Measurement of the Starch Content

of Commercial Starches
Analytical Working Party of the Starch
Experts Group (STEX) of the European
Starch Associations (ESAY
For the EC starch regulation 2169/1986 a minimum purity of 97%
starch on dry substance is required to obtain the new end-use
refunds. Three different methods for the determination of starch
were tested. Two polarimetric methods (EC L123/72, AOAC
14031-032) and an enzymatic merhod (EC L158/86) were used to
determine the high starch content of commercial unmodified starches. The results of a ring test between six laboratories shows that
the AOAC method, solubilizing the starch in the presence of calcium
chloride, has the smallest standard deviation. An explanation for the
differences observed between the three methods is given, based on
the sugar spectrum of the hydrolyzed starch. An independent ring
test in France shows a modified enzymatic method, developed by
AFNOR (the French standards organization) produces satisfactory
results.

Messung des Starkegehaltes handelsublicher Starken. In der

EG-Starkeregelung 2169/1986 wird eine Mindestreinheit von 97%
Starke in der Trockensubstanz gefordert, um die neue Endverbrauchs-Erstattung zu erhaltcn. Fur die Bestimmung der Starke wurden drei verschiedene Methoden getestet. Zwei polarimetrische (EC
L123/72, AOAC 14031-032) und eine enzymatische Methode (EC
L158/86) wurden angewendet, urn den hohen Starkegehalt handelsiiblicher nichtmodifizierter Starken zu ermitteln. Die Ergebnisse
einer Ringuntersuchung zwischen sechs Laboratorien zeigen, daR die
AOAC-Methode, bei der die Starke in Gegenwart von Calciumchlorid gelost wird, die geringste Standardabweichung hat. Auf der
Grundlage des Zuckerspektrums der hydrolysierten Starke. Ein
davon unabhangiger Ringtest in Frankreich zeigt, daR eine von
AFNOR (die franzosische Standardisierungs-Organisation) modifizierte enzymatische Methodc zu befriedigenden Ergebnissen fuhrt.

1 Introduction

Table 1.
Starch Content (dry basis) of Commercial Starches (%).

On 1st July 1986 a new E C starch regime came into force whereby
substantial refunds are given for starch used in certain agnculturally unprotected applications (EC regulation 2169/86).
In order to qualify for the new end-use refunds, a minimum
purity of 97% starch o n a dry substance basis is required.
Consequently it is essential to have an analytical method that
allows the measurement of this high starch content in a reliable,
reproducible and accurate way. The Starch Expert Committee
of the European Starch Associations decided to take a critical
look at existing methods. All the methods chosen were intended to some extent to measure the relatively low starch content
of various starch containing products.
To meet the high purity requirements of 97% on a dry basis, the
standard deviation of the applied method should be less than 1%.
To evaluate the accuracy and repeatability between different
laboratories, the Starch Industries organised a ring test amongst
some of its members. The methods tested were:
1) Ewers method
EC L123172
2) Calcium chloride method
A O A C 14031-32
3) Enzymatic method
E C L1.58186
The methods were evaluated by analysis of the starch content of
wheat starch, potato starch and maize starch (Tables 1 and 2).
The analyses were carried out in triplicate by 2 o r 3 analysts
within one laboratory and repeated on the same samples by
different laboraties. The results were collected and the statistics
were calculated according to the A O A C Statistical Manual [ 11.

Method

Wheat

Maize

Potato

Ewers
00

98.94
0.36
2.07

98.67
0.48
1.41

100.68
0.58
1.73

CaCI?
00
ax

97.26
0.30
1.11

97.84
0.28
0.83

98.88
0.40
1.05

Enzymatic
00

97.45
1.53
4.06

97.68
1.84
3.69

99.78
I .46
4.90

OX

OX

00

= repeatability

OX =

standard deviation
reproducibility standard deviation

Table 2.
Non-Starch Components Perccntages on Dry Basis.
Product
Protein
I S 0 3188

