2

Analytical Methods for Determining Pectin Composition

Landis W. Doner

Agricultural Research Service, North Atlantic Area, Eastern Regional Research Center,
U.S. Department of Agriculture, Wyndmoor, PA 19118

Chemical and modern chromatographic methods have been

applied to the determination of pectin composition and
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structure. The dominant structural feature of pectin

Publication Date: June 5, 1986 | doi: 10.1021/bk-1986-0310.ch002

is a linear chain of galacturonic acid residues, some

of which are esterified with methyl groups. Chemical
methods (including reactions with carbazole and sub­
stituted phenols) and chromatographic methods (GLC and
HPLC) are available for galacturonic acid determination.
Methyl ester levels are determined either chemically
(after oxidation to formaldehyde) or by GLC after pectin
ester saponification. A variety of neutral sugars are
present in pectin, mainly rhamnose, galactose, arabinose,
and xylose. Effective GLC procedures are available for
their determination, and applicable liquid chromatographic
methods have been developed. Measurements of less the
less-common substituents, such as O-acetyl and O-feruloyl
esters have been achieved by colorimetric and titrimetric
13
procedures. Developments in infrared and C-NMR spectro­
scopy have resulted in these being applied to structural
analysis in pectin.
Pectin polysaccharides and the hemicelluloses are matrix components
i n the c e l l walls of higher plants. T r a d i t i o n a l l y , these classes
of carbohydrates have been defined operationally by t h e i r presence
i n fractions obtained by sequential extraction of c e l l walls.
The p e c t i c substances are extracted with water, d i l u t e a c i d , or
with calcium chelating agents, such as EDTA, ammonium oxalate, or
sodium hexametaphosphate. But c l a s s i f i c a t i o n of polysaccharides
i s best based on s t r u c t u r a l components rather than on the method
used f o r i t s i s o l a t i o n . According to structure, the pectic
substances would include galacturonans, rhamnogalacturonans,
arabinans, galactans, and arabinogalactans which possess a l i n e a r
β-1,4-Ι) -galactan backbone.
A recent c l a s s i f i c a t i o n (1) describes the p e c t i c polysaccha­
rides as those polymers found i n covalent a s s o c i a t i o n with galactu-
ronosyl-containing polysaccharides. The hemicelluloses are those
carbohydrate polymers which are noncovalently associated with
c e l l u l o s e . Diverse categories of pectic polysaccharides occur
not only among plant sources, but among tissues i n a given source.

This chapter not subject to U.S. copyright.

Published 1986, American Chemical Society

In Chemistry and Function of Pectins; Fishman, M., et al.;

ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
 14 CHEMISTRY AND FUNCTION OF PECTINS

A comprehensive (2) review on the structure of p e c t i n has recently

been published. Pectin was described as consisting of a branched
block, i n which the main galacturonan chain i s interrupted by
rhamnose u n i t s . Many of these rhamnoses carry arabinan or galac-
tan chains; the galactan chains are sometimes further substituted
with arabinan segments. These heavily branched galacturonan
chains alternate with unbranched blocks i n which rhamnoses are
r a r e l y present. Methyl esters of galacturonic acid are also
present as blocks, a l t e r n a t i n g with sequences of n o n - e s t e r i f i e d
galacturonic acid.
This review w i l l describe the a n a l y t i c a l methods a v a i l a b l e
to determine the s t r u c t u r a l components of p e c t i n . These features
determine the important p h y s i c a l , chemical, and b i o l o g i c a l proper-
t i e s of p e c t i n . Included w i l l be discussion of galacturonic acid
determinations, degree of e s t e r i f i c a t i o n with methyl groups, the
neutral-sugar composition, and analysis of some less-common
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Publication Date: June 5, 1986 | doi: 10.1021/bk-1986-0310.ch002

e n t i t i e s , such as 0-acetyl and O-feruloyl linkages.

