Trends in Food Science & Technology 11 (2000) 245±253
Review
Strategies for the
manufacture of
resistant starch
Donald B. Thompson
Department of Food Science, Penn State University,
111 Borland Laboratory, University Park,
PA 16802, USA
(tel:+1-814-863-0481; fax: +1-814-863-6132;
e-mail: dbt1@psu.edu)
Starch not hydrolyzed in the small intestine is considered to
be enzyme-resistant starch (RS). Because individuals di€er in
their ability to digest starch, there is no absolute distinction
between RS and digestible starch. A method for in vitro
determination of RS should be validated using a human
population average value. In any starch material, the con-
stituent molecules will have a range of susceptibility to
amylolytic activity in vitro. For a starch or starch-containing
ingredient it is possible to alter this range by judicious
selection of processing conditions to increase the propor-
tion that tests as RS. The starch material will also have a
range of thermal stabilities before and after processing,
which may or may not re¯ect the range of susceptibility to
hydrolysis. The manufacture of RS may be thought of as
enhancement of the proportion of the starch that tests as
RS. To compare the e€ect of di€erent processing schemes, it
is critical to stipulate the method used to estimate RS con-
tent. This review focuses on strategies for increasing the
proportion of types 2 and 3 RS in a starch-containing
ingredient. Special emphasis is given to increasing RS levels
of granular starch. # 2001 Elsevier Science Ltd. All rights
reserved.
Introduction
Digestion and absorption of glucose from starch
occurs in the small intestine. Starch not hydrolyzed in
the small intestine is considered resistant starch (RS), in
the sense that it has resisted hydrolysis by the amylolytic
enzymes elaborated by the healthy human [1]. RS is
0924-2244/00/$ - see front matter Copyright # 2001 Elsevier Science Ltd. All rights reserved.
PII: S 0 92 4 - 2 24 4 ( 0 1 ) 0 0 00 5 - X
considered to have bene®cial e€ects similar to those of
some forms of dietary ®ber [2±4]. Depending on its
form, this RS may be further hydrolyzed in the colon,
where the glucose will be rapidly fermented. The func-
tional division of the human digestive tract into small
intestine and colon is at the essence of the concept and
de®nition of RS. The rate and the extent of hydrolysis in
the small intestine are both important nutritionally [5].
An altered rate of digestion may lead to an altered gly-
cemic index, and an altered extent of digestion means an
altered level of RS. (If RS makes up part of the ®xed
amount of carbohydrate in the meal employed to deter-
mine the glycemic index, a lower blood glucose response
would be anticipated on that basis alone.) In the colon,
the rate and the extent of hydrolysis of the RS are also
important nutritionally [5]. An altered rate of hydrolysis
in the colon may lead to the desired RS fermentation
occurring in a di€erent portion of the colon, with any
physiological e€ects of the fermentation products accru-
ing to the speci®c colonic region. An altered extent of
hydrolysis will lead to an altered amount of fermentation
of RS and thus an altered amount of RS excreted in feces.
Starch may be subjected to physical or chemical
treatments to alter the level of RS in a food or ingre-
dient. This review will focus on strategies to increase the
RS levels in starch ingredients.
Individuals vary widely in their responses to dicult-
to-digest starch, and what behaves as RS in one person
may not behave as RS in another [6]. Consequently,
there are no absolute distinctions between the RS and
digestible starch in a particular starch sample. For a
given starch-containing sample we can only determine
the mean level of RS for a population of individuals [6].
Because using a human population to monitor RS
levels is inconvenient, much work has been done to
develop in vitro procedures for determining RS (for a
recent review, see [1]). There is currently no generally
accepted procedure for determining RS. To be useful,
any of these procedures should be validated based on
comparison to results from human subjects [7]. This
validation may be done using ileostomists, or by other
methods [7]. Several procedures for RS determination
have been developed in an attempt to mimic starch
digestion in the human [1]. Commonly these procedures
employ pancreatic a-amylase digestion at 37 C. Even
when the quantitative results agree with studies in
humans, the nature of the RS recovered in an in vitro
246 D.B. Thompson / Trends in Food Science & Technology 11 (2000) 245±253

procedure has been shown to di€er from that recovered
in humans in vivo [8]. Another approach that has been
utilized in the literature is based on application of the
AOAC method for total dietary ®ber (TDF) [9]. This
procedure involves digestion of starch with heat-stable
a-amylase during boiling. Because the boiling treatment
itself will lead to variable losses in RS, RS determined
by this procedure will generally be lower than that
determined by a more physiological 37 C digestion.
