C H A P T E R

20
Melanoidins in Coffee
José A. Rufián-Henares, Silvia Pastoriza
Department of Nutrition and Food Sciences, School of Pharmacy, University of Granada, Campus Cartuja S/N,
Granada, Spain

List of Abbreviations
MR  Maillard reaction
HMW  High-molecular weight
LMW  Low-molecular weight
GMNs Galactomannans
AGLs Arabinogalactans
AGPs  Arabinogalactan proteins
MRPs  Maillard reaction products
20.1 INTRODUCTION
Coffee, one of the most consumed drinks in the world
and with a distinct aroma and flavor, is prepared from the
roasted seeds of the Coffea plant.1 Different compounds
found in green coffee beans are degraded/transformed
to some extent, giving rise to a decrease of the amount of
carbohydrates and chlorogenic acids and the appearance
of melanoidins.2 Melanoidins are a heterogeneous group
of nitrogen-containing brown polymers produced during
the last steps of the Maillard reaction (MR).3 Coffee mela-
noidins are responsible for coffee’s characteristic brown
color because they can absorb light at high wavelengths
such as 420 nm.4 This absorbance is useful in evaluating
the melanoidins content in coffee brew,5 although other
methods, such as quantification by difference, also are
possible.6 In the case of roasted coffee beans, the melanoi-
din content accounts for up to 25%,2 although this content
increases with stronger roasting processes.7 This chapter
focuses on the structure and composition of melanoidins
and the mechanisms of their formation.
20.2  CHEMICAL COMPOSITION
As stated above, coffee melanoidins are considered to
be high-molecular-weight (HMW) brownish compounds.
However, their chemical structure is not fully understood
because of both the large diversity of structures that couple
to build the different melanoidin fractions and the difficulty
in isolating and characterizing such fractions. A first step
in the elucidation of coffee melanoidin structure relies on
estimating their molecular weight. This has been done by
different separation techniques that isolate melanoidins by
using their heavy molecular weight. Among these, ultra-
filtration, diafiltration, and dialysis are the most suitable
for isolating melanoidins, using membranes with different
molecular weight cut offs, ranging from 3 to 100 kDa.8–10
Although no general consensus regarding the real molecu-
lar weight of coffee melanoidins exists, it is more accepted
that they have a mass ranging from 3 to 22 kDa.8 In addi-
tion, these melanoidins could be linked in a noncovalent
manner to build a small fraction with a molecular mass
higher than 100 kDa. Other authors also state that in addi-
tion to the HMW fraction, a low-molecular-weight (LMW)
fraction is also found.9 According to this article, 69% of
coffee melanoidins has a molecular weight higher than
12 kDa and 29% are LMW melanoidins. In addition, the
increasing formation of HMW melanoidins as a response
to a severe roasting process has been demonstrated.11
A second purification step relies on the different physi-
cochemical properties of the HMW fraction previously
isolated. For example, depending on their hydrophobic-
ity, other LMW fractions were obtained using solid phase
extraction.9 A different approach uses the metal chelating
ability of coffee melanoidins, separating the HMW com-
plex into fractions with different copper-chelating strengths
using copper affinity chromatography.12 Related to this,
because of the anionic behavior of coffee melanoidins, they
can also be isolated by anion exchange chromatography.7
In the case of melanoidin solubility, ethanol partitioning
can be used to obtain fractions with different polysaccha-
ride composition.13 At ethanol concentrations of 40–50%,
galactomannans (GMNs) precipitate, whereas arabinoga-
lactans (AGLs) precipitate at 75–80%; the remaining solu-
ble compounds have the lowest carbohydrate content.

Coffee in Health and Disease Prevention 183

http://dx.doi.org/10.1016/B978-0-12-409517-5.00020-6 © 2015 Elsevier Inc. All rights reserved.
184 20.  COFFEE MELANOIDINS