Quantitative Analysis of Galacturonic Acid i n Pectin

The dominant and u n i f y i n g s t r u c t u r a l feature i n pectins i s a

l i n e a r 1^4-a-linked D-galactopyranosyl-uronic acid chain. a-L-Rham-
nosyl residues are inserted at i n t e r v a l s i n the chain, and v a r i a b l e
proportions of the uronic acid residues are e s t e r i f i e d with
methanol. Neutral sugars other than rhamnose are present, and
neutral sugar l e v e l s often t o t a l about 20%. The other neutral
sugars are mainly D-galactose, L-arabinose, and D-xylose, and
these are l i k e l y to be attached i n branches to the rhamnose
residues i n the main chain. In a l a t e r section, various chromato-
graphic approaches f o r determining the l e v e l s of i n d i v i d u a l
neutral sugars w i l l be described.

Chemical Methods. Determinations of galacturonic acid of p e c t i n

usually includes both the free and e s t e r i f i e d forms, since strongly
a c i d i c media are employed i n the c o l o r i m e t r i c methods. Procedures
which continue to be used widely are modifications of those
described e a r l y by Dische. One i s based upon reaction with
cysteine (3) and the other with carbazole (4). One modification
of the carbazole method, which gave a doubling i n s e n s i t i v i t y ,
along with an increased s t a b i l i t y i n color and greater reproduci-
b i l i t y , was reported i n 1962 (5). Inclusion of borate into the
assay medium was responsible f o r the enhancement of the method.
In a l l the c o l o r i m e t r i c methods, galacturonic acid i s l i b e r a t e d
during the assay by hydrolysis of polymeric p e c t i n . An applica-
t i o n of the carbazole method, a f t e r e x t r a c t i o n of p e c t i n from
various f r u i t s and vegetables, has been described (6), as has
been an automated carbazole method f o r monitoring uronic acid
l e v e l s i n p e c t i n f r a c t i o n s (7).
A more rapid and somewhat simpler procedure f o r uronic acid
determination was described i n 1973 (8). This method i s advan-
tageous f o r determining galacturonic acid i n p e c t i n , as i n t e r f e r -
ence by neutral sugars i s reduced. This method i s based on the
color formation which accompanies the addition of m-hydroxybi-
phenyl to heated solutions of uronic acids i n s u l f u r i c acid/boric

In Chemistry and Function of Pectins; Fishman, M., et al.;

ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
 2. DONER Pectin Composition Determination 15

acid. This assay has been applied to the determination of gala-

cturonic acid i n food pectins (9,10), and interference by neutral
sugars was minimized (9). Another phenol, 3,5-dimethylphenol,
has been found (11) to be more s e l e c t i v e than m-hydroxydiphenyl
when large amounts of neutral sugars are present i n the sample.
This method has recently been employed i n studies of the degree
of methylation of pectin i n plant c e l l walls (12).
A procedure employing c o l l o i d a l t i t r a t i o n has been used f o r
the determination of galacturonic acid i n p e c t i n , and i n d i r e c t l y ,
also for determining the degree of e s t e r i f i c a t i o n (13). Samples
are t i t r a t e d with poly-N,N-dimethylallylammonium chloride, and a
d i s t i n c t f l o c c u l a t i o n occurs, the endpoint of which i s determined
by use of t o l u i d i n e blue i n d i c a t o r . In a duplicate sample, ester
methyl groups can be saponified, and t o t a l galacturonic acid
determined; by difference, the degree of methyl e s t e r i f i c a t i o n i s
calculated. The quantitation of t h i s c o l l o i d a l t i t r a t i o n method
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i s more precise with pectins of high degrees of polymerization.

In another t i t r i m e t r i c method, t o t a l galacturonic acid and the
degree of e s t e r i f i c a t i o n i s determined by copper-binding before
and a f t e r s a p o n i f i c a t i o n (14). The bound copper i s determined by
atomic absorption spectrometry. A p p l i c a t i o n of t h i s copper-binding
approach to the analysis of c e l l - w a l l polysaccharides i n many
f r u i t s and vegetables has been reported (15).
-
Decarboxylation with hydroiodic acid [16) was the basis f o r
a procedure used i n determining uronic acid l e v e l s i n dietary
f i b e r fractions (17). The carbon dioxide from decarboxylation
was p u r i f i e d , trapped i n a c e l l containing standard sodium hydro-
xide, and conductivity changes were measured using an Ingold
electrode.
Studies comparing the d i s t r i b u t i o n of free carboxyl groups
i n enzymatically and chemically d e - e s t e r i f i e d pectins are impor-
tant because the g e l l i n g behavior of r e s u l t i n g products i s a
function of the method used. Enzymatically d e - e s t e r i f i e d pectins
have a blockwise d i s t r i b u t i o n of non-esterified galacturonic acid
residues, and gel with calcium at higher degrees of e s t e r i f i c a t i o n
than do acid d e - e s t e r i f i e d pectins, which possess a more random
d i s t r i b u t i o n of free carboxyl groups. Free carboxyl d i s t r i b u t i o n
has been studied (18) by f i r s t e s t e r i f y i n g by reaction with
ethylene oxide ( g l y c o l a t i o n ) , and then t r e a t i n g the sample with a
mixture of pectin enzymes. The glycolated fragments are unreactive
toward these enzymes. F i n a l l y , the hydrolysis products are
separated from the glycolated fragments by ion exchange chromato-
graphy, and a f t e r deglycolation, chain s i z e i s determined by g e l
f i l t r a t i o n . An a p p l i c a t i o n of t h i s approach i n studies of orange-
peel pectin has been reported (19).