Moreover, the nature of the RS recovered by the TDF
procedure will likely di€er from the nature of the RS
recovered by an in vitro digestion procedure at 37 C. As
will be discussed below, certain treatments may cause
the RS observed by the TDF approach to increase even
as the RS observed by a 37 C digestion decreases [10].
This observation shows that when one reads the literature
related to manufacturing of RS, one must be cautious in
comparing processes that are evaluated using di€erent
methods of determination of RS.
A starch material will be heterogenous with respect to
susceptibility to amylolytic enzymes. Consequently, for
any RS-containing sample we expect a distribution of
enzyme susceptibility, as illustrated schematically in Fig.
1. Although Englyst et al. [11] have shown that raw
potato starch is poorly hydrolyzed, the in vitro data
indicate that there is no clear biochemical distinction
between the resistant and the non-resistant portions, as
hydrolysis continues after the 120 min digestion period
in their in vitro method [10, 11]. Thus, for this sample
the RS level is determined by the precise conditions
chosen, rather than resulting from a portion of the
sample being completely resistant. The level of RS
might be considered to be illustrated by the fraction of
the area to the right of the vertical line shown in Fig. 1
(a line which would be located based on typical hydro-
lysis as observed in humans, just as the 120 min diges-
tion period for the Englyst method was validated).
When potato starch is freshly cooked, enzymatic sus-
ceptibility is much enhanced, and none of the starch
appears as RS (Fig. 1). On cooling and storage, ele-
ments of the starch decrease in enzyme susceptibility,
such that a portion of the starch can be categorized as
RS. For a particular starch material, the shape and
location of the curve will result from the precise condi-
tions of processing and storage. Strategies for manu-
facturing RS from cooked potato may be considered as
selection of conditions that shift at least part of the dis-
tribution to the right of the vertical line in Fig. 1.
Starch in a particular ingredient exists in a range of
metastable states. The manufacture of RS may be con-
sidered to be due to an enhancement of the portion of
the starch being in a metastable state that is less sus-
ceptible to hydrolysis. Because arriving at a particular
metastable state is under kinetic control, the speci®c
path, i.e. conditions of processing, with determine the
metastable state. It does not necessarily follow that
enhanced thermodynamic stability will confer enhanced
enzyme resistance. Nevertheless, one issue in the manu-
facturing of RS is the thermal stability of the particular
RS-containing material. For example, the RS of potato
and banana has limited practical utility due to its sensi-
tivity to loss by thermal treatments. Likewise, RS from
retrograded amylopectin [12] is also of limited practical
utility.

Fig. 1. Schematic distribution of susceptibility to amylolytic hydroysis. The vertical line represents the typical conditions encountered during
passage through the human small intestine. A more aggressive enzyme (perhaps from the colon ¯ora), higher temperatures, of other changes
in conditions could lead to a variable portion of the material designated as RS being hydrolyzed.
 D.B. Thompson / Trends in Food Science & Technology 11 (2000) 245±253
Figure 1 illustrates that a RS-containing food or
ingredient is generally not composed solely of RS, but
contains a portion of RS and a portion that is digestible
starch. This concept is important because in practice the
food technologist is concerned with the physical and
nutritional properties of the RS-containing material,
not just the RS component of it.
Starch is understood to be resistant to digestion for
several distinct reasons, leading to the initial classi®cation
of three types of RS: type 1, due to physical inaccessibility,
e.g. in whole grains; type 2, due to the refractory nature
of starch granules from certain plants; and type 3, due
to the reassociation, or retrogradation, of starch mole-
cules subsequent to cooking, often leading to some
crystallinity [11]. An additonal type of RS, type 4 RS, is
considered to result from certain types of chemical
modi®cation [13, 14]. All four RS types can be manipu-
lated by processing treatments, and combinations of RS
types are possible as well. In types 2 and 3, the resistance
to digestion is due to the physical state of the starch
molecules. Evidence is accumulating that the precise
nature of a RS material may lead to di€ering nutritional
e€ects [15, 16].
RS was ®rst encountered as a puzzling contaminant in
the determination of non-starch polysaccharides. Prior
to this work [17], starch was not expected to be able to
resist a boiling treatment followed by amylolytic digestion.
