Caballero B. (ed.) Encyclopaedia of Food Science, Food Technology and Nutrition. Ten-Volume Set

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room. Paris: UNESCO.

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Breast-feeding See Infants: Nutritional Requirements; Breast- and Bottle-feeding; Weaning; Feeding

Problems; Lactation: Human Milk: Composition and Nutritional Value; Physiology

BROWNING

Contents

Nonenzymatic

Toxicology of Nonenzymatic Browning

Enzymatic – Biochemical Aspects

Enzymatic – Technical Aspects and Assays

Nonenzymatic

Jennifer M Ames, University of Reading, Reading, UK

Introduction

0001 The term ‘nonenzymatic browning’ refers to the

chemical reactions that result in the formation of

brown color when food is heated. In contrast to

enzymatic browning, no enzymes are involved in

nonenzymatic browning reactions. The most import-

ant nonenzymatic browning reaction is the Maillard

reaction, which encompasses the cascade of reactions

that occur when reducing sugars are heated with

compounds possessing a free amino group (e.g.,

amino acids, amines, and proteins) and which result

in thousands of reaction intermediates and products.

The Maillard reaction is named after the French

scientist, Louis Camille Maillard, who first investi-

gated reducing sugar–amino acid interactions in

1912. Other nonenzymatic browning reactions in-

clude Maillard-type reactions between amino com-

pounds and other compounds possessing a free

carbonyl group, e.g., ascorbic acid and lipid oxida-

tion products. A further type of nonenzymatic

browning is caramelization, which occurs when

sugars discolor in the absence of amino compounds.

In all types of nonenzymatic browning mentioned,

the chemistry involved is very similar. In this article

(unless stated otherwise), the terms ‘nonenzymatic

browning’ and the ‘Maillard reaction’ are used inter-

changeably and refer to reactions between reducing

sugars and amino compounds.

0002As well as giving brown colors, the Maillard reac-

tion results in various other outcomes in foods. These

include development of flavor, increase in antioxida-

tive capacity, loss of nutritional and functional prop-

erties, and formation of compounds with potential

adverse effects on human health. Since the Maillard

BROWNING/Nonenzymatic 665

reaction has so many important consequences for

food quality, it is without doubt one of the most

significant reactions that occur in food. Various

factors affect the course of the Maillard reaction.

These include the nature of the reactants, tempera-

ture, and time of heating, the pH and water activity

) of the food and the presence of reaction inhibi-

tors, e.g., sulfite. Due to the importance of the

reaction, its control is important for food quality

and kinetic approaches to modeling and control are

showing considerable promise.

Chemistry

0003 A summary of the chemical pathways comprising the

Maillard reaction is given in Figure 1. The early stage

of the reaction is well understood. It involves the

condensation of the reducing sugar with the amino

compound to give an N-substituted glycosylamine

which rearranges to the 1-amino-1-deoxy-2-ketose,

also known as the Amadori rearrangement product

(ARP). The relative importance of the subsequent

pathways largely depends on the pH of the food. At

a pH below 5, the route involving 3-deoxyosone as

the intermediate prevails but, if the pH is above 7,

the pathways involving the 1- and 4-deoxyosones

and the 1-amino-1,4-dideoxyosone are more import-

ant. Since most foods are in the pH range 4–6,

the 3-deoxyosone route usually predominates. The

3-deoxyosone reacts with amino acids to give mela-

noidins (the final products of the Maillard reaction

which are macromolecular and colored), Strecker al-

dehydes via the Strecker degradation (Figure 2) and

pyrroles and furfurals and their derivatives. Enami-

nols, which are products of the Strecker degradation,

subsequently condense to give pyrazines (Figure 2).

The other deoxyosones are much less stable than the

3-deoxyosone. They are converted into reductones

which may react with other intermediates to give

melanoidins. In addition, these deoxyosones (as

well as other Maillard intermediates, e.g., the N-

substituted glycosylamine) readily fragment via retro-

aldolization. The low-molecular-weight products

that result are highly reactive and may participate

Fragmentation

products

Low-molecular-weight colored compounds

and melanoidins

Reductones

Cyclic flavor

compounds

1-Deoxyosone,

4-deoxyosone,

1-amino-1,4-dideoxyosone

Strecker

aldehydes

3-Deoxyosone

Furfurals

Pyrroles

N-substituted

glycosylamine

Reducing sugars +

amino compound

1,2-Eneaminol

Amadori

rearrangement product

(ARP)

2,3-Enediol

Heterocyclic

amines (HAs)

fig0001 Figure 1 Outline of the chemical pathways of the Maillard reaction.