Wheat starch
Maize starch
Potato starch

0.16
0.33
0.10

Method
Ash
Total lipids
I S 0 3593 after hydrolysis
CIRF 8.14
0.25
0.07
0.21

0.31
0.51
0.02

2 Principles
2.1 Ewers method EC L123/72
The method consists of a double determination. The sample is first
heated with diluted hydrochloric acid. By heating, the starch is gelatinized and by the action of hydrochloric acid the starch is hydrolyzed to
smaller oligosaccharides and glucose as is shown in Fig. 1. After
clarification and filtration the rotatory power is measured by polarimetry. Secondly, a sample is extracted with 40% ethanol. The

414

starch/starke 39 (1987) Nr. 12. S. 414-416

filtrate is acidified with hydrochloric acid and after clarification and

filtration, the rotatory power of the free sugar content is measured
under the same conditions as in the first determination.
The starch content is given by difference between the two measurements, multiplied with a known factor. This factor depends on the
nature of the starch: For corn starch the factor F = 184.6, for wheat
starch F = 182.7 and for potato starch F = 195.4.

* Participants:

Amylum, Avcbe, Cargill, C P C E . Roquette, ZBB.

0 VCH Verlagsgesellschaft mbH. D-6940 We~nheim.1987

0038-9056/87/1212-0414$02.50/0

3.2 Calcium chloride AOAC 14031-032

As the solution of the starch does not rely on hydrolysis but
rather on the formation of a soluble complex, the heating step is
much less critical. The preparation of the sample for polarimetry is straight forward and the polarimetric measurement is
reproducible between laboratories.
The factor used in the calculations is perhaps inappropriate for
high starch concentrations. This explains the systematic low
results.

3.3 Enzymatic EC L158/86

Flg.

Fig 2

Fig. 3

Figure 1-3. Sugar analyses were performed on a Bio-Rad HPX-87C

column at 85C with water as eluent and a flow rate of 0.4 mYmin.
I . Glucose 2. Maltose 3. Triose 4. Tetraose 5 . Pentaose 6. High
molccular weight polysacharides 7. CaCI2 8. Acetic acid 9. Sodium
salts.

2.2 Calcium chloride method (AOAC 14031-32)

The calcium chloride method is a polarimetric method for the determination of the starch content. The method is based on a differential
measurement of the optical rotation of the ,,treated" sample and on an
ethanol extracted blank. The calcium chloride method, unlike the
Ewers method is based on the solubilisation of the starch in a 33%
calcium chloride solution at neutral or slightly acidic pH by the addition
of acetic acid. The high ionic strength causes the starch molecule to
unfold and to dissolve without the formation of oligosaccharides or
glucose (Fig. 2). The free sugar content, present in the sample before
treatment. can be detected by an ethanol extraction. In the ringtest this
has been deleted.

2.3 Enzymatic method EC L158/86

This method is based on the hydrolysis of the starch by an amyloglucosidase, at 60C, of which the endproduct is glucose (Fig. 3). Before
hydrolysis the starch is dispersed in a 0.5 N sodium hydroxyde solution
at 60C. The glucose content, after hydrolysis, is determined by an
enzyme oxidation-reduction system, detected at U. V. wavelengths.

The benefit of an enzymatic method is that enzymes are specific

in their action and, therefore, in the case of starch, hydrolyse
only the starch and not other material present. In the present
case we have nearly 100% starch so the specificity is less
important. Indeed it is a disadvantage because amyloglucosidase, for instance, is not suited to break all 1,6 linkages.
The alcohol content must be as low as possible before enzyme is
added or the amyloglucosidase will be denatured.
The reagents used are biologically unstable and this can cause
problems in the hands of inexperienced operators. The use of
fresh reagents is highly desirable but not always possible.
At the end of the procedure 0.1 ml of the test solution is taken
and this very small aliquot is a further source of error. Also the
standardization of spectrophotometers between laboratories is
notoriously difficult.
Dissolving starch in a 0.5 N sodium hydroxide solution at 60C is
very difficult and could cause errors in the end determination.
A n improved enzymatic method was tested by the French
starch producers. This method combines parts of the official E C
method with a French A F N O R method. The new method is
proposed as a new A F N O R project V.036.06. Reproducible
results were obtained this way (Table 3).
Table 3.
Enzymatic Determination of YO Starch Content (dry basis) According
to the AFNOR Method (Project V.036.06).
Method