Subsequently, RS was observed in a di€erent procedure,
determination of total dietary ®ber (TDF) based on an
initial simultaneous boiling/amylolytic digestion step
intended to remove starch [18], a procedure that came
to be the AOAC method for TDF [9]. These two early
approaches may have contributed to an excessively spe-
ci®c idea that RS was material that survived boiling and
enzyme treatment, and the related idea that resistance
was likely due to retrogradation of amylose. While the
surviving material was undoubtedly RS, it likely was
not all of the RS initially present.
The in vitro analytical approaches designed to be
consistent with physiological processes (i.e. digestion at
37 C) allow determination of type 2 RS from potato
and banana starch, as there is no gelatinization at this
temperature. Although this strategy has been described
as allowing the analyst to distinguish among types 1, 2,
and 3 RS [11], that end can only be achieved by assuming
that boiling eliminates all RS2, an assumption that does
not hold for high-amylose maize starch (HAMS) [5, 19].
Furthermore, if RS2 is to be distinguished from RS3 in
a mixture by comparing the result of digestion following
boiling with that of digestion without boiling, one must
assume that no RS3 is lost on boiling, an assumption
that is also incorrect for the RS3 of HAMS [20], as well
as for the RS3 that is observed in retrograded waxy
maize starch [12]. Thus, distinguishing between RS2 and
RS3 is not straightforward analytically, especially with
HAMS.
247
Strategies for the manufacture of RS
Type 1 RS
The literature describes size-reduction treatments to
decrease type 1 RS, but contains little about how to
increase type 1 RS by processing treatments. The
enzyme resistance of type 1 RS is based on the inacces-
sibility of the enzyme to the starch, and it does not
depend on the intrinsic enzyme resistance of the starch
itself. In theory, any source of starch could be trapped
in a food in such a way to make it inaccessible to
enzymes. If this type of RS were part of an ingredient to
be used in a manufactured food, the ultimate RS level in
the food would depend on the stability of the entrap-
ping material to the conditions during manufacturing.
Type 2 RS
Type 2 RS in native granules
It is well known that native starch granules from
potato or banana are highly resistant to digestion [11].
There is little information available about how the spe-
ci®c cultivars of these species might be selected for high
or low RS levels. In any event, RS from these species is
something of a curiosity, since most potato products are
eaten cooked, and ripe bananas contain little or no
starch. (Bjorck has pointed out that plantain bananas
will contain appreciable starch [4].) Little information
about the level of type 2 RS in other sources is available,
although it is often assumed that normal maize and
wheat starch have little RS. We have suggested recently
that RS in both starches may be substantial, although it
is readily lost on cooking [10].
For some time it has been known that HAMS from a
variety of di€erent mutant maize genotypes are not well
digested [21, 22]. HAMS is the result of one or more
endosperm mutations that alter the nature and proportion
of the amylose and amylopectin fractions [23]. While the
adjective `high-amylose' implies that the amylose fraction
is enriched, the longer chains of the constituent amylo-
pectin also contribute to a higher apparent amylose
content [23]. Most HAMS are from endosperm con-
taining the amylose-extender (ae) gene, and it has been
known for some time that for ae-type maize starches the
level of amylose varies considerably with the genetic
background [24]. Selection of a starch cultivar for an
altered amylose content may be a means of modulating
the RS level. One company has been marketing a maize
starch with an exceptionally high amylose content as a
high-RS ingredient [14]. It is clear that judicious choice
of genetic make-up of the plant is a strategy for
manipulating type 2 RS.
Some insight into the structure of HAMS granules
and their digestion has been provided by electron
microscopy [25]. These granules appear to be attacked
from within, with partially digested granules appearing
intact from the exterior [25]. It is possible that the
248 D.B. Thompson / Trends in Food Science & Technology 11 (2000) 245±253
resistance is due to structures located in the exterior
regions of the granule. It also appears that there is great
variation in digestion among a particular population of
HAMS granules [25].
Berry ®rst showed that the amylose level in maize
starches generally correlates with RS levels [26]. Because
it was reported that high-amylose starch granules with
decreasing granule size have higher amylose content and
lower digestibility [27], fractions of high-amylose starches
were prepared in an attempt to modulate RS. As deter-
mined by the TDF method, McNaught et al. [28] found
that there is an optimal granule size for RS, and that
this size is not necessarily associated with the fraction
with the highest amylose content.