Regarding the physicochemical properties of coffee
melanoidins, their solubility depends on the size of the
polymer and its chemical nature. In coffee, melanoi-
dins are quite hydrophilic because of their intermedi-
ate molecular weight (3–33 kDa; see Table 20.1 for more
information). However, in other foods with high protein
content, carbohydrates react through the MR to build a
cross-linked HMW network that is insoluble.3 In addi-
tion, the LMW fraction of coffee melanoidins has a
hydrophobic nature.9 Another interesting property of
melanoidins is their polyanionic behavior.14 Coffee mel-
anoidins are negatively charged, but the distribution of
such charges is heterogeneous15 in that the HMW frac-
tion is more charged than the LMW one.9 According to
this, almost half of the anionic fractions have copper-
chelating properties.12 In the case of coffee melanoidins,
the source of these negative charges seems to rely on the
presence of chlorogenic acids,16 although melanoidins
from other foods without chlorogenic acids also show
such anionic behavior.14 Coffee melanoidins consists of
a mixture of carbohydrates, amino acids, and phenolic
compounds that interact during coffee roasting. In the
case of polysaccharides, the predominant species in green
coffee beans are GMNs and AGLs, which comprise up
to 50% of the bean’s dry weight.2 AGLs are covalently
linked to proteins in both green and roasted coffee17 in
the form of arabinogalactan-proteins (AGPs), and differ-
ent melanoidin fractions also contain 50% of brownish
AGPs.15 Both GMNs and AGLs undergo different chemi-
cal (caramelization, MR, oxidation, etc.) and structural
modifications (e.g., AGL debranching and GMN polym-
erization) because of the thermal treatment that occurs
during the roasting process.13,18 These polysaccharides
are covalently linked to the different melanoidin popu-
lations,7,8,12 although the exact types of linkages is yet
unknown (for a more detailed explanation see Ref. 2).
Proteins are especially rich in the melanoidin fraction
with a higher solubility at an ethanol concentration of
80%.7,8 The proteins linked to melanoidins have a high
concentration of aspartic and glutamic acids, as well as
alanine and glycine, whereas the amounts of histidine,
lysine, methionine, tyrosine, and arginine are lower than
those in green coffee proteins.16 This could be explained
by taking into account the protein degradation that
occurs as a consequence of the roasting process and the
extent of the MR. Then, some MR products derived from
these amino acids, such a Nε-(fructosyl)lysine or Nε-
(carboxymethyl)lysine, also are included in the melanoi-
din structure.12
The amount of phenolic compounds in green coffee is
high (up to 10%), so it is clear that some of them can
be incorporated within the melanoidin structure. The
main part of phenolic compounds in coffee is chlo-
rogenic acids, which includes different acids such as
caffeic, coumaric, or ferulic acid.19 Less than 1% of
the chlorogenic acids found in green coffee are incor-
porated into melanoidins, although during roasting
the phenolics linked to melanoidins increase to up to
29% of the overall phenolic compounds found in the
coffee brew.20 It is noteworthy that these chlorogenic
acids incorporated during the first stages of the pro-
cess are later degraded. These acids have been demon-
strated to be noncovalently linked to the melanoidin
skeleton in soluble coffee21 through hydrogen, hydro-
phobic, or ionic bonding and can be released during
gastrointestinal digestion,22 which allows their possible
absorption at the intestinal level. In addition, the pres-
ence of phenolic compounds covalently linked to the
melanoidin core has been speculated using the Folin-
Ciocalteu reagent15 and the detection of phenols by
thermal degradation of coffee melanoidins.23,24 How-
ever, the presence of chlorogenic acids in the melanoi-
din structure was confirmed by alkaline fusion, which
released different chemical species derived from caffeic
and ferulic acids.12 The linking site of such phenols is
still being studied, although it has been proposed that

TABLE 20.1  Physicochemical Properties of Melanoidins

Foods Containing Melanoidins Color Intensity Appearance Water Solubility Hygroscopicity