666 BROWNING/Nonenzymatic

in the Strecker degradation, condensation reactions

resulting in cyclic products, and reactions leading to

melanoidins.

0004 At temperatures above 130



C or at pH values of

below 3 or above 8, sugars degrade in the absence

of amino acids to give key intermediates that also

form in the Maillard reaction (Figure 1), as shown

in Figure 3.

Flavor

0005 Hundreds of flavor compounds formed by the Mail-

lard reaction have been identified. The compounds

concerned are largely those that contribute to aroma

(i.e., they are volatile and are thus perceived by the

nose), although a few components that possess

specific tastes (and are therefore perceived by the

tongue) have also been reported. Roasted coffee,

cooked meat, bread, cookies, breakfast cereals,

roasted nuts, and baked potatoes are among the

foods that owe their particular flavor to the Maillard

reaction. Also, the Maillard reaction is employed by

the flavor industry for the production of process or

reaction flavors. In addition to the development of

desirable flavors, the Maillard reaction is responsible

for off-flavors in certain foods, particularly in com-

modities in which the original fresh flavor of the

product is desired. Thus, it can lead to cooked or

stale flavors in processed orange juice or dried milk

powder. (See Flavor (Flavour) Compounds: Struc-

tures and Characteristics.)

0006The volatile Maillard flavor compounds may be

divided into three groups: those formed from the

sugar, those formed from the amino acid, and those

requiring both the sugar and the amino acid for their

formation (Table 1). Strictly speaking, it is only the

compounds in the latter group that are formed by the

Maillard reaction, i.e., reducing sugar–amino com-

pound interactions. Members of a further category

of aroma compounds are formed by reactions involv-

ing intermediates of both the Maillard reaction and

lipid degradation. These components often possess

fried odors and contribute fried aromas to foods

such as potato chips. Much less is known about the

products of the Maillard reaction that may contribute

to taste. However, it has been established that some

pyrrole derivatives contribute to the bitter taste of

roasted coffee and malt. The structures of some spe-

cific compounds, together with their odor (or taste)

threshold values and contributions to food flavor, are

given in Table 2.

Color

0007The compounds formed early on in the Maillard re-

action are colorless and absorb in the ultraviolet (UV)

region of the electromagnetic spectrum. As the reac-

tion progresses, absorption in the UV region intensi-

fies and begins to tail into the visible region, resulting

in the appearance of yellow, orange, and brown

R C COOH + R' CO CO R"

+ R CHO + R' CO

Amino acid

Dicarbonyl, e.g.,

3-deoxyosone

Strecker

aldehyde

Enaminol

A pyrazine

2 R' CO C R"

−2H

fig0002 Figure 2 Strecker degradation and pyrazine formation.

Aldose

reducing sugars

1,2-Enediol

Ketose

reducing sugar

2,3-Enediol1-Deoxyosone

3-Deoxyosone

4-Deoxyosone

fig0003 Figure 3 Degradation of reducing sugars at temperatures

above 130



C, or at pH values below 3 or above 8, and in the

absence of amino compounds.

tbl0001Table 1 Volatile compounds derived from the sugar, the amino

acid, and sugar/amino acid interactions

Compounds derived

from the sugar

Compounds derived

from sugar^amino

acidinteractions

Compounds derived

from the amino acid

Furans Pyrroles Sulfur compounds

Pyrones Pyridines Aldehydes

Cyclopentenes Pyrazines Thiazoles

Carbonyls Oxazoles Cyclic polysulfur

compoundsAcids Thiophenes

BROWNING/Nonenzymatic 667

tbl0002 Table 2 Properties of selected flavor compounds

Compound Odor threshold value (mgkg

1

)Odor

Butanedione (diacetyl) CH

COCOCH

7 Buttery

3-(Methylthio)propanal (methional) CH

SCH

CHO 200 Cooked potatoes

2,5-Dimethyl-4-hydroxy-3(2H)furanone (furaneol) 0.04 Caramel, burnt pineapple, strawberries

Methylpyrazine 6 10

Nutty

2-Acetyl-1-pyrroline 0.1 Freshly baked wheat bread crust

2-Acetyl-1,4,5,6-tetrahydropyridine 1.6 Crackers

Trimethyloxazole H

C 5 Green, nutty, sweet

2-Acetylthiophene 0.1 Onion-link, mustard-like

bis(2-Methyl-3-furyl) disulfide 2  10

5

Cooked meat

2-Actylthiazole 10 Nutty, popcorn

3-Methyl-5-pentyl-1,2,4-trithiolane Fried chicken

5–10 10

Bitter

Taste threshold.