Unlike potato or banana, a good portion of the RS of
HAMS survives boiling in excess water and thus is
stable to common processing conditions. Thus this starch
is of practical interest as a type 2 RS source. Moreover,
this di€erence in physical behavior suggests that the
nature of the type 2 RS in HAMS is likely to be di€erent
than the nature of the type 2 RS of potato or banana.
Gelatinization of normal starches is well understood
to involve loss of granule order, swelling, and selective
leaching of amylose from the granule, among other
behaviors. When HAMS is cooked in excess water, it is
dicult to document appreciable swelling or selective
amylose leaching [29]. When heating of starch is mon-
itored by di€erential scanning calorimetry (DSC), a
portion of the enthalpy is not observed on immediate
reheating. For normal starches, this portion of the
initial enthalpy is understood to be associated with loss
of granule order from amylopectin crystallites immedi-
ately prior to granule swelling. However, for HAMS
this portion of the initial enthalpy is largely observed
below 90 C, far below the temperature necessary to
disperse the granule. Thus it would seem that for
HAMS the loss of original granule structure during
dispersion is not closely associated with disordering of
the crystallites of branched-chain components, as is the
case for normal starch. Recent work from our laboratory
suggests that retrogradation of amylopectin depends
upon the temperature to which the material was heated
in cooking [30±32], and we have observed a similar
behavior for HAMS from three di€erent genotypes [33].
We suggest that HAMS granules do not gelatinize in the
same sense as do normal starch granules, but rather
they lose their structure gradually over a broad tem-
perature range, thus leaving only the increasingly ther-
mostable elements as the heating temperature increases.
Enhancement of RS levels in granular starch
Various combinations of temperature, moisture, and
time may be employed to enhance RS levels while
maintaining discrete granule structure. Granules may be
either caused to swell without loss of integrity and the
constituent molecules allowed to retrograde to form RS,
or the granule may be subjected to conditions that alter
RS levels without gelatinization or melting. The latter
strategy could be considered to enhance type 2 RS,
while the former generates material more like type 3 RS,
albeit in granular form.
The physical treatment of granular starch to modify
structure without gelatinization or melting has been
termed ``hydrothermal treatment'' [34, 35]. The native
structure of the granule is semicrystalline. Hydro-
thermal treatments may be considered to either improve
the order of the crystalline fraction or enhance the pro-
portion of this fraction. Since mobility of the amor-
phous regions is thought to be a prerequisite for these
changes, hydrothermal treatments will be e€ective only
above the glass transition temperature (Tg) of the
amorphous components. Thus, hydrothermal modi®ca-
tions only occur when the component polymers of the
amorphous regions within the granules are in a rubbery
and mobile state [35]. Since Tg varies with moisture, the
moisture content dictates what minimum temperature is
required. Although the original component polymers of
the granules may be highly ordered and packed as
semicrystalline material in the native granule, they exist
in a metastable state; consequently, there is the prospect
of enhancing the stability of this physical state.
Jacobs and Delcour [35] have divided hydrothermal
treatments into heat-moisture treatments (HMT), those
performed at moisture levels below 35%, and annealing
treatments (ANN), those performed at moisture levels
greater or equal to 40%. By either type of hydrothermal
treatment it is possible to enhance the RS level without
destroying granular structure. Because both approaches
generate more highly ordered structures based on the
initial structural order, they would both be considered
annealing as the term is used in the polymer literature
[30]. Starch annealing has recently been shown to in¯u-
ence enzyme susceptibility [36]. HMT has also been
shown to in¯uence amylase adsorption onto starch [37].
At moisture contents above about 60%, loss of the
granule structure occurs at or near the gelatinization
temperature; therefore, at these moisture levels, ANN
processing temperatures must remain below this tem-
perature to maintain or improve the level of type 2 RS.
When moisture is below about 35%, the initial loss of
the granule structure occurs at higher temperatures as
moisture content decreases, as a result of what has
loosely been considered a melting transition as observed
by DSC. Thus HMT can be accomplished at these low
moisture contents even at temperatures above the gela-
tinization temperature observed with excess water. At
intermediate moisture levels (between 40 and 60%), the
granule structure is lost by a combination of gelatiniza-
tion and melting [38±40], and therefore the process has
also been considered ANN because the temperature
must be held below the gelatinization temperature if
initial granule structure is to be maintained.