Coffee ++++ Large, iridescent, flat flakes ++++ ++

Chocolate +++ Hardened agglomerate ++ ++++

Balsamic vinegar +++ Short, flat flakes +++ ++

Sweet wine ++++ Short, flat flakes ++ +++

Dark beer +++ Powder +++ ++

Pilsner-style beer ++ Powder +++ ++

Bread crust + Spongy mass + ++

Breakfast cereals + Spongy mass + ++

Biscuits + Spongy mass + +

I.   INTRODUCTORY AND GENERAL TEXT

 20.3  Mechanistic Formation and Structure of Coffee Melanoidins
chlorogenic acids could be linked to the melanoidin
skeleton through proteins25 or through the arabinose
residues of GMNs and AGLs.16 The most recent results
in the field26 indicate that phenolic compounds interact
with coffee storage proteins (legumin-like seed stor-
age proteins of the 11S class) thanks to polyphenol oxi-
dase activity. These chlorogenic acids react covalently
with lysine at the α chain and undergo oxidation by
polyphenol oxidase, giving rise to colored compounds
ranging from light yellow to dark green. The develop-
ment of dark pigments because of the presence of poly-
phenols also has been seen in melanoidins from other
foods such as sweet wine or grape syrup.4
Other LMW compounds have been found in cof-
fee melanoidins and melanoidins from different foods
and model systems. Some of the LMW compounds
attached to melanoidins are odor-active thiols, such
as 2-furfurylthiol, 3-methyl-2-butenthiol, 3-mercapto-
3-methylbutyl formate, 2-methyl-3-furanthiol, and
methanethiol, that influence the flavor of the coffee
brew.27 Others are furans accompanied by pirazines,
pyrroles, pyridines, and carbonyl compounds.4 Most of
these compounds are chromogenic, with different colors
such as blue (pyrrolopyrrole dimer), red (pyrrolopyrrole
azepine), and yellow (acetyl pyrimidine and acetyl
azepinone). These colorants have an extraordinary
polymerizing activity, which suggests that they are
reaction intermediates for melanoidin formation.28 The
involvement of such LMW colored compounds in the
formation of melanoidins is discussed in the next section.
All the fractions mentioned above—LMW and HMW
proteins, polysaccharides, and phenolic compounds—give
TABLE 20.2  Biological Activities of Coffee Melanoidins
Biological Activity Active Structure
Prebiotic AGLs and GMNs
Anticariogenic Polymeric backbone
Food preservation Polymeric backbone
Fiber-like antioxidant activity AGLs and GMNs
Antioxidant activity in coffee HMW and LMW fractions
brew
Plasma antioxidant capacity N-methylpyridinium
Toxic compound binding HMW fraction
Aroma compound binding AGLs, GMNs, or chlorogenic acids
Cation binding HMW fraction
Chemopreventive HMW fraction
Anticarcinogenic HMW fraction
Antihypertensive HMW and LMW fractions
AGL, arabinogalactan; GMN, galactomannan; HMW, high-molecular weight;
LMW, low-molecular weight.
I.   INTRODUCTORY AND GENERAL TEXT
185
rise to many different biological effects such as antioxi-
dant,29 antimicrobial,30 or antihypertensive activities31
that depend strongly on fraction’s chemical composition
(Table 20.2), as reviewed elsewhere.2–4
20.3  MECHANISTIC FORMATION AND
STRUCTURE OF COFFEE MELANOIDINS
The formation of melanoidins is quite complicated
in real foods and has been studied using model sys-
tems. In these systems, reaction time and temperature
dictate that the molecular weight of coffee melanoi-
dins, since pH influences the development of chromo-
phores.4 In general, there are three theories about how
HMW melanoidins are formed. The first theory states
that the HMW complexes are formed from LMW inter-
mediates coming from the MR (pyrroles, furans, etc.)
that polymerize during the later stages of the reaction.28
As stated in the previous section, the LMW chromo-
phores are reaction intermediates, which means that
their polymerizing activity depends strongly on the
heating time. A second theory32 proposes that the HMW
melanoidin complexes are formed by the cross-linking
activity of chromophoric LMW compounds and the
reactive side chains of several amino acids (lysine,
arginine, cysteine). Different chromophores have been
isolated through the reaction of α-dicarbonyl com-
pounds, furans, or their nitrogen-containing deriva-
tives with arginine, lysine, or casein.4 The nitrogen
atoms of such colored compounds can then be included
within proteins via the furan-derived structures pro-
duced upon the thermal degradation of carbohydrates
or the amino and guanidine groups of amino acids.
The third theory postulates that the melanoidin core is
made of sugar derivatives (degradation products such
as α-dicarbonyl compounds, oligosaccharides, and/
or polysaccharides) branched with amino acids.33 The
main structure of such saccharides remains uncharged
in low-moisture media, reacting with amino acids at
their reducing end. In addition, a small part of the
glycosidic bonds is broken, allowing transglycosyl-
ation reactions during the first stages of the MR that
increase the branching state of the melanoidin skel-
eton. This is the situation in real foods such as coffee
or cocoa, where the MR takes place at low moisture
content and high temperature (>150 °C) in short time
periods (ranging from minutes to a few hours). On the
other hand, α-dicarbonyl compounds react with pro-
teins via aldolic condensation in high-moisture media
to increase the complexity of the melanoidin back-
bone.4 This is how melanoidins form in high-moisture
foods such as beer or sweet wine, where the MR
takes place for longer periods (fermentation or stor-
age for days or months) at low temperatures (<50 °C).
186 20.  COFFEE MELANOIDINS