Physical Methods. Infrared (IR), Raman, and nuclear magnetic

resonance (NMR) spectroscopic methods have been applied to struc-
t u r a l analysis of polysaccharides such as pectin. These applica-
tions have been reviewed (20), and reference IR spectra of p e c t i c
substances have been published (21). Quantitative IR has been
used to estimate acid d i s s o c i a t i o n constants of polyuronides from
the r a t i o of -CCLH to -CO^- as a function of pH (22). Also, by

In Chemistry and Function of Pectins; Fishman, M., et al.;

ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
 16 CHEMISTRY AND FUNCTION OF PECTINS

use of s o l u t i o n IR i n D 0,2 the r a t i o of -C0 H to -C0 CH (free to

2 2 3

e s t e r i f i e d ) groups i n pectins can be determined (23). The ester-

1
carbonyl stretching band i s observed at 1740 cm and a carboxylate
1 3
-1
stretching band at 1650 cm . C-NMR spectroscopy has also been
useful i n determining r e l a t i v e proportions of free and e s t e r i f i e d
carboxyl groups i n pectin (24). In t h i s approach, the r a t i o of
peak areas at 172.8 ppm (-C0 H) i s determined r e l a t i v e to the
2

areas e i t h e r at 171.3 ppm (-C0 CH ) or 53.7 ppm (-0CH ). In

2 3 3

13
addition, pectin from sugar-beet has been examined by C-NMR,
and 0-acetyl, carbons-one of the minor constituents galactose and
arabinose, and carbon-six of rhamnose can be discerned (25).
Chromatographic Methods. The high l e v e l s of galacturonic acid
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(free and e s t e r i f i e d ) i n p e c t i n has resulted i n the development

of chemical methods which are rather non-specific. With correc-
tions applied for neutral sugars however, reasonable estimates of
i t s l e v e l s can be made. The c o l o r i m e t r i c procedures are conducted
i n strongly a c i d i c media, which r e s u l t s i n some loss of galactu-
ronic acid by decarboxylation to L-arabinose. Also, a l l uronic
acids respond to the various c o l o r i m e t r i c t e s t s , so for analyzing
mixtures of uronic acids, gentler h y d r o l y t i c steps and f a r more
s e l e c t i v e assays are needed. To t h i s end, several g a s - l i q u i d
(GLC) and high-performance l i q u i d chromatographic (HPLC) methods
have been developed for determining galacturonic acid. Some have
employed s p e c i f i c enzymes f o r depolymerization as an a l t e r n a t i v e
to acid hydrolysis. Combinations of p e c t i c enzymes and acid
c a t a l y s i s are required to q u a n t i t a t i v e l y hydrolyze p e c t i n . This
section w i l l describe applications of modern chromatographic
methods for determining galacturonic acid i n pectin. For excel-
lent and comprehensive descriptions of sugar chromatography i n
general, review a r t i c l e s on GLC (26) and HPLC (27,28) have been
published.
A s e n s i t i v e GLC procedure for determining galacturonic acid
i n p e c t i n has been developed (29) from an e a r l i e r described
method for analyzing uronic acids (30,31). Pectins, a f t e r extrac-
t i o n from plant tissues with ammonium oxalate, are depolymerized
with pectinase. The l i b e r a t e d galacturonic acid i s then reduced
by sodium borohydride to galactonic a c i d , which i s converted to
L-galactono-1,4-lactone. The t r i m e t h y l s i l y l d e r i v a t i v e of the
lactone gives a sharp peak on SE-30 stationary phase, and p e r - t r i -
m e t h y l s i l y l x y l i t o l i s used as an i n t e r n a l standard. Often, the
decomposition of monomeric sugars which r e s u l t from the resistance
to acid hydrolysis of polysaccharides containing a c i d i c groups
such as p e c t i n i s overcome by f i r s t reducing the uronic acids to
neutral sugars. This reduction method (32), which should be
repeated at l e a s t twice for quantitative conversion, i s widely
used i n polysaccharide s t r u c t u r a l a n a l y s i s , and has been applied
i n a GLC procedure for galacturonic acid (33). A f t e r a c t i v a t i o n
of p e c t i n carboxyl functions with a water-soluble diimide, reduc-
t i o n with sodium borodeuteride converts the galacturonic acid
residues to 6,6-dideutero-galactose d e r i v a t i v e s . Then, a f t e r
acid h y d r o l y s i s , standard gas chromatography-mass spectrometry