 D.B. Thompson / Trends in Food Science & Technology 11 (2000) 245±253
If the starch resistance to a-amylase action is in some
way related to the metastability of ordered regions in
the granule, it is reasonable to suppose that an
improvement of the granule structure to a more stable
physical state can be associated with a more enzyme-
resistant starch granule. As a consequence of an
enhanced granule stability produced by hydrothermal
treatments, an enhanced RS content might be expected.
Wursch described two procedures for generating RS
from HAMS by annealing [41]. One procedure was for
annealing of HAMS below 100 C, generating a product
with 42% RS by digestion at 37 C. The second procedure
was for annealing of HAMS followed by debranching,
which led to a product with 30% RS by the TDF method.
Shi and Trzasko [42] described treatment of HAMS at
temperatures from 60 to 160 C (preferably 90±120 C),
with the temperature varying according to the moisture
content. Temperature/moisture combinations were chosen
to ensure that birefringence was not lost. On heating
HAMS at 100 C at 37% water for 1±4 h, about 40% RS
was determined by the TDF analysis, as compared to
12% RS in the initial HAMS. In a subsequent
improvement [43], inorganic salts were added to inhibit
swelling, thus helping retain granular structure at a
higher temperature for a particular moisture content.
Haralampu and Gross [44] heated HAMS to swell but
not rupture the granules, and then debranched the
starch and allowed it to retrograde. The product was
then annealed at 90 C, leading to about 30% RS by
TDF analysis.
Recently, we have shown that partial acid hydrolysis
of HAMS prior to ANN or HMT can improve the RS
yield (as determined by the TDF method) as compared to
ANN or HMT without partial acid hydrolysis [10]. We
believe that limited acid hydrolysis can enhance the mobil-
ity of the molecules and allow more ecient rearrange-
ment. For 120 or 140 C HMT temperatures, RS by the
TDF method increased (from about 18% for the
HAMS to as high as about 60%) even as RS by the
Englyst method decreased. We ascribed this behavior to
creating a higher proportion of thermally stable RS at
the expense of the total RS [10]. We have highlighted
the advantages of producing `boiling-stable' granular
RS, as this form of granular RS should be stable to
many subsequent thermal treatments.
Type 3 RS
The production of type 3 RS involves retrogradation
of starch molecules subsequent to gelatinization or dis-
persion of the native starch granules. Some recently
proposed strategies involve partial depolymerization
either before or after native granule structure is lost.
Selective hydrolysis before thermal treatment is done to
allow increased polymer mobility for molecular rear-
rangement, and hydrolysis after thermal treatment is
done to enhance the proportion of RS in the ingredient.
249
Colonna et al. [45] reviewed the molecular e€ects of
physical modi®cation of starch and then reviewed the
factors which limit a-amylolysis in heterogeneous phases:
di€usion of the enzyme, porosity of the starch matrix,
adsorption of enzyme to substrates, and catalysis.
Although type 3 RS is often attributed to amylose
retrogradation [46, 47], retrograded amylopectin has also
been shown to contribute to type 3 RS [12, 48]. As noted
above, RS from retrograded amylopectin is completely
lost on heating to 100 C. Klucinec and Thompson [50]
have shown that dispersions of amylose and amylo-
pectin apparently interact on cooling. Although it may
be that RS would result from this interaction, this RS
would not be stable at 100 C. Nevertheless, the presence
of amylopectin has been shown to detract from ordering
of amylose as observed by DSC [49, 50]. Debranching
of amylopectin was ®rst shown by Berry to be e€ective
in generating RS from amylopectin [26]. This treatment
has the additional advantage that the adverse e€ect of
amylopectin on amylose ordering is apparently mini-
mized as well.
Most amylose-containing starches can be processed
by heat and moisture to produce some type 3 RS. For
example, wheat starch will produce low levels of RS as
a result of gelatinization and cooling in bread baking
[51, 52].
