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Coffee Carbohydrates

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This review summarises recent advances in the chemistry, physiology and molecular properties of coffee carbohydrates with a particular focus on the cell wall polysaccharides. The results of detailed chemical studies have demonstrated novel structural features of both the galactomannans and the arabinogalactan polysaccharides of the green and roasted coffee bean. For the first time immunological probes based on monoclonal antibodies for specific polysaccharide epitopes were used to reveal the patterns of distribution of the galactomannans, arabinogalactans and pectic polysaccharides in the coffee bean cell wall. Finally, the results of physiological and molecular studies are presented which emphasise the growing awareness of the potential role the metabolic status of the green bean may play in final coffee beverage quality.
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M I N I R E V I E W
Coffee carbohydrates
Robert Redgwell* and Monica Fischer
Nestlé Research Centre, Lausanne, Vers-Chez-Les-Blanc, P.O. Box 44, CH-1000 Lausanne 26, Switzerland. *Corresponding author:
robert.redgwell@rdls.nestle.com

This review summarises recent advances in the chemistry, physiology and molecular properties of coffee carbohydrates with a
particular focus on the cell wall polysaccharides. The results of detailed chemical studies have demonstrated novel structural
features of both the galactomannans and the arabinogalactan polysaccharides of the green and roasted coffee bean. For the
first time immunological probes based on monoclonal antibodies for specific polysaccharide epitopes were used to reveal the
patterns of distribution of the galactomannans, arabinogalactans and pectic polysaccharides in the coffee bean cell wall. Finally,
the results of physiological and molecular studies are presented which emphasise the growing awareness of the potential role
the metabolic status of the green bean may play in final coffee beverage quality.
Key words: Coffea, cell wall, beverage quality, seed.
Carboidratos do café: Este artigo sumariza os mais recentes desenvolvimentos nas áreas de química, fisiologia e propriedades
moleculares dos carboidratos do café, com um particular interesse nos polisacarídeos presentes nas paredes celulares. Os
resultados dos estudos químicos detalhados demonstraram novas características estruturais tanto nos galactomananos como
nos polissacarídeos dos arabinogalactanos do grão de café verde ou torrado. Pela primeira vez, estudos imunológicos baseados
em anticorpos monoclonais foram usados para revelar a distribuição dos galactomananos, dos arabinogalactanos e dos
polissacarídeos pectídicos na parede celular do grão de café. Finalmente, os resultados dos estudos fisiológicos e moleculares
são apresentados de maneira a sublinhar a influência do status metabólico do grão verde do café na qualidade final da bebida.
Palavras-chave: Coffea, parede celular, qualidade da bebida, semente.
INTRODUCTION
development. However, to date there is very little published
The importance of carbohydrates in coffee can be
literature relating to the carbohydrate physiology of coffee.
attributed to not only their high concentration in the bean but
What there is will be covered in the present review but of
also to the complex changes they undergo during the roasting
necessity the major focus will be the polysaccharides of
process which contribute to the organoleptic appeal of the
the coffee bean cell wall, where significant advances in
coffee beverage. In this review we will focus on the coffee
understanding their chemistry, biochemistry and distribution
carbohydrate literature of the last 5 years. Before this time,
within the endosperm have been made in the last 5 years.
an excellent summary of coffee carbohydrates was reported
in the volume, “Coffee, Recent developments” (Bradbury,
Carbohydrate status of developing grains
2001). It included a detailed report of the low molecular
weight sugars in green and roasted coffee beans of many
Free sugars: The driver for coffee research has been to
varieties of Arabica and Robusta, which will not be repeated
consider the various components of the coffee grains as
in the present review. In the last few years interest has grown
potential precursors of coffee beverage flavour and aroma.
in the physiology and biochemistry of green coffee bean
Whether focusing on the low molecular weight sugars or
development and the role this could play in coffee quality.
the polysaccharides, the great majority of such studies have
This includes work to understand sugar metabolism in
been done on the mature grains of Arabica and Robusta
coffee in relation to sink-source relationships during bean
coffee. There is a lack of information on the evolution of
Braz. J. Plant Physiol., 18(1):165-174, 2006

166
R. REDGWELL and M. FISCHER
the carbohydrate profile of the grain during its growth and
than invertase functioning more towards sucrose degradation
development and the postharvest stages of bean processing.