In Chemistry and Function of Pectins; Fishman, M., et al.;

ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
 2. DONER Pectin Composition Determination 17

methods to resolve neutral sugars are applied. The galactose

(dideutero) which had been generated from galacturonic acid i n
pectin i s distinguished from galactose ( d i p r o t i o ) n a t u r a l l y
present i n p e c t i n by mass spectrometry. These GLC methods are
c h a r a c t e r i s t i c a l l y very s e n s i t i v e and e f f i c i e n t .
Liquid chromatographic procedures have been developed more
recently, and some are quite e f f e c t i v e f o r determining galactu­
ronic acid i n p e c t i n . An automated anion-exchange chromatographic
system (34) allows the separation of i n d i v i d u a l uronic acids,
including galacturonic acid. Column e f f l u e n t s were s e n s i t i v e l y
analyzed for uronic acids by post-column reaction with o r c i n o l
and monitoring at 420 nm. More rapid HPLC approaches have been
described. By using strong anion-exchange columns, 0.7M acetic
acid mobile phase, and r e f r a c t i v e index detection, galacturonic
acid was separated from mannuronic and glucuronic acids i n less
than 15 minutes (35). S i m i l a r conditions were employed to separate
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oligogalacturonic acids of up to DP-8, and Bio-Gel P-2 has been

used f o r the same purpose (36). In another report (37), strong
anion-exchange HPLC also was used, with a mobile-phase consisting
of b o r i c acid/potassium hydroxide b u f f e r , and post-column f l u o r o -
metric detection (2-cyanoacetamide), allowing the r e s o l u t i o n and
s e n s i t i v e detection of four uronic acids, including galacturonic
acid. Polygalacturonic acid has been subjected to methanolysis
(reaction with methanolic hydrogen c h l o r i d e ) ; multiple peaks
r e s u l t i n HPLC chromatograms, due to the presence of α,β-mixtures
of the methyl glycosides (38). Although the procedure i s not
i d e a l f o r q u a n t i t a t i o n , i t i s useful f o r q u a l i t a t i v e analysis of
uronic acid composition i n polysaccharides. In a recent report
(39) polygalacturonic acid was subjected to both acid and polygalac­
turonase catalyzed hydrolysis. The hydrolyzates were analyzed by
+
HPLC on a cation-exchange column of HPX-87-H , and galacturonic
acid was eluted i n 8.5 minutes. The advantages of enzyme over
acid-catalyzed hydrolysis were apparent. The y i e l d of monomer
was greater, no monomer degradation products were present, and a
far lesser quantity of oligogalacturonic acid chains were produced.