HAMS is the preferred starting material for producing
high-RS ingredients. One challenge with this starch is
heating to accomplish complete gelatinization, and the
question may remain whether RS produced is type 2,
type 3, or a combination of both. For HAMS the DSC
endotherm on initial heating in excess water has two
readily identi®able components: one below about 90 C,
lost after initial heating, and attributed to the branched
molecules in the raw starch; and one between 90 and
110 C, observed on an immediate reheating, and attributed
to melting of amylose/monoacyl lipid complexes. In an
RS-containing material the ®rst of these two endo-
therms may be absent without the presence of an endo-
therm at or above 120 C. Since a marker to indicate the
absence of gelatinization can be lost without creating a
positive marker for one subcomponent of type 3 RS,
this high-temperature endotherm cannot be used to dis-
tinguish between types 2 and 3 RS for HAMS. Thus one
cannot assume that thermal processing of HAMS at
100 C will convert its type 2 RS to type 3 RS.
DSC endotherms above 120 C have been observed for
some RS preparations from HAMS, and have been
attributed to amylose retrogradation [46]. When this
high-temperature endotherm is present in a treated
HAMS, it clearly indicates the presence of type 3 RS.
However, it is important to note that type 3 RS may
exist without this endotherm. Because monoacyl lipid
has been shown to provide the central stabilizing ligand
for helical amylose inclusion complexes, some have
shown that monoacyl lipid interferes with amylose
250 D.B. Thompson / Trends in Food Science & Technology 11 (2000) 245±253
double-helical association, and consequently with RS
formation [53±55]. In recent work, we have shown that
amylose association on cooling, as observed by DSC, is
enhanced when native monoacyl lipid levels are reduced
prior to heating [33].
Use of the TDF method to quantitate type 3 RS after
processing treatments might lead one to conclude that
type 3 RS is entirely stable to boiling. In one recent
review focusing on type 3 RS [13], the authors state that
type 3 RS was shown to be ``thermally very stable,'' but
this RS was that isolated by the TDF method. These
authors subsequently noted that ``highly resistant''
fractions are those that resist hydrolysis at 100 C. The
nature and amount of type 3 RS isolated by the 37 C
digestion has been compared to that isolated using a
boiling/digestion treatment [56]. Type 3 RS was pre-
pared from maize, isolated using a 42 C digestion, and
then subjected to a boiling/digestion treatment. They
observed that about one-third of the initially isolated
RS was further hydrolyzed by the boiling/digestion
treatment. Both the bacterial enzyme activity and the
high temperature contributed to the additional hydro-
lysis [56]. Others have compared the amount of RS from
HAMS that could be determined by the two types of
analysis. One group found about 29% RS by the 37 C
treatment but only 14% RS using a boiling treatment
[20]. In a separate study of RS from autoclaved HAMS,
the 37 C procedure produced a RS value of 39%,
whereas the boiling treatment gave a value of only 21%
RS [57]. Consequently, for these samples the boiling/
digestion procedure allows more complete amylolytic
digestion.
Several reports describe the manufacture of material
assumed to be type 3 RS from HAMS. Sievert and
Pomeranz [46] described the use of autoclave/cooling
cycles to produce RS. They investigated autoclaving at
temperatures of 121, 134, and 148 C, and explored up
to 20 autoclave/cooling cycles. By this approach they
were able to produce up to 40% RS as determined by
the TDF method. They studied the autoclaved starches
and RS isolated by the TDF method, and showed that
the high-temperature DSC endotherm of the intact
starches correlated with a similar endotherm from the
RS. The authors noted that although they showed a
positive association between RS and the high-temperature
endotherm, RS is not a well de®ned material. Moreover,
one must bear in mind that the TDF procedure would
select for that portion of the RS that would be most
likely to contribute to this endotherm. It would be
inappropriate to use the DSC as a means of evaluating
the levels of RS in the processed starch material unless
one knew that the particular RS sample was one known
to have a high level of highly ordered amylose. For
example, Sievert and Pomeranz isolated RS from raw
HAMS by the same procedure and yet did not observe a
high-temperature endotherm [46]. Subsequent work by
this group employed re®ned autoclave/cooling strategies
for preparing RS [57±60].
Based on polymer crystallization theory, Eerlingen
et al. [61] hypothesized that the formation of type 3 RS
can be considered as a crystallization process of amylose
in a partially crystalline system. To test that hypothesis,
they autoclaved wheat starch in excess water at 120 C
for 1 h. Then, they controlled the crystallization condi-
tions by cooling the gelatinized starch to 0, 68, or 100 C
and followed the production of RS over time by the
TDF method. They found that initially (15 min) the
yield of RS was favored by the lowest crystallization
temperature (0 C), while for long storage times (>10 h)
the yield of RS was favored by the highest crystal-
lization temperature (100 C). The authors interpreted
these results based on the idea that at 0 C (just above
the starch Tg of about 5 C) the nucleation rate of amy-
lose crystals was favored and the propagation rate was
limited; therefore, the RS increased rapidly initially but
leveled o€ quickly. On the other hand, when the gelati-
nized starch was stored at 100 C (below the temperature
of the starch melting transition region, 150 C) the
propagation rate of amylose crystals was favored even
though the nucleation rate was rather limited. Once a
few nuclei were formed, they were extensively propa-
gated; therefore, the initial yield of RS was low but this
strategy produced the highest RS over time.