COCH

668 BROWNING/Nonenzymatic

colors. The Maillard reaction is responsible for the

desirable yellow or brown colors in foods, including

bread crust, potato chips, popcorn, cookies, and

other flour confectionery, but it can also lead to

discoloration, e.g., in heated fruit juices and milk

powder.

0008 The colored Maillard reaction products may be

divided into two groups, i.e., the low-molecular-

weight compounds that possess up to about five

linked rings, and the macromolecular melanoidins.

Most work on colored Maillard reaction products

has concerned the low-molecular-weight compounds

isolated from model food systems. A selection of

compounds isolated from aqueous sugar–amino acid

systems, heated with or without additional furfural,

is shown in Figure 4. Progress in the field of low-

molecular-weight colored Maillard products has

advanced rapidly over the last 5 years. Nevertheless,

much remains to be done.

0009 No homogeneous melanoidin has been isolated

from a food or model food (sugar–amino acid) system

to date and no structures for melanoidins isolated

from food are available. Progress concerning the

structures of high-molecular-weight melanoidin-like

structures has been made by investigating the

polymers produced on polymerization of first,

2-hydroxymethyl-N-methylpyrrole in trichloro-

methane and, second, N-methylpyrrole with 2-for-

myl-N-methylpyrrole (or furfural) in methanol.

Figure 5 gives an example of the structures identified

C nuclear magnetic resonance spectroscopy,

fast atom bombardment–mass spectrometry and

matrix-assisted desorption ionization–time of flight–

mass spectrometry. Although the formation of such

regular homopolymers in food systems is unlikely, it

is reasonable to expect that the type of structure

depicted in Figure 5 represents a domain of a food

melanoidin.

0010It has been demonstrated that heating food pro-

teins (casein) in the presence of glucose results in

protein cross-linking and a parallel increase in color

III IV

CHO

H COOH

fig0004 Figure 4 Selected low-molecular-weight colored compounds.

I, from xylose and glycine; II, from xylose and glycine or xylose

and lysine; III and IV, from xylose, alanine and furfural. (I:2-

Acetyl-6-(hydroxymethyl)-5,6-dihydro-4H-pyridinone; II: 2-furfury-

lidene-4-hydroxy-5-methyl-3(2H)-furanone; III: 4-[1-formyl-2-(2-

furyl)ethenyl) ]-5-(2-furyl)-2-[(2-furyl)methylidene]-2,3-dihydro-a-

methyl-3-oxo-1H-pyrrole-1-acetic acid; IV: 4-(2-furyl)-7-[(2-furyl)-

methylidene]-2-hydroxy-2H,7H,8aH-pyrano[2,3b]pyran-3-one.)

CHO

VVI

+V, +VI

Up to about 26 units

+ V

+ VI

fig0005Figure 5 Structure of the polymer prepared from N-methylpyr-

role and 2-formyl-N- methylpyrrole.

BROWNING/Nonenzymatic 669

intensity of the reaction products. This color develop-

ment is due to the incorporation of colored Maillard

reaction products into the cross-linked protein.

Therefore it is likely that, in foods, a portion of

the brown color formed on heating is due to low-

molecular-weight colored Maillard products binding

to colorless protein backbones giving protein cross-

linking and browning.

Antioxidant Activity

0011 The Maillard reaction may influence the oxidative

stability of foods because some Maillard reaction

products are able to retard lipid oxidation. An in-

crease in stability may be achieved by modifying the

heat process applied, e.g., to milk, adding Maillard

reaction precursors, e.g., to cookie dough, prior to

heat treatment, or addition of Maillard reaction

products to susceptible foods, e.g., margarine.

0012 An increase in pH and the amino compound–

sugar molar ratio enhances the formation of nondia-

lyzable melanoidins and the inclusion of nitrogen in

the Maillard reaction products. There is a positive

correlation between all these factors and antioxi-

dant activity. Nondialyzable melanoidins can scav-

enge hydroxyl radicals and other active oxygen

species. Antioxidant activity increases with formation

of melanoidins up to a maximum and then decreases

with further heating and browning. Reducing activity

and metal chelating ability are factors that may

account for antioxidant activity. A strong candidate

for such activity is the reductone group. However,

enediol-like reductones and amino reductones

(examples of the latter being those formed by the

Amadori rearrangement) may act as antioxidants or

prooxidants, depending on the reaction conditions,

and the structures responsible for antioxidant activity

remain uncertain. (See Antioxidants: Natural Anti-

oxidants; Role of Antioxidant Nutrients in Defense

Systems.)