Determination of Methyl, A c e t y l , and F e r u l o y l S u b s t i t u t i o n i n Pectin

Pectin consists mainly of polygalacturonate chains, and the

carboxyl groups are s i g n i f i c a n t determinants of i t s chemical and
b i o l o g i c a l properties. In plant c e l l w a l l s , more than 50% of the
carboxyl groups are often e s t e r i f i e d with methanol. The degree
of e s t e r i f i c a t i o n l a r g e l y determines the ion-exchange,
water-binding, c r o s s - l i n k i n g , and hydrogen-bonding capacities of
pectin. S i m i l a r l y , properties of p e c t i n i n c e l l walls are some­
times modified by low l e v e l s of hydroxyl e s t e r i f i c a t i o n with
acetyl groups. The d i s t r i b u t i o n of a c e t y l groups i n p e c t i n i s
unknown, but i n sugar beet, pear, and apricot p e c t i n , a c e t y l
levels approach 4%. In addition, a l k a l i - l a b i l e f e r u l i c acid
groups are found i n ester linkage to p e c t i n ; they are believed to
be c a r r i e d by arabinose and/or galactose residues on neutral side
chains. This section w i l l describe recent methods to determine
p e c t i n s u b s t i t u t i o n with methyl, a c e t y l , and f e r u l o y l groups.

In Chemistry and Function of Pectins; Fishman, M., et al.;

ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
 18 CHEMISTRY AND FUNCTION OF PECTINS

Methyl Esters In Pectin. A t i t r a t i o n method has been reported

(40), i n which methyl ester l e v e l s are calculated from the number
of equivalents of standard sodium hydroxide required to neuturalize
the pectin sample before and a f t e r saponification. The copper
t i t r a t i o n procedure described e a r l i e r for determination of galac-
turonic acid residues i n pectin (15), i s also used to determine
methyl ester l e v e l s from the increase i n copper-binding a f t e r
hydrolysis of the esters. An accurate and sensitive colorimetric
method (41) i s rather time-consuming, but several samples can be
run i n p a r a l l e l . Samples are saponified, the released methanol
oxidized to formaldehyde, and the formaldehyde determined by
spectrophometric assay (4l2nm) of i t s condensation product with
pentane-2,4-dione.
GLC procedures are widely used for methyl ester determina-
t i o n s ; a f t e r saponification with 0.5N base, methanol i s measured
by GLC on columns of Poropak Q at 120°C (42), or on Carbowax 1500
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at 125°C (12). In the l a t t e r study, analyses were conducted on

small samples of i s o l a t e d plant c e l l w a l l preparations. In
studies on the enzymatic incorporation of methyl groups from
S -adenosyl-L-methionine into pectin (43), ^C-methyl-labelled
14
substrate was used. The C-methanol, a f t e r release from p e c t i n ,
was determined by GLC on Carbowax 300 using a r a d i o a c t i v i t y
counting detector. The coupling of a n a l y t i c a l p y r o l y s i s to GLC
has resulted i n the detection of c h a r a c t e r i s t i c fragments from
macromolecules, such as pectin. This topic has been reviewed
(44), and correlations between degree of methyl e s t e r i f i c a t i o n
and i n t e n s i t y of some of the peaks have been made (45).
Acetyl and F e r u l o y l Esters i n Pectin. A colorimetric method f o r
determining degrees of acetylation i n pectins from various sources
(46), has been shown to be rapid and quite s e n s i t i v e . Hydroxy-
lamine i s reactive toward both the methyl and acetyl esters i n
p e c t i n , and f e r r i c ion complexes with the r e s u l t i n g hydroxamic
acids are red. The pectin complex i s insoluble and removed by
f i l t r a t i o n ; the i n t e n s i t y at 520nm i n the soluble f r a c t i o n ,
consisting of the f e r r i c complex with acetohydroxamic a c i d , i s a
measure of acetyl content. The accuracy of the method was demon-
strated i n determinations of 0-acetyl l e v e l s i n standard per-ace-
t y l a t e d polysaccharides. Another method (47) involves a l k a l i n e
hydrolysis of the acetyl groups from p e c t i n , followed by d i s t i l -
l a t i o n of acetic acid and i t s t i t r a t i o n with standard base.
In a study of the structure and functions of feruloylated
pectins i n primary c e l l walls i n spinach, about one f e r u l o y l
group was found per s i x t y sugar residues (48). F e r u l i c acid was
determined a f t e r a l k a l i n e hydrolysis by the Folin-Ciocalteu
phenol reagent.