Then, because of the extensive polymerization, food by the authors is that the formation of the melanoidin
melanoidins (especially those coming from liquid itself and the brown compounds during coffee roast-
foods) are richer in HMW compounds than in LMW ing have distinct origins. On one hand, the presence
ones. of soluble sugar in coffee is one of the main factors
The different chemical composition of coffee mela- for the development of LMW brownish compounds
noidins suggests that they can be formed through dif- linked to proteins and polysaccharides. On the other
ferent pathways. In this sense, previously published
articles seem to indicate that the above three theo-
ries may occur during coffee roasting (Figure 20.1).
The presence of volatile compounds, such as furans
and carbonyl compounds, in thermally degraded
coffee melanoidins,23 as well as the formation of the
polymerizing CROSSPY radical during coffee roast-
ŽīĞĞ
ing, reinforce the plausibility of the first theory. In ŵĞůĂŶŽŝĚŝŶƐ
the case of the second and third theories, other stud-
ies have pointed out the presence of different popula-
tions of modified and nonmodified proteins12,16,26 and
polysaccharides7,8,12,15 on the melanoidin backbone
(Figures 20.2 and 20.3). Nunes and coworkers34 FIGURE 20.1  The mechanisms of melanoidin formation in
recently studied these several formation pathways coffee. Adapted with permission from Nunes and Coimbra.25 Copyright©
through a novel “in bean” model. The first idea stated 2010, Springer.

FIGURE 20.2  Relevance of proteins and phenolic compounds in the formation of coffee melanoidins. Adapted with permission from Nunes
and Coimbra.25 Copyright© 2010, Springer.

FIGURE 20.3  Relevance of arabinogalactan proteins and galactomannans in the formation of coffee melanoidins. Adapted with permission
from Nunes and Coimbra.25 Copyright© 2010, Springer.

I.   INTRODUCTORY AND GENERAL TEXT

 20.5 Summary Points 187

ŵĂƚĞƌŝĂů

ŽīĞĞ
ŵĞůĂŶŽŝĚŝŶƐ

FIGURE 20.4  Main chemical species of coffee melanoidins. Adapted with permission from Nunes and Coimbra.25 Copyright© 2010, Springer.

hand, sugars such as sucrose do not increase the final
amount of HMW melanoidins. In the same way, AGPs
were the other source (in addition to chlorogenic acid)
of brown compounds, in accordance with other arti-
cles.15 This could be related to the reaction of the arab-
inose side chains with reducing sugars upon roasting.
Finally, Nunes and coworkers also found that the cell
wall material is responsible for generating a large
amount of melanoidins. Then, the roasting-induced
pyrolysis of cell wall polysaccharides such as GMNs
accounts for 47% of the melanoidin formed. All the
chemical species responsible for the formation of cof-
fee melanoidins are depicted in Figure 20.4.
20.4  FINAL REMARKS
In the past decade the elucidation of melanoidins’
structure, composition, and formation pathways cen-
tered the interest of many research groups worldwide.
The European COST Action 919 “Melanoidins in food
and health” was the driving force behind melanoidin
research in Europe. In the past 10 years research activity
moved toward the biological activity of these polymers.
Therefore, the knowledge about the amount and forma-
tion of melanoidins in different foodstuffs will allow the
formulation of new foods with improved sensorial or
technical and biological activities.
20.5  SUMMARY POINTS
• C offee melanoidins are brown nitrogen-containing
polymers produced during coffee roasting.
• The amount of coffee melanoidins accounts for up
to 25% of the roasted coffee and increases with the
severity of roasting.
• Coffee melanoidins are composed of different
chemical species such as GMNs, AGLs, proteins,
chlorogenic acids, and MR products.
• The formation of melanoidins is explained by three
different theories involving the polymerization of
LMW MR products, cross-linking activity of LMW
compounds over proteins, and/or the reaction of
polysaccharides with proteins.
• Coffee melanoidins are generated by these three
mechanisms, including the additional reaction of
phenolic compounds.

I.   INTRODUCTORY AND GENERAL TEXT

188 20.  COFFEE MELANOIDINS
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