Wheat

Maize

Potato

AFNOR

97.78
0.47

98.60
0.34

98.36
0.91

(3X

3 Critique of the Methods

4 Conclusion

3.1 Ewers EC L123/72

A n examination in detail of the results of the collaborative test

(which will be reported in detail in due course) indicates that of
three methods studied, the calcium chloride method has the
best reproducibility. It is also a simple method to carry out and
needs a minimum of equipment.
It must be remembered that none of the methods tested has
been devised for the determination of starch contents at such
high levels. The E C enzymatic and Ewers methods were
originated for measuring the starch content of animal feeding
stuffs and similar products with 20 to 50% starch content on dry
basis. The calcium chloride method is intended for the determination of the starch content of flour, that is up to a maximum of
85% starch on dry basis. It is a principle of regulatory bodies
that a subtance refered to in legislation should be the component to be determined, whenever possible. The results obtained
by the calcium chloride method are low in relation to the known
starch content of the samples tested. It is likely that this is due to
the factors used in the calculation of the results. The coefficients

This method relies on hydrolysis of starch by hydrochloric acid

at atmospheric pressure using a boiling water bath for 15 min.
Under these conditions hydrolysis of the starch is quite incomplete [2,3]. Therefore the degree of hydrolysis will depend on:
- Heat transfer characteristics of water batidflask system used.
For instance the thickness of the flask, the extent of agitation
of the contents of the flask, the thermal capacity of the water
bath.
- The precision of timing of the hydrolysis before cooling.
- Heat transfer characteristics of the flask during cooling [4,5,
61.
- Influence of clarification reagents and their amounts [7, 81.
Thus, the preparation of the sample for polarimetry presents a
number of sources of error. The polarimetric measurement
itself is reproducible between laboratories, and standardization
using sucrose is recommended.
starchktarke 39 (1987) Nr. 12, S . 414-416

415

of variation of the Ewers and enzymatic methods as currently

written are unacceptably high. Due to the economic significance of starch purity determinations, a method is required
which will give clear. indisputable results. In relation to a purity
requirement of 97%, no method with a repeatibility between
2% and k 5% is likely to meet this requirement.
The results obtained with the proposed AFNOR enzymatic
method show that improvements can and should be made
before a final analytical method can be agreed upon.

Bibliography
[ l ) Youden, W. J.,and H . Sreiner: AOAC Statistical Manual 1975.
[2] Ewers, E. : Ztschr. f . offentliche Chemie 14 (1908), 150-157.
[3] Witikler, S.,and G . Luckow: StarchlStarke 19 (1967), 110-115.
[4] Dudas, F.. Starch/Starke 28 (1976), 127-129.
F . : StarcNStarke 26
157-159.
[6] Dudas, F.: StarchEtarke 23 (1971), 390-393.
[7] Dudas, F.: StarchlStarke 24 (1972), 367-369.
[8] Dudas, F.: StarchlStarke 22 (1970). 385-388.
Correspondence to: MI. E. Moreels. Amylum N . V., Burchtstraat
10, B-9300 Aalst (Belgium).
(Received: May 6, 1987).

Vergleichende Untersuchungen zur Erbsenstarkeisolierung auf nantechnischem Wege*

N. U. Haase, W. Kempf, G. Tegge
und U. Dheur, Detmold
Kornererbsen enthalten Starken, die abhangig von der Sorte iibcr
sehr hohe Amyloseanteile verfiigen. Fur die bisher weitgehend fehlenden analytischen Untersuchungen derartiger Starken miissen
vorab moglichst gute Isolierungen durchgefuhrt werden. Deshalb
wurden in der vorliegenden Untersuchung die drei Verfahren der
Flutung, der Zentrifugation und der Extraktion einander gegeniibergestellt, wobei die Proteinabtrennung besonderes Gewicht einnahm.
Ein kurzzeitiger Kontakt der vermahlenen Erbsen mit Natronlauge
fuhrte sowohl bei der Flutung als auch bei der Zentrifugation zu
einer vergleichbar hohen Starkequalitat, doch lag der Wirkungsgrad
der Starkeabtrennung bei der Flutung um 30% hoher. Die Extraktion fie1 gegeniiber den beiden anderen Methoden beziiglich der
verwendeten MeRkriterien deutlich ab. Ein zusatzlich vorgenommener Sortenvergleich wies darauf hin, daR die Starke aus Palerbsen
besser abtrennbar ist als die aus Markerbsen.