Eerlingen et al. [47] also studied the in¯uence of amy-
lose chain length on type 3 RS formation, as isolated by
the TDF method. They found that the chain length of the
crystalline regions of type 3 RS was independent of the
amylose chain lengths originally used to form the RS.
As DPn increased to 260, the yields of RS increased with
the amylose chain length, reaching a maximum value of
28% RS. Longer chain lengths than DPn 260 yielded
less than the 28% RS. The explanation proposed by
Eerlingen et al. [47] for the formation of RS in amylose
solutions was that amylose with DPn lower than 100
would not have enough chains with the minimum length
required to form resistant crystallites. In contrast, based
on entropic considerations, amylose with DPn higher
than 300 would not easily reach the required alignment
of polymer chains to form resistant crystallites.
Cairns et al. [62] prepared amylose gels and studied
the enzyme-resistant portion after 24 h at 37 C. They
suggested that resistance was due to discontinuous
crystals, with amorphous regions contained within the
crystals and protected by them. Gidley et al. [63] pro-
duced type 3 RS from HAMS by autoclaving at 134 C
and subsequently isolating the RS using a 42 C amylolytic
digestion according to Berry [26]. They then studied the
nature of the isolated RS and concluded that it was 60±
70% double helical but only 25±30% crystalline. The
authors suggested that double helices were aggregated
imperfectly into B-type crystallites, with single chain
material present as imperfections. Crystallinity is also
 D.B. Thompson / Trends in Food Science & Technology 11 (2000) 245±253
clearly not the primary explanation for enzyme resis-
tance of type 2 RS, as the crystallinity of raw potato
starch is high, while that of raw HAMS is much lower,
and yet both are high in RS. One must be cautious in
generalizing from the work of Gidley et al. [63], because
the particular manufacturing conditions may have pro-
duced a RS with a particular nature. The 134 C auto-
claving treatment likely allowed some type 2 RS to be
retained, as some minor birefringence and granule frag-
ments were observed [63].
We have recently shown that when HAMS is heated
to 160 C, the high-temperature endotherm is no longer
observed on reheating [33], and we interpret this behavior
as an indication that the 160 C temperature allowed
melting of the most stable structures. Others have
shown that RS heated to 200 C in water eliminates the
characteristic high-temperature endotherm on reheating,
whereas treatment at 100 C does not eliminate this
endotherm [60]. Still others have shown that on cooling
of RS after heating to 180 C, an exotherm is observed
at about 60 C, which was attributed to reassociation of
the chains previously comprising RS [55]. In our work
with HAMS we have been unable to observe a high-tem-
perature endotherm after HAMS thermally processed to
contain that endotherm was heated to 180 C, and so we
do not believe that the exotherm on cooling represents
reassociation of the chains in the same form as before
heating to 180 C [33].
Iyengar et al. [64] developed a method for manu-
facturing type 3 RS that involved the dispersion of
HAMS by cooking an aqueous suspension, incubating
at 60±120 C, and then holding at 4 C. Because intact
amylopectin otherwise reduced the retrogradation rate,
a enzymatic debranching step was suggested prior to the
60±120 C incubation in order to accelerate crystallization.
The level of RS could be increased by subjecting the retro-
graded material to enzyme or acid hydrolysis, resulting in
a predominately crystalline material that was about
92% resistant to digestion at 37 C.
A related method for manufacturing type 3 RS was
described by Chiu et al. [65] and Henley and Chiu [66].
This method followed the same general debranching
strategy of Iyengar et al. [64]; the innovation of Chiu et al.
[65] was that cooling by extrusion could replace subject-
ing the dispersion to cooling and heating cycles to pro-
duce RS, thus dramatically reducing the process time.