Neutral Sugar Composition of Pectin

The neutral sugars, with the exception of L-rhamnose, are attached

e x c l u s i v e l y i n sidechains, and include D-galactose, L-arabinose,
D-xylose, and less frequently, D-glucose, D-mannose, L-fucose,
2-0-methyl-D-xylose, 2-0-methyl-D-fucose, and D-apoise. Whether

In Chemistry and Function of Pectins; Fishman, M., et al.;

ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
 2. DONER Pectin Composition Determination 19

the sugars are determined by GLC or HPLC methods, i t i s e s s e n t i a l

that the polysaccharide f i r s t be hydrolyzed to i t s monomeric
sugars. Acid-catalyzed h y d r o l y t i c methods are most often used,
but the various linkages have d i f f e r e n t s u s c e p t i b i l i t i e s , as do
the various sugars when released upon hydrolysis. These problems
have been discussed, along with a review of the sugar GLC l i t e r a -
ture p r i o r to 1973 (49). I t was stated that "no one method of
hydrolysis w i l l necessarily cleave every linkage and give each
component i n quantitative y i e l d . " The Saeman hydrolysis (50),
which employs 72% s u l f u r i c a c i d , or 2N t r i f l u o r a c e t i c acid"T51)
are used most often. When p o s s i b l e , enzymatic approaches i n
combination with acid hydrolysis are preferred f o r polysaccharide
hydrolysis. A f t e r h y d r o l y s i s , the most widely used methods for
sugar determination are based on GLC of s u i t a b l y v o l a t i l e deriva-
t i v e s . The derivatives i n which the anomeric center i s eliminated
so that s i n g l e peaks r e s u l t are most e f f e c t i v e .
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Single-peak sugar derivatives which allow the r e s o l u t i o n of

sugar constituents i n p e c t i n include the t r i m e t h y l s i l y l a t e d
methyloximes (52), acetylated a l d o n o n i t r i l e s (53), t r i m e t h y l s i l y -
lated a l d i t o l s " T 5 4 ) , and acetylated a l d i t o l s (51). A comprehen-
sive review a r t i c l e on GLC of sugars has been published (26).
In studies of polysaccharides structure, the a l d i t o l acetate
procedure remains the most widely used GLC procedure. The advent
of high-resolution glass c a p i l l a r y columns has allowed very
e f f i c i e n t separations. Recent applications of these columns to
a l d i t o l acetate separations have been described (55-57). The
a l d i t o l acetate procedure requires reduction of the sugars with
sodium borohydride. A f t e r removal of boric a c i d , the sample i s
acetylated by conventional means. Various polar stationary phases
have been used i n GLC separation of a l d i t o l acetates, i n both
packed and c a p i l l a r y columns. A low-polarity phase was used i n a
report (54) which demonstrated the separation of t r i m e t h y l s i l y l a t e d
a l d i t o l s , and the neutral sugars i n a hemicellulose sample were
resolved.
A l i q u i d chromatographic system has been applied i n a study
of monomer composition i n c e l l - w a l l polysaccharide hydrolyzates
(58). A strong-base anion-exchange column was eluted with a
borate buffer step-gradient. Post-column reaction with o r c i n o l
allowed the s e n s i t i v e determination of the sugars rhamnose,
xylose, arabinose, glucose, galactose, and mannose. An improved
two-step HPLC procedure f o r t o t a l r e s o l u t i o n of the above neutral
sugars has been published (59). On aminopropyl s i l i c a with
acetonitrile-water as mobile phase, rhamnose i s separated, but
the pentoses xylose and arabinose are not w e l l resolved, nor are
the hexoses glucose, galactose, and mannose. These two m u l t i -
component peaks are c o l l e c t e d and re-chromatographed on Aminex
HPX-87P, a heavy metal cation-exchanger i n the lead form. Total
resolution of the f i v e monosaccharides r e s u l t s .

Acknowledgment

R e f e r e n c e to a brand or f i r m name does not c o n s t i t u t e endorsement by

the U.S. Department of A g r i c u l t u r e over o t h e r s of a s i m i l a r n a t u r e
not mentioned.

In Chemistry and Function of Pectins; Fishman, M., et al.;

ACS Symposium Series; American Chemical Society: Washington, DC, 1986.
 20 CHEMISTRY AND FUNCTION OF PECTINS

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RECEIVED December 20, 1985

In Chemistry and Function of Pectins; Fishman, M., et al.;

ACS Symposium Series; American Chemical Society: Washington, DC, 1986.