Nutritional Effects

0013 The Maillard reaction can result in serious loss of

availability of essential amino acids, both in the free

form and in peptide chains. Loss of the available

amino acid occurs as soon as the ARP is formed. In

most foods, lysine destruction is most significant, due

to its e-amino group. Ascorbic acid and also vitamins

possessing a free amino group, e.g., thiamine, can

participate in Maillard-type reactions leading to loss

in activity of the vitamin concerned. Various Maillard

reaction products may complex nutritionally im-

portant metals, e.g., copper, zinc, iron, thus making

them unavailable. ARPs or more advanced reaction

products fed orally to humans increase zinc excretion

via urine. The effect is more marked in patients fed

intravenously.

0014The posttranslational modification of proteins and

the development of protein cross-links as a conse-

quence of the Maillard reaction reduce protein digest-

ibility (as well as functionality). Pyrraline (identified

in heated milks) and pentosidine (identified in roasted

coffee) are examples of such modifications and their

structures are shown in Figure 6. Reduction in protein

digestibility may be due to decreased access by digest-

ive enzymes to the modified protein or by the direct

inhibition of the enzymes. The main effect is to block

normal tryptic hydrolysis on the carboxyl side of

lysine.

Mutagen Formation

0015More than 20 heterocyclic amines (HAs) that are

products of the Maillard reaction have been identified

at the p.p.b. level in cooked beef, pork, lamb, chicken,

and fish muscle. Amino acids, sugars, and bio-

chemicals present in muscle tissue, e.g., creatinine

and creatine, are all required for their formation.

The structures of a selection of HAs are shown in

Figure 7. The amounts produced increase with tem-

perature and time of cooking; temperatures encoun-

tered during, for example, grilling and frying favor

their formation. Most HAs cause mutations in bac-

teria, mammalian cells and cancer in animals. They

are the most potent mutagens examined by the Ames

test and are able to produce frameshift mutations in

arginine

lysine

Pentosidine

HOH

(CH

)

CHO

NH CH C

Pyrraline

fig0006Figure 6 Structures of pyrraline and pentosidine. The peptide

backbone is represented by a solid line.

670 BROWNING/Nonenzymatic

Salmonella and base pair substitutions in hamster

ovary cells. The risk to humans of consuming foods

containing measurable amounts of HAs is difficult to

estimate and requires epidemiological studies. Esti-

mates based on animal cancer data and surveys of

human food intake suggest that the average exposure

to cooked food containing HAs gives a 10

4

risk of

lifetime cancer. This average value is subject to sub-

stantial variation due to differences in diet, cooking

methods, and genetic factors.

0016 Although HAs are the mutagens formed during

the Maillard reaction that have received most

attention, other Maillard products are also reported

to be mutagenic, including methylglyoxal and

5-hydroxymethylfurfural (HMF). (See Mutagens.)

Factors Affecting the Maillard Reaction

0017 The extent and course of the Maillard reaction are

affected by the profile of components in the food

capable of acting as reaction precursors, temperature

and time of heating, pH, and a

. The reaction is also

affected by metals, oxygen, and the presence of in-

hibitory agents, e.g., sulfite. When attempting to con-

trol the Maillard reaction, the aim may be to stop it or

to steer it towards a particular outcome in terms of

flavor or antioxidant activity. In any case, the reac-

tion is difficult to stop once it has started.

0018 The Maillard reaction may be minimized by re-

moving one of the required reaction partners, i.e.,

the reducing sugars for protein-rich foods or the

amino compounds for carbohydrate-rich foods. In

jam-making, limiting the cooking time after addition

of sucrose reduces its inversion to glucose and fruc-

tose in the acidic medium. An alternative to removing

the reducing sugars or amino compounds is to process

separately the carbohydrate and protein components

of a food, e.g., cream-style soups.