Vasanthan and Bhatty [67] described a type 3 RS-
containing product made from HAMS and other starches
using an initial heating at 100 C and retrogradation at
4 C. Samples were then subjected to partial acid hydro-
lysis prior to hydrothermal treatment at 30% water by
heating to 100±140 C. By the TDF method, a RS con-
tent of over 25% was achieved for HAMS.
Haynes et al. [68] described manufacture of a particu-
larly thermostable type 3 RS, with a DSC endothermic
peak above 140 C. The process was designed to avoid
251
lower-melting crystals and amylose±lipid complexes.
The initial heating step is above the amylose±lipid complex
melting temperature, but said to be below the melting
point of the type 3 RS (also below the temperature for
complete dispersion of HAMS). A minimum of one cycle
(but preferably four) of nucleation and propagation, ®rst at
a temperature (about 60 C) that precludes amylopectin
crystallization, then at a temperature above amylose±
lipid complex formation (above 120 C) is called for. The
latter temperature would remove any amylose±lipid
complexes formed during nucleation. They produced a
material with up to 35% RS by the TDF method. The
authors also described use of debranching enzymes after
preparation of the high-melting RS product, use of
high-melting RS seed crystals for nucleation, or a ®nal
annealing treatment. One might consider this temperature
cycling protocol to be an improvement over the original
temperature cycling of Sievert and Pomeranz [46], as the
earlier treatment would have had a similar e€ect of
removing amylose±lipid complexes and amylopectin
crystals while retaining the more stable amylose±amy-
lose interactions with each heating.
Kettlitz et al. [69] produced highly fermentable RS of
largely DP 10-35 by beginning with a potato or tapioca
maltodextrin, and simultaneously debranching and
allowing retrogradation. The product was about 55%
RS as determined by 37 C incubation, and had a DSC
peak at about 105 C. The advantage of this product is
in its enhanced production of butyrate (observed in
vitro), which is thought to contribute to colon health.
Type 4 RS
The digestibility of modi®ed starches has been studied
from the perspective of caloric availability and safety [70],
but not from the point of view of using these techniques to
intentionally modulate RS levels. The use of chemical
modi®cation speci®cally for manufacture of RS is described
by Seib and Woo [71], who explain how generation of
distarch phosphodiester cross-linkages could be modulated
to produce products of 40±98% RS by the TDF method.
Overall considerations for the manufacture of RS
RS is generally found as a component of a RS-con-
taining ingredient or food. The goal of including an
ingredient high in RS should be to combine physical
functionality, processing stability, and nutritional func-
tionality. The physical functionality of the RS-containing
ingredient is required for appropriate physical charac-
teristics of the food, such as texture, water holding
capacity, etc. The processing stability of RS is important
in order to preserve the nutritional functionality of the
RS-containing ingredient. The nutritional functionality
of the RS-containing ingredient can involve both resis-
tance to digestion in the small intestine and resistance to
fermentation in the colon. To fully take advantage of
the prospects for use of RS in foods, it will be important
252 D.B. Thompson / Trends in Food Science & Technology 11 (2000) 245±253
to understand the physical bases for enzyme resistance in
the small intestine and in the colon. With such knowledge it
might be possible to develop a wide range of RS-con-
taining materials for use in manufactured foods.
Categorization of RS into types 1, 2, 3, and 4 has
provided a conceptual basis for di€erentiating among RS
materials. However, these categories may be a constraint
to better understanding of RS because they may con-
tribute to the idea that there is homogeneity within a
type, and because they may contribute to the idea that
the di€erentiation among the types is straightforward.
RS is a concept, not a precise physical entity.
Although it is de®ned with reference to the human
digestive tract, it must also be de®ned for a population
of healthy individuals, not for a speci®c individual. The
lack of an accepted in vitro method for RS determina-
tion is a major current limitation to the clarity of the
®eld of research on RS manufacture. However, if there
were such an accepted method, it might obscure our ability
to perceive the diverse nature of materials contributing to
RS. There is likely a distribution of enzyme susceptibilities
in starch, and processing treatments may be considered to
in¯uence this distribution. Manufacturing of RS should be
considered in this general context. Eventually we should be
able to produce starch materials having a desired rate and
extent of digestion (in terms of mean population respon-
ses), and (for any RS that might be present) a desired rate
and extent of hydrolysis and fermentation in the colon.
Acknowledgements
The author thanks Jorge Brumovsky for assistance with
the literature search, and Koushik Seetharaman and
Annette Evans for review of the manuscript and helpful
suggestions.
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