccinate groups as well as crosslinking
cause the decrease of digestibility [8, 17, 18, 19]. Unfortunately, the comparison of the data
published by various research groups is difficult, due to the fact that obtained results are
strongly influenced by the method applied for the determination of digestibility. The
differences could be the effect of application merely different alpha-amylases (rabbit or
human) [9]. Generally, all in vitro methods of the estimation of digestibility of starch consist
in the simulation of digestion process in the human alimentary tract. However all these
simulations are very imperfect due to the fact that only the activities of alphaamylase and
glukoamylase are taken into consideration, and other enzymes occurring in enteron are
omitted. As a source of enzymes are proposed not only pure enzymes – alpha-amylases and
glucoamylases but also mixtures such as pancreatin [3, 8]. Older methods, for the digestion of
starch, employed only one enzyme i.e. alpha-amylase. The amount of released reducing
saccharides were determined spectrophotometrically with 3,5-dinitrosalicylicacid. The extent
of the hydrolysis was expressed as a ratio of the amounts of released maltose to analysed
starch [9, 19]. Essential progress was made during the research programme of the European
Communitiy Commission of regarding resistant starch [2]. As a result of the work of
EURESTA group the methods for the determination of rapid digestible starch (RDS), slow
digestible starch (SDS) and resistant starch (RS) were proposed. The principle of the method
base on hydrolysis of starch with the mixture of pancreatic alpha-amylase and glucoamylase
followed by the measurement of the released glucose using glucose oxidase [7]. Rapidly
digested and slowly digested starch are defined as the amount of glucose released after
hydrolysis at the temperature of 37°C for a period of 20 and 100 min respectively. Resistant
starch is the starch not hydrolysed after 120 min of incubation. The newest standards for the
determination of resistant starch postulate the hydrolysis with the mixture of pancreatic alpha-
amylase and glucoamylase at the temperature of 37°C for the time of 16 h [1, 4, 5].
The chemical modification of starch for food application is precisely restricted not only in
terms of type of employed chemical reactions but also the extent of changes in starch
macromolecules [6]. The above mentioned restrictions are recommended by Joint FAO/WHO
Expert Committee on Food Additives (JECFA) with the aim of protecting consumers against
intake of objectionable food. However changes in molecular and supermolecular structure of
starch, caused by chemical modification and changing rate of digestion could be
advantageous, for example in manufacturing functional food designed for the diabetics.
The aim of the work was to contribute to the understanding of the relationship between
structure, physicochemical properties and the digestibility of food grade modified starches.
Native starch was used as a reference material.
MATERIALS AND METHODS
Materials
Native potato starch Superior Standard as well as food grade modified starches listed below
kindly provided by WPPZ Luboń SA. (Poland) were studied i.e.:
bleached starch E 1403 – “Skrobia Budyniowa”
oxidised starch E 1404 – “Skrobia Żelujšca”
distarch phosphate E 1412 – “Luboterm”
acetylated distarch phosphate E 1414 – “Creamix 1050”
acetylated starch E 1420 – “Zagęstnik AC”
acetylated distarch adipate E 1422 – “Zagęstnik AD”
X-ray diffractometry
X-ray diffractometry was carried out with a TUR 62 Carl Zeiss X-ray diffractometer under
the following conditions: X-ray tube CuKα (Ni filter); voltage 30 kV; current 15 mA;
scanning from Θ = 2° to 18°. To avoid the influence of relative humidity on relative
crystallinity, starch samples were placed in a desiccator and conditioned in the atmosphere of
relative humidity of 92% for 48 h. The desiccator was filled with a sodium carbonate
saturated water solution.
Microscopic examination
The starch samples were prepared for light microscopy examination using the smear method.
The 8% starch suspension were heated at 90°C for 15 min. A drop of paste was placed on a
microscope slide and, on cooling, the smear was stained with iodine (I2 in KI) and studied
using an Olympus BX60 light microscope.
Viscosity measurements
The course of gelatinisation was monitored with a Brabender viscograph under the following
conditions: measuring cartridge 0.07 Nm; heating/cooling rate 1.5° C/min; thermostating 30
min.
Chemical composition
The carboxyl as well as acetyl groups content of starch derivatives were determined according
to Joint FAO/WHO Expert Committee on Food Additives (JECFA) recommendation [6].
Carboxyl groups determination involves leaching of modified starch with mineral acid to
convert carboxyl salts to the acid form. Cations and excess acid are removed by washing with
water. The washed sample was gelatinised in water and titrated with standard alkali. Acetyl
groups content determination consists in hydrolysing them with sodium hydroxide and
titration of the excess of alkali with hydrochloric acid. Results compiled on tables are means
of four measurements with standard deviation considered.