0019Pentose sugars, e.g., ribose, react faster than hexose

sugars, e.g., glucose and fructose, which in turn react

faster than reducing sugars that are disaccharides,

e.g., maltose and lactose. Amino acids also react at

different rates. Of the free amino acids, when color

formation is measured, lysine and glycine are among

the fastest while cysteine is the slowest. In peptide

chains, the amino acid residues with reactive

side chains, e.g., lysine and arginine, react the fastest.

The amino acid has a large effect on the flavor

formed. Cysteine gives meaty notes, methionine

results in a potato aroma, and cracker and bready

odors are produced by proline.

0020The Maillard reaction increases with temperature.

values range from 2 to 8 according to the system

and parameter being measured. Therefore, if the aim

is to stop the reaction, the temperature should be

reduced to as low a level as possible. The profile of

reaction products formed varies according to the tem-

perature and time of heating and it is not possible to

produce the profile obtained at a high temperature by

heating at a lower temperature for a longer time. This

MeIQ

MeIQx

DiMeIQx PhIP

fig0007 Figure 7 Structures of selected heterocyclic amines. MeIQ, 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline; MeIQx, 2-amino-3,

8-dimethylimidazo[4,5-f]quinoxaline; DiMeIQx, 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline; PhIP, 2-amino-1-methyl-6-phenyli-

midazo[4,5-b]pyridine.

BROWNING/Nonenzymatic 671

can be illustrated by comparing the aroma of meat

resulting from boiling at 100



C and from roasting at

200



C. The two aromas are easily distinguished be-

cause different reaction pathways, and thus different

aroma compounds, are favored by the different

cooking methods.

0021 The pH of the system plays a key role in determin-

ing the relative importance of the reaction pathways

followed (see section on Chemistry, above). At pH

values below about 5, the 3-deoxyosone route is

favored while routes involving the other deoxyosones

are favored above pH 7. In fact, there is no sharp cut-

off point. All the pathways operate at all pH values of

relevance to food, the ratio of products being influ-

enced by the pH. Thus, pH is a parameter that can

be manipulated to achieve the required profile of

reaction products.

0022 Since water is a product of the Maillard reaction,

the reaction occurs less readily in high-a

foods.

In low-a

foods, the reactants are less mobile and

this also impedes the reaction. In practice, the Mail-

lard reaction is favored by intermediate a

values

(0.5–0.8). When the aim is to prevent the Maillard

reaction, a

is of great concern during the drying or

concentration of food. In such cases, the food must be

taken through the intermediate a

stage as quickly as

possible, especially if heat is used. The formation of

individual Maillard reaction products is favored by

different a

values and therefore a

represents

another means of optimizing the profile of reaction

products formed.

0023 Since the Maillard reaction is so difficult to stop

once it has begun, the food industry may resort to the

use of sulfite (normally a solution of sodium bisulfite

or sodium metabisulfite) to control it. The sulfite

combines with reducing sugars and Maillard inter-

mediates possessing carbonyl and dicarbonyl groups

to inhibit the reaction.

Kinetics and Modeling

0024 The Maillard reaction is much more complex than

most reactions that have been modeled by physical

chemists. However, since the reaction is of such inter-

est to the food industry, attempts have been made to

describe it using kinetics and mathematical equations,

in order to make valid predictions concerning color,

flavor, and antioxidant activity for various food com-

positions processed and stored under different condi-

tions. As a result, it is possible to predict accurately

the color development (absorbance at a single wave-

length of a model system) at a particular time. Rate

constants have also been published for a limited

selection of Maillard aroma compounds. However,

since the Maillard reaction is qualitatively and quan-

titatively affected by many factors, the direct applica-

tion of models may be of very limited use. There is a

great need for further application of kinetics prin-

ciples to the Maillard reaction so that various aspects

of the reaction may be predicted and, ultimately,

controlled in foods.

See also: Antioxidants: Role of Antioxidant Nutrients in

Defense Systems; Browning: Toxicology of

Nonenzymatic Browning; Casein and Caseinates: Uses

in the Food Industry; Flavor (Flavour) Compounds:

Structures and Characteristics; pH – Principles and

Measurement

Further Reading

Ames JM (1992) The Maillard reaction. In: Hudson BJF

(ed.) Progress in Food Proteins – Biochemistry,

pp. 99–153. London: Elsevier Applied Science.

Ames JM and Hofmann T (eds) (2000) Chemistry and

Physiology of Food Colorants. American Chemical

Society Symposium Series 775. Washington, DC: ACS.

Eriksson C (ed.) (1981) Maillard Reactions in Food.

Oxford: Pergamon Press.