than synthesis. This was supported by the fact that no expres-
The metabolic status of the green bean at these stages will
sion of invertase-encoding genes was observed in any of the
affect the final chemical composition of the mature green
tissues tested. The Susy activity peaked in the final stages of
bean and the influences which modulate this metabolic status
perisperm development suggesting that the tissue played a
are factors which impinge on coffee bean quality.
role in controlling bean size and in the build up of sugars in
Rogers et al., (1999) conducted a study of the changes
the green bean. At the molecular level 2 cDNAs were cloned
to the content of sugars, and sugar alcohols in developing
encoding Susy isoforms which showed differences in their
grains from varieties of Robusta and Arabica. The grains
spatial and temporal expression in coffee fruits.
were harvested between 12–30 weeks after flowering (WAF)
The importance of seed metabolism during postharvest
for Arabica and 18-40 WAF for Robusta. In the early stages
processing methods as it relates to coffee quality has been
of development, up to the halfway stage of maturation,
emphasised by Mazzafera and Purcino (2004). They drew
glucose and fructose were the major free sugars with glucose
attention to the postharvest physiology of the coffee bean
consistently twice the concentration of fructose. Glucose
and described changes to several groups of metabolites,
levels were higher in the Arabica varieties (between 8-12 %
including the carbohydrates, which are likely to be influenced
dry weight) than in Robusta (2-4 % dry weight). At the end
by the nature of the postharvest processing methods. Just
of grain development concentrations of glucose and fructose
to what extent the final beverage quality is influenced by
had decreased for both species to 0.03 and 0.04 % dry weight
the metabolic status of the bean at harvest has yet to be
respectively, while sucrose at 5-12 % of the dry weight was
determined but it remains an area which needs more attention
essentially 100 % of the total free sugars in mature grains.
from coffee researchers.
A more detailed analysis in one variety of Arabica was
done by separating the perisperm tissues from the endosperm
Polysaccharides: The polysaccharides which make up ~50
and analysing the free sugar concentration in each. The
% of the green bean’s dry weight, consist of three major
higher concentrations of glucose and fructose compared
types: mannans or galactomannans, arabinogalactan-proteins
to sucrose in the early stages of development were always
(AGPs) and cellulose. In addition, there are small amounts of
associated with perisperm tissue. In the endosperm even at
pectic polysaccharides (Redgwell et al., 2002a) and recently
the earliest stages of maturation, sucrose was the dominant
xyloglucan was also shown to be present (Oosterveld et al.,
sugar.
2003).
The authors speculated that the catabolism of sucrose
in the perisperm would be consistent with the requirement
Galactomannans: If there is little information on the biosyn-
by the tissue for an increase in osmotic pressure to enable
thesis of free sugars during coffee bean development there is
both the initial expansion within the locular space and a sink
even less on the biosynthesis of the cell wall polysaccharides.
function.
Nevertheless, with the advent of molecular techniques there
Results of the analysis of sugar alcohols (mannitol)
is now heightened interest in the mode of biosynthesis of
and oligosaccharides (raffinose, stachyose) did not show
coffee cell wall polymers. This is particularly pertinent in
any discernible trend that would indicate fundamental
relation to the galactomannans, the solubilisation of which
changes to the metabolism of these compounds during grain
is a critical factor in determining the yield of soluble coffee
development.
powder during commercial extraction (Clifford, 1985). Cel-
Another recent study (Geromel et al., 2004) investigated
lulose apart, the most resistant polymers to solubilisation are
the biochemical and molecular characterisation of sucrose
the galactomannans. One of the principal determinants of
synthase (Susy EC2.4.1.13) and invertase during coffee
galactomannan solubility is the frequency of substitution of
bean development. The objective of this work was to under-
the mannan backbone with galactose residues. In theory an
stand sugar metabolism in coffee in relation to sink-source
increase in the degree of galactosylation of the mannans may
relationships during bean development. To this end they
increase the degree of solubilisation of the galactomannans.