Digestibility
The rate of digestion of starch was determined by its hydrolysis with the mixture of pancreatic
alpha-amylase and glucoamylase at the temperature of 37°C, at particular periods of time,
followed by the measurement of the released glucose using glucose oxidase. Porcine
pancreatic alpha-amylase type VI-B (Sigma) as well as glucoamylase AMG 300L
(Novozymes) were used for the analyses. The amount of released glucose was determined
colorimetrically at the λ = 500 nm using Liquick Cor-Glucose diagnostic kit (Cormay,
Poland). Four replicates were made for each probe and standard deviation were calculated.
RESULTS AND DISCUSSION
Native potato starch, one of the most important sources of energy in human nutrition, was, as
it could be expected, fully digested after 16 h (Table 1). It should be emphasised that almost
full digestion of native starch was achieved already after 8 h. Similar digestibility, as well as
digestion rate profile, revealed distarch phosphate E 1412 preparation. Distarch phosphate, as
other cross-linked starches, is obtained as a result of very fine changes in molecular structure
of starch i.e. on cross-linked bond occurs per 5001000 anhydroglucose units [13, 14]. In
other words, it could be expressed as a degree of substitution DS = 0.001 – 0.002. Starch
modified by using other methods i.e. obtained by incorporation of carboxyl or acetyl groups
revealed reduced digestibility-up to about 10%. The extent of this decrease was correlated
with the degree of substitution of modified starch, but this correlation seems to be not linear.
Slightly oxidised E 1403 starch containing 0.04 of carboxyl groups (what could be also
expressed as DS = 0.0014) revealed the decrease of digestibility of about 5.6%. The increase
of carboxyl groups content up to 0.1 % (DS = 0.036) caused the decrease of digestibility only
of additional 1%. The lowest digestibility revealed acetylated starch E 1420 containing 1.7%
of acetyl groups (DS = 0.065). Acetylated distarch adipate E 1422 containing 1.2% of acetyl
groups (DS = 0.046) showed digestibility higher than E 1420 of about 2%. Acetylated
distarch phosphate E 1414 containing the lowest amount of acetyl groups 0.8% (DS = 0.030)
revealed relatively high digestibility, even higher than oxidised starches.
Table 1. The rate of digestion of chemically modified starches and modifying groups content
Modifying groups content
Starch Digestibility after different time of digestion [%]
[%]
preparation
0.5 h 1.0 h 2.0 h 4.0 h 8.0 h 16.0 h carboxyl acetyl
Native 40.7±0.7 62.4±0.2 72.0±0.3 84.7±0.2 99.4±0.2 100.4±0.3 – –
E1403 36.3±0.6 48.2±0.3 58.0±0.2 75.5±0.4 90.9±0.2 94.4±0.4 0.038±0.001 –
E1404 36.9±0.5 46.9±0.3 57.9±0.6 73.3±0.2 85.0±0.8 93.5±0.3 0.10±0.01 –
E 1412 39.1±0.8 62.6±0.7 73.0±0.6 82.2±0.3 96.8±0.7 100.1±0.1 – –
E 1414 38.5±0.7 52.9±0.4 71.5±0.8 87.7±0.7 95.6±0.6 96.2±0.1 – 0.82±0.02
E 1420 30.7±0.6 43.7±0.3 53.0±0.1 65.9±0.7 88.4±0.5 90.9±0.2 – 1.71±0.02
E 1422 35.5±0.9 39.9±0.3 52.9±0.1 64.7±0.7 88.0±0.3 93.0±0.2 – 1.21±0.01
The data presented above pointed out that degree of substitution of modifying groups might
not be the only reason for the decrease of digestibility, and some physicochemical or
structural factors could be of important significance. Modification of starch for food
application are fulfilled under very mild condition. Chemical reactions are carried out in water
suspension, then the product is separated by filtration. Next, the filter cake is dispersed in
water and filtered in order to removing by-products and unreacted raw materials. The
purification procedure is repeated twice and then the product is dried in a flash drier [16]. As
a consequence, these mild processing conditions did not cause any changes in crystal structure
of the starch, what could be clearly observed at the Figures 1 and 2.
Fig. 1. X-ray diffraction pattern of: a – native potato starch; b – bleached starch E 1403; c – oxidised
starch E 1404
Native potato starch revealed typical for tuber starches B type of X-ray diffraction pattern
(Fig. 1). All modified starches show not only the same B type of X-ray diffraction pattern but
also the same relative crystallinity (Fig. 1 and 2).