Comparative Investigations on the Wet-Separation of Pea

Starch. Peas contain starches with partly hrgh amylose fractions. As
exact analytical investigations do not exist yet, the first thing is to
develop an efficient technique to isolate these starches. Therefore, in
this study three laboratory procedures of wet milling which are based
on settling tables, centrifugation and extraction are compared. In this
connection the efficiency of protein separation was of importance. A
short contact of the ground peas with sodium hydroxide resulted in a
comparably high starch quality when using tables as well as centrifuges. However, the recovery rate of starch was 30% higher when
using tables. Compared to the other procedures the extraction
method led to starches of much lower quality with regard to the
tested quality criteria. An additional comparison of different pea
species indicated that the starch separation from smooth peas is more
efficient than that from wrinkled peas.

1 Einleitung

derartige Starken ohne vorgeschaltete Aufarbeitungstechniken

zu nutzen. Als Rohstoff konnen dabei neben Amylomais [4]
verschiedene Leguminosenarten [9] eingesetzt werden, die sortenabhangig - ebenfalls sehr hohe Amylosegehalte besitzen.
Eine Eingliederung dieser Leguminosenstarken in das vorhandene Starkeangebot erfordert in jedem Fall Starken mit
bestimmten Qualitatseigenschaften. D a bislang gewerblich vertriebene Leguminosenstarken in der Regel nur iiber Windsichtung [l, 21 abgetrennt werden und damit die fur die anderen
Starkearten bereits bestehenden Qualitatsanforderungen in
der Bundesrepublik Deutschland [ 101 nicht erfullen konnen,
muB zunachst ein praktikables Verfahren zur Starkeisolierung
im LaboratoriumsmaRstab erarbeitet werden, um im Anschlurj
daran qualitative Studien an diesen Starken moglich zu
machen. Die vorliegende Arbeit berichtet uber Versuche, die
Starkeabtrennung auf naBtechnischem Wege vorzunehmen.

Die sich verscharfende Situation der Agraruberschusse der

Europaischen Gemeinschaft hat bereits zu einer Ausweitung
des Anbauspektrums der Kulturarten gefiihrt. Mit der Bereitstellung neuartiger Rohstoffe fur die industrielle Nutzung sol1
dabei eine Entspannung der Uberschurjsituation auf dem Nahrungs- und Futtermittelsektor erreicht werden. Diese Sachlage
tnfft neben verschiedenen 0 1 - und Faserpflanzen auch einige
Leguminosenarten, bei denen es Ansatze gibt, neben dem
Proteinanteil auch die Starke zu nutzen [l,2 , 31.
In der industriellen Verwertung finden iiberwiegend Starken
Verwendung, die Amylosegehalte zwischen 20 und 25% aufweisen. Der Einsatz amylosereicher Starken, die heute noch
ausschlierjlich aus Amylomais staqmen [4] bzw. uber Fraktionierungsschritte angereichert werden [ S ] , findet hingegen trotz
vielfaltiger Einsatzmoglichkeiten [6, 7, 81 nur in sehr begrenztem Umfange statt. Die verstarkte Nutzung dieses Potentials
wird voraussichtlich auch allein dann erfolgen, wenn es gelingt,

2 Material und Methoden

* Nr. 5579 der
**

Veroffentlichungen der Bundesforschungsanstalt fiir

Getreide- und Kartoffelverarbeitung. Detmold.
Vortrag von N . U. Haase anlaBlich der 38. Starke-Tagung der
Arbeitsgemeinschaft Getreideforschung in Detmold vom 22. bis 24.
April 1987.

416

starchklrke 39 (1987) Nr. 12. S. 416-421

Die Starkeabtrennungen wurden aufgrund ihres orientierenden Charakters im Laboratoriumsmarjstab mit Erbsen bzw.
Erbsenmehlen (vermahlen in einer Schlagkreuzmuhle) durchgefuhrt.

8 VCH Verlagsgesellschaft mbH. D-6940 Weinheim. 1987

0038-9056/87/1212-0416$02.50/0