measured the sugar concentrations and activities of the two
In order to manipulate the final structure of the ga-
enzymes in the pulp, perisperm and endosperm from Coffea
lactomannans it is necessary to understand the metabolic
arabica at stages of bean development. Susy was more active
steps involved in their synthesis. In particular, what are the
Braz. J. Plant Physiol., 18(1):165-174, 2006

COFFEE CARBOHYDRATES
167
crucial steps which dictate the final degree of galactosyla-
reasonable assumption that the arabinose and galactose are
tion? It is known that in some plants, the final degree of
derived mostly from the arabinogalactans and the rhamnose
galactosylation is determined by the action of an ?-galac-
and galacturonic acid are structural components of the pectic
tosidase which cleaves galactosyl residues from the prima-
polysaccharides, some discernible trends can be deduced
ry synthetic product. If such a mechanism operated in the
for the structural features of these polymers during grain
coffee bean, then down regulation of the ?-galactosidase
development. The Gal/Ara ratio of the arabinogalactans
gene could result in coffee beans with a higher Gal/Man
in the earliest stage of growth was 1.3:1 but this gradually
ratio. Fischer et al. (1999) determined the monosaccharide
increased during grain growth and reached 2.6:1 at maturity.
composition of coffee bean cell walls 12, 17 and 29 WAF
In addition, at the earliest growth stage the arabinogalactan
and reported that early in development the galactomannans
accounted for ~50 % of the total polysaccharides but this
were more highly substituted than at maturity. In a more
decreased to 34 % in the mature grain.
detailed study Redgwell et al. (2003), isolated and charac-
In the case of the pectic polysaccharides the endosperm
terised galactomannans from the endosperm of coffee beans
of the youngest growth stage contained ~20 % of its weight
11, 15, 21, 26, 31 and 37 WAF. At the earliest stage of de-
as pectic polymers, which dropped dramatically in the mature
velopment the galactomannans accounted for ~10 % of the
grain (~4 %). This is to be expected, as the earliest formed
polysaccharides but were highly substituted, with Gal/Man
layer during the period of rapid growth and cell division is
ratios between 1:2 and 1:7. At maturity the galactomannan
the cell plate, which is rich in pectic polymers.
became the predominant polysaccharide accounting for ~50
In summary during growth and development of the
% of the total endosperm polysaccharides but their degree
coffee bean cell wall there is a progressive change in
of substitution decreased with Gal/Man ratios between 1:7
both the relative content of the different polysaccharide
and 1:40. The decrease in the Gal/Man ratio of the galacto-
types and their structural features. At the earliest stages
mannans commenced between 21 and 26 WAF and was in
of growth, cellulose and arabinogalactan appear to be the
synchrony with a rise in free galactose. It was concluded
primary products of cell wall synthesis with the former the
that the final Gal/Man ratio was to an extent the result of
predominant polysaccharide (Fischer et al., 1999). During
galactose removal from the primary synthetic product by an
the middle stages of growth, cellulose synthesis appears to
?-galactosidase.
cease and there is a progressive increase in mannan synthesis
relative to the other wall polysaccharides as the grain
Other polysaccharides: Of the other polysaccharides in the
approaches maturity. The close stereochemistry of cellulose
coffee bean cell wall little has been reported on the subject
and mannan prompts the speculation that perhaps the same
of developmental changes to their structural features. In the
catalytic membranes, which lead to cellulose synthesis, are
same study which reported developmental changes to the
also involved in mannan synthesis later in the growth of the
galactomannans (Redgwell et al., 2003) data was published
endosperm with the additional intervention of an enzyme
for the monosaccharide composition of the cell wall material
capable of interconverting GDP-mannose and GDP-glucose
at different stages of development (table 1). Making the
such as GDP-mannose-2-epimerase.
Table 1. Monosaccharide composition of alcohol-insoluble residue from coffee bean endosperm at several stages of
development
WAF
Monosaccharide composition (mole %)
Rha
Ara
Xyl
Man
Gal
Glc
GlcA
GalA
11
3.7
18.9
1.9
9.3
23.9
21.7
6.6
14.0
15
3.7
17.5
3.0
13.4
25.7
20.8
4.7
11.0
21
1.7
14.7
0.5
19.6
33.1
21.7
3.3
5.3
26
0.5
10.1
0.4
37.0
27.7
18.2
2.4
3.6
31
0.5
8.9
1.1
45.4
23.0
15.9
2.1
3.2
37
0.6
9.1
1.0
45.6
23.2
15.7
1.9
3.1
(Redgwell et al., 2003)
Braz. J. Plant Physiol., 18(1):165-174, 2006

168
R. REDGWELL and M. FISCHER
Molecular and biochemical characterisation of polysac-
probes showed that mRNA transcripts for both cDNAs
charide modifying enzymes
were present during the same periods of bean germination,
Once the function and expression of the enzymes
with expression peaking 20 days after imbibition of water.
responsible for the biosynthesis and breakdown of coffee
Activity and mRNA levels appeared to be tightly coordinated
polysaccharides is understood, targeted manipulation of the
and unlike the reports of Dirk et al. (1995) they stated that
morphological properties of the bean becomes a realistic
enzyme activity did not exist in grains prior to germination.
proposition. To date there has been little published data
Thus, transcripts of the enzyme were not detected during
on the biochemical or molecular characterisation of the
grain maturation or in other tissues of the plant (roots,
endogenous enzymes governing polysaccharide metabolism
leaves, stems, flowers). The enzyme showed no activity with
in the coffee bean. Those that have been targeted include
mannotriose or mannobiose and required oligomers with at
endo-?-mannanase and ?-galactosidase, two enzymes which
least 5 or more units for maximum efficiency.
mediate changes to the galactomannans. Biochemical studies
Despite the obvious potential to manipulate the physico-
involving endogenous enzymes of coffee, which specifically
chemical properties of coffee beans by using transformation
promote metabolic changes to the arabinogalactans, have not
technology, which can change the polysaccharide structure of
been reported.
the grain cell wall, it is unlikely that the approach will yield
The galactomannans play a dominant role in the physi-
any significant benefits to the coffee industry in the short
cochemical properties of the coffee grain, a major factor
to medium term. Primarily this is because the relationship
influencing industrial extractability. The two major enzymes
between genetically induced changes and the biochemistry,
concerned with modification of the galactomannans are ?-
physiology and quality traits of the coffee bean is far from
galactosidase and ??(1?4) endo-mannanase. Both have
being elucidated. In addition, since the FLAV SAVR tomato
been fairly well characterised in relation to their biochemical
became the first genetically engineered whole fruit to become
and molecular properties. Zhu and Goldstein (1994) reported
commercially available, the application of biotechnology in
the cloning and functional expression of a cDNA encoding
agriculture has been intensively discussed, and consumer
coffee bean ?-galactosidase and demonstrated that the en-
polls, particularly in Europe, have shown a general ambiva-
zyme had a preference for ?-1,3- and 1,4-glycosidic link-
lence and some hostility in attitudes (Schibeci et al., 1997).
ages. If, as the results of Redgwell et al. (2003) suggested,
?-galactosidase is involved in the determination of the final
Chemistry of the polysaccharides
galactose content of coffee endosperm galactomannans, then
Advances in our knowledge of the chemistry of coffee
potentially, coffee plants transformed by down regulation
bean polysaccharides during the last 5 years has focused
of the ?-galactosidase gene could contain galactomannans
primarily on elaborating the detailed structural features of
with a higher degree of substitution than the wild type. The
the arabinogalactans and the (galacto)-mannans. Some ad-
opposite effect has already been demonstrated by Joersbo et
ditional information has also been revealed on the structural
al. (2001), who cloned and transformed the ?-galactosidase
features of the pectic and hemicellulosic polysaccharides.
gene expressed in immature senna seeds into a species of
The availability of a range of immunological probes, which
guar (Cyamopsis tetragonoloba). Approximately 30 % of the
are specific for certain epitopes of the polysaccharides has
guar transformants produced endosperm with galactoman-
permitted localisation studies to reveal additional informa-
nans where the galactose content was significantly reduced.
tion on the architecture of the coffee bean cell wall. To date
Until recently reports on endo-?-mannanase activity
there has been no published work on the characteristics of
in coffee grains were limited to two studies. Dirk et al.,
coffee cellulose.
(1995) reported multiple isozymes of the enzyme in dry and
imbibed seeds while Giorgini and Comoli (1996) measured
Arabinogalactans: Several structural studies have revealed
the effect of growth regulators on the activity of the enzyme
that the type II arabinogalactans in the coffee bean consist
during germination. The first molecular characterisation
for the most part of a backbone of ? ?(1?3) - linked galac-
was reported by Marraccini et al. (2001) who cloned and
tosyl residues, substituted at intervals in the 0-6 position with
sequenced two endo-?-mannanase cDNAs (man A and
various combinations of arabinosyl and galactosyl residues
man B) from germinating coffee grains (Coffea arabica L.).
(Bradbury, 2001). Redgwell et al., (2002a) reported two
Northern hybridizations with man A- and man B-specific
important additional pieces of structural information. Firstly,
Braz. J. Plant Physiol., 18(1):165-174, 2006

COFFEE CARBOHYDRATES
169
that the polysaccharides carried a negative charge due to the
possibility is that the compact structure of the coffee cell wall,
presence of up to 10 % of their structure as glucuronosyl
which is made up mostly of the insoluble polymers cellulose
residues which occurred as non-reducing terminal units on
and mannan, entraps much of the AGP within its structure,
a 1?6 linked galactosyl side chain. Secondly, all or some
rendering it effectively insoluble. Evidence to support this
of the arabinogalactans are in fact arabinogalactan-proteins
idea was provided by Redgwell et al. (2002a) who were able
(AGPs). The existence of a covalent link between the arabi-
to solubilise almost all the AGPs in green beans by treating
nogalactan moiety and protein was premised on the contin-
the insoluble residue remaining after 8 M KOH treatment
ued association of carbohydrate and protein during purifica-
with a mixture of cellulase and mannanase enzymes. The
tion, the positive reaction to the ?-glucosyl-Yariv reagent and
8 M KOH treatment was necessary to render the mannan/
the amino acid composition of the protein moiety which was
cellulose polymers more susceptible to the enzymes and
hydroxy-proline rich, a characteristic of many reported AGPs
probably did this by causing the cellulose/mannan fibrils to
(Clarke et al., 1979). The protein content of three different
swell, making them more accessible substrates.
AGP fractions isolated from Arabica Yellow Caturra was 0.4,
Redgwell et al. (2002a) reported that the AGPs existed
1.1 and 1.9 %. The AGPs were shown to exist in several vari-
as an extremely heterogeneous mixture containing between
eties of both Arabica and Robusta coffee beans.
6-10 % glucuronic acid and possessing a Mw average of
Previous studies characterised coffee arabinogalactans,
~650 kDa. The heterogeneity related particularly to their de-
which represented only fractions of the total cell wall polymer.
gree of branching and monosaccharide composition of their
This can be attributed to the fact that the arabinogalactans
side chains. Five different AGP fractions were isolated with
are not readily extractable from green beans despite the fact
Gal/Ara ratios which varied markedly. For Arabica Caturra,
that they and AGPs in general are extremely water-soluble.
Catimor and Sarchimor the Gal/Ara ratios ranged from 0.9 to
Bradbury and Halliday (1990) used 20 % NaOH at 100°C to
3.1, 1.5 to 3.2 and 1.2 to 3.0, respectively. For Robusta Indes,
extract arabinogalactan from green beans and reported that 45
Conillon and Ivoire the values were 0.9 to 3.1, 1.1 to 3.0 and
% remained in the insoluble fraction. Fischer et al. (2001a,b)
0.9 to 3.1, respectively. The putative structural features of the
used a sequence of extractants, which included 8M KOH
major arabinogalactan fraction which was liberated only fol-
and NaClO with similar results. Oosterveld et al., (2003)
lowing enzyme treatment of the insoluble residue of CWM
2
used water, EDTA and 4 M NaOH and released less than
isolated from Arabica Yellow Caturra, is given in figure 1.
10 % of the polysaccharides. Bradbury (2001) speculated
The wide heterogeneity of coffee arabinogalactans
that the insolubility of coffee arabinogalactan was evidence
supported the earlier findings of Fischer et al. (2001a,b)
that it was covalently linked to a less soluble component of
which reported that Robusta contained a highly soluble ara-
the cell wall (e.g. cellulose or mannan). However, a second
binogalactan which possessed more branch points and more
Figure 1. Possible structure of arabinogalactan moiety of coffee AGP (Redgwell et al., 2002a).
Braz. J. Plant Physiol., 18(1):165-174, 2006

170
R. REDGWELL and M. FISCHER
extended side chains than those found in Arabica and argued
of other moieties (e.g. acetyl groups) could also disrupt
that this may be the reason why the arabinogalactans from
interchain hydrogen bonding and may be an explanation for
Robusta are more easily solubilised than those of Arabica.
why some galactomannans with an apparently low degree
of galactose substitution are relatively soluble. Oosterveld
Galactomannans: The galactomannans are the predominant
et al. (2004) reported that galactomannans extracted from
components in the coffee bean cell wall accounting for 50
green Arabica coffee beans were acetylated. In this study the
% of the polysaccharides. Recent research has provided
galactomannans were extracted in water at 90°C for 1 h. The
information on the degree of galactosylation of the mannans,
galactomannans were separated into two neutral fractions
the presence and distribution of other substituents (e.g.
by anion-exchange chromatography which suggested that
acetyl groups), the possibility that other sugar residues exist
other mechanisms such as molecular weight must also
in the primary structure of the molecule (e.g. arabinose and
have played a part in the separation. One neutral fraction of
glucose) and the location of the mannans in the endosperm
average molecular weight of 2000 kDa possessed both a high
of the cell wall.
degree of galactosylation (30 %) and acetylation (9 % of the
Coffee bean mannan consists for the most part of linear
mannose groups were acetylated). The second neutral fraction
chains of ?1?4-mannosyl residues with single galactose
had a molecular weight average of 20 kDa and the degree of
units ?-linked at C-6 of a mannosyl residue. The literature
galactosylation and acetylation was much lower (11 % and
reports wide-ranging degrees of substitution from 47:1
4 % respectively). The results of Oosterveld et al. have been
(Wolfrom and Patin, 1961), 130:1 (Bradbury and Halliday,
reinforced in a study by Nunes et al. (2005) who reported that
1990), 30:1 (Fischer et al., 2001a), 1:7 and 1:40 (Redgwell
the galactomannans in hot water infusions of green coffee
et al., 2003) and 3:1 and 9:1 (Oosterveld et al., 2004). It
beans are acetylated at a level of 11 mole %. They provided
is likely that the mannan molecules in coffee consist of a
evidence that the acetyl groups were attached to the O-2 and
heterogeneous mixture of substituted and unsubstituted
O-3 positions (sometimes both) on the mannose residue.
polymers but definitive data on the exact nature of this
Contiguously acetylated mannosyl residues were also found.
mixture is not available. One of the reasons for this is that
Another possibility for disrupting interchain hydrogen
most of the linkage analyses for coffee mannans have been
bonding would be the interruption of the mannan backbone
done only on fractions of the total wall mannan, usually
with glucose residues and/or the substitution of the mannan
those which can be solubilised by various forms of solvent
backbone with sugar residues other than galactose. Navarini
extraction. Invariably they consist of the more galactosylated
et al., (1999) reported the possibility that the galactomannans
galactomannan fractions which are more readily soluble than
were substituted with arabinose. These structural features
the less substituted molecules which remain in the insoluble
were looked for, but not found, in the Oosterveld et al. study
residue. This is confirmed by the study of Redgwell et al.,
(2004). However, the Nunes et al. report (2005) provided
(2003) where almost all the mannan was solubilised from
evidence that terminally linked arabinosyl residues (2 mole
mature grains by a combination of chemical and enzymatic
%) were attached at O-6 of the mannose residues. In addition,
treatments. The chemically solubilised polymers (8 M KOH
they stated that ?-(1?4)-linked glucosyl residues (6 mole
) possessed Man/Gal ratios of 7:1. The 8 M KOH-insoluble
%) were present in the mannan backbone and concluded that
residue was solubilised by enzymic hydrolysis with a
green coffee mannans extracted with hot water contained
mixture of cellulase and mannanase. The arabinogalactan
acetylated arabinogalactoglucomannans.
which was solubilised at the same time was easily removed
by dialysis allowing the Man/Gal ratio (40:1) of the
Cytochemical and immunolabelling of cell wall polysac-
galactomannan fragments derived from the insoluble residue
charides
to be determined.
Chemical analysis has revealed an increasingly sophisti-
A pure mannan is able to form a hard insoluble structure
cated picture of the coffee cell wall polymers. Just how these
much like cellulose because of interchain hydrogen bonding.
different types of polysaccharide and different structural
One of the principal effects of galactose substitution is to
forms of the same type of polysaccharide, contribute to the
disrupt the interchain hydrogen bonding and this can lead
architecture of the wall has until recently been largely un-
to increased solubility. However, galactose is not the only
known. Sutherland et al. (2004) used a range of cytochemical
substituent, which could induce such an effect. The presence
and immunological probes to reveal the spatial arrangement
Braz. J. Plant Physiol., 18(1):165-174, 2006

COFFEE CARBOHYDRATES
171
of the arabinogalactan-proteins, galactomannans and pectic
Mannans: When mannans were labelled with the ?-1,4-man-
polysaccharides in the cell wall of the endosperm of green
nan-specific monoclonal antibody BGM C6, the antibody
coffee beans (Coffea arabica L.Yellow Caturra).
labelled across the entire wall. However, there was a vari-
ation in intensity of the labelling across the wall with more
Arabinogalactan-proteins: AGPs were localised by labelling
intense staining adjacent to the lumen of the cell and the
with the AGP-specific ?-glucosyl Yariv reagent and the
middle lamella. These two zones were separated by a re-
monoclonal antibody LM2 (Sutherland et al., 2004) which
gion of only moderately intense staining (Sutherland et al.,
recognises a carbohydrate epitope containing glucuronic acid.
2004). Evidence that galactomannans with different degrees
Glucuronic acid has been shown to occupy terminal positions
of galactosylation were located at different sites in the wall
on some of the side chains of coffee AGPs (Redgwell et
was provided by the use of BS-1 lectin, which is specific for
al., 2002a). Both forms of labelling showed a widespread
distribution of the AGP across the cell wall. However, there
was more intense staining with the Yariv reagent in the region
adjacent to the cell lumen. The labelling pattern for LM6,
a monoclonal specific for several contiguous arabinosyl
residues, was quite different to that of LM2. Whereas LM2
labelled across the whole wall, LM6 was found in two
specific locations (figure 2). LM6 gave intense labelling of
the epidermal cells across the whole width of the cell wall
indicating that these cells were enriched in 1,5-?-arabinan
compared to the endosperm cells. The second location was in a
compact band adjacent to the cell wall lumen of the endosperm
cells. No label was found in the main body of the cell wall.
This indicated the existence of a different structural form of
arabinan polymer in the region adjacent to the cell wall lumen,
which was absent in the rest of the cell wall. Since the Yariv
reagent also showed increased staining in this location one
explanation could be that the AGPs in this region have more
1,5-?-arabinosyl residues incorporated into their side chains.
However, all antibody data must be interpreted with caution,
as there is a possibility that the antibody LM6 is reacting with
similar epitopes on completely unrelated molecules (e.g.
rhamnogalacturonans and AGPs). An alternative explanation
is that LM6 was labelling a rhamnogalacturonan type molecule
which carried side chains of 1,5-?-arabinosyl residues.
The presence of rhamnogalacturonans in coffee bean cell
walls which contain moderate amounts of 1,5-?-arabinosyl
residues has been demonstrated (Redgwell et al., 2002a). The
argument for the arabinosyl residues being structural features
of some pectic polysaccharides is supported by the fact that
LM6 does not cross react with the AGP in gum acacia. On the
other hand chemical analysis of the coffee bean does indicate
the presence of a mixture of AGPs which are polydisperse with
Figure 2. Localisation of 5-linked arabinan with monoclonal
regard to their 1,5-arabinosyl residue content. In addition, the
antibody LM6 (Sutherland et al., 2004. Upper: Low
inhibition of LM6 labelling by similar concentrations of pure
magnification showing intense labelling of epidermal
1,5 arabinan and a pectin-depleted AGP fraction from coffee,
layer. Lower: High magnification showing labelling of
layer adjacent to the cell lumen.
suggested that coffee AGPs do cross-react with LM6.
Braz. J. Plant Physiol., 18(1):165-174, 2006

172
R. REDGWELL and M. FISCHER
terminal ?-galactose (Hayes and Goldstein

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