No changes in the crystal structure due to chemical modification pointed out that the structure
of the granule did not influenced digestibility of starch. This conclusion is plain because
starch is eaten after gelatinisation, and the determination of digestibility was also performed
after gelatinisation. Probably, different level of digestibility is caused by the variability of
interactions of water with modified starch macromolecules in solution.
Fig. 2. X-ray diffraction patterns of: a – distarch phosphate E 1412; b – acetylated distarch phosphate E
1414 ; c – acetylated starch E 1420; d – acetylated distarch adipate E 1422
Differently than in crystal structure, the changes in molecular structure of starch caused by
chemical modification brought about significant differences in starch pasting characteristics
(Fig. 3-5). When analysed the course of starch gelatinisation with Brabender viscograph it
should be taken into consideration that the data gained does not depend only on applied
temperature profile and measuring cartridge, but also on concentration of the investigated
starch suspension.
Fig. 3. Brabender viscosity curves for 6% suspensions of native and oxidised starches
A typical concentration value usually applied to potato starch is 3.3%, and most of our
measurements were done under such conditions. However hypochlorite modification (applied
for manufacturing of the E 1403 and 1404 preparations) caused so significant decrease in the
viscosity of starch that it was in that case immeasurably low. Consequently the pasting
characteristics of oxidised starches E 1403 and E 1404 were determined at a concentration of
6%. Under these experimental conditions native potato starch (Fig. 3) revealed a high type of
swelling characteristics, typical of tuber starch, with a rapid increase in viscosity within a
narrow temperature range and the occurrence of a viscosity peak. Modified starches E 1403
and E 1404 revealed similar type of swelling characteristics. Drop in viscosity, mentioned
above, depended on the degree of oxidation and was observed within almost all range of the
measurement (Fig. 3). The only exception was final value of viscosity of the E 1403
preparation which was higher than in case of native starch. High type of swelling
characteristics presented by all sorts of starch shown at fig. 1 pointed that process of
dissolution were relatively easy and did not clarify changes in digestibility. Also a drop in
viscosity accompanying the oxidation of starch could not be the reason of the decrease of
digestibility.
Unlike oxidised starches characterised by a very low degree of substitution, acetylated starch
E 1420 contained 1.7% of modifying groups, and revealed a course of swelling characteristics
similar to native starch, accompanied by similar viscosity values (Fig. 4). It should be
mentioned that acetylated starch revealed gelatinisation temperature significantly lower than
native starch, which pointed to easier dissolving process. However this observation did not
explain the differences in digestibility.
Fig. 4. Brabender viscosity curves for 3.3 % suspensions of native and acetylated
starches
Cross-linked starch: distarch phosphate E 1412, acetylated distarch phosphate E 1414
containing 0.8% of acetyl groups, and acetylated distarch adipate E 1422 containing 1.2% of
acetyl groups, were also investigated. These group of modified starch showed a changed type
of swelling characteristic with a progressive rise in viscosity over a wide temperature range
and lack of a viscosity peak (Fig. 5). The more restricted type of swelling characteristic
indicated a more difficult dissolving of the starch and could be the reason of the reduction of
digestibility. Paradoxically, distarch phosphate E 1412 revealing the most restricted type of
swelling characteristics revealed almost unchanged digestibility in comparison with native
starch. Consequently the solubility of starch could not be regarded as the most important
factor influencing digestibility. Acetylated distarch adipate E 1422, standing out from other
cross-linked types of starch with higher acetyl groups content, likewise with acetylated starch
E 1420 it revealed lower pasting temperature. These observation indicated that the acetylation
process caused a decrease in pasting temperature of starch, but did not explain the differences
in digestibility.
Fig. 5. Brabender viscosity curves for 3.3% suspensions of cross-linked starches
Light microscopy investigation are a very useful tool for analysing starch water interaction
in solution [10, 11, 12]. It makes it possible, for example, to observe the mechanism of
gelatinisation of starch and interaction between starch macromolecules and other substances
present in water solution. Light microscopy pictures of all modified starches were presented
in Figures 6-8. The images represents the structure of the solution formed in food products
without sterilisation process. It is known that the gelatinisation process of starch started with
swelling of starch granules and the leakage of amylose [11]. The pictures formed at the
temperature of 90°C (Fig. 6 a) showed almost homogenous medium formed mainly with
amylose and amylopectin solved in water with suspended swollen granules remnants. These
remnants were formed mainly by amylopectin which is more difficult to dissolve. The images
of hypochlorite modified starches (Fig. 6 b and 6 c) only slightly differed from that of native
starch. The most important difference was that granule remnants were more gentle what was
probably connected with the lower viscosity and consequently lower molecular mass of
starches. The analysis of light microphotographs of oxidised starches did not explain the
differences in digestibility.
Fig. 6. Light microscopy pictures of starch solutions
a. native potato starch
b. bleached starch E 1403
c. oxidised starch E 1404
Fig. 7. Light microscopy pictures of acetylated starch E 1420
solution
Light microscopy pictures of acetylated starch E 1420 (Fig. 7) represented a quite different
image in comparison to native and oxidised starches. At the microphotograph of acetylated
starch it was impossible to discriminate the amylose and amylopectin fractions. Neither
amylose molecules formed the blue complex nor amylopectin the red one. All starch
macromolecules created quite different brown complex. This observation seems to be crucial
to understand the differences in digestibility of modified starches. Modifying acetyl groups
changed the conformation of starch macromolecules so much that forming standard blue and
violet complexes of starch with iodine became impossible. These relatively deep changes in
the conformation of starch macromolecules probably hindered the enzymatic action of
amylases and caused drop in the digestibility of starch. It could not be excluded that, also in
the case of oxidised starches, the conformational changes of starch macromolecules hindered
the enzymatic action of amylases, though these changes did not disturb forming complexes
with iodine.
The light microscopy pictures of cross-linked starch (Fig. 8) indicate more difficult dissolving
process of these preparations in comparison with native starch. The images showed mainly
remnants of swollen granules. The suspending medium made from solution of amylose and
amylopectin was more difficult to observe. Distarch phosphate E 1412 which revealed almost
the same digestibility as the native starch seemed to be the most difficult to dissolve.
Nevertheless, as it was stated before, solubility of starch could not be regarded as the most
important factor influencing digestibility. Acetylated and cross-linked starch was better
soluble in water than distarch phosphate. This observation pointed that incorporation of acetyl
groups to starch macromolecules facilitated starch dissolving, but lowered their digestibility.
Fig. 8. Light microscopy pictures of cross-linked starch solutions: a –
distarch phosphate E 1412; b – acetylated distarch phosphate E
1414; c – acetylated distarch adipate E 1422
distarch phosphate E 1412
acetylated distarch phosphate E 1414
acetylated distarch adipate E 1422
CONCLUSIONS
Investigated food grade modified starch revealed reduced digestibility – up to about 10%.
The extent of decrease of digestibility depends on the degree of substitution of modified
starch, but this correlation is not linear.
Granular food grade modified starch reveals unchanged crystal structure, identical as native
starch.
Modification of starch changes its physicochemical properties i.e. pasting profile,
gelatinisation temperature and solubility in water. However these changes do not correlate
with digestibility.
Changes in conformation of starch macromolecules in solution, caused by incorporation of
modifying groups, hinder enzymatic action of amylases and cause drop in digestibility of
starch.
ACKNOWLEDGMENTS
This work was partly supported by postgraduate students fellowship No US/60/06 funded in
scope of the project “Fellowships for the best PhD Students in Wielkopolska Region
conducting research in strategic areas supporting regional development” financed by the
European Social Fund (75%) and Polish budget (25%).
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Accepted for print: 28.08.2007
Joanna Le Than
Department of Biotechnology and Food Microbiology,
The August Cieszkowski Agricultural University of Poznan, Poland
Wojska Polskiego 48, 60-627 Poznan, Poland
Phone: (+48 61) 846 60 05
Fax. (+48 61) 846 60 03
Wioletta Błaszczak
Institute of Animal Reproduction and Food Research of Polish Academy of Sciences,
Division of Food Science, Olsztyn, Poland
Grażyna Lewandowicz
Department of Biotechnology and Food Microbiology,
The August Cieszkowski Agricultural University of Poznan, Poland
Wojska Polskiego 48, 60-627 Poznan, Poland
Phone: (+48 61) 846 60 05
Fax. (+48 61) 846 60 03
email:
[email protected]
Responses to this article, comments are invited and should be submitted within three months of the publication
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![Table 1. The rate of digestion of chemically modified starches and modifying groups content The data presented above pointed out that degree of substitution of modifying groups might not be the only reason for the decrease of digestibility, and some physicochemical or structural factors could be of important signifi ficance. Modification of starch for food application are fulfilled under very mild condition. Chemical reactions are carried out in wate suspension, then the product is separated by filtration. Next, the filter cake is dispersed in water and filtered in order to removing by-products and unreacted raw materials. The purification procedure is repeated twice and a consequence, these mild processing conditi then the product is dried in a flash drier [16]. As ons did not cause any changes in crystal structure of the starch, what could be clearly observed at the Figures 1 and 2.](https://figures.academia-assets.com/45657438/table_001.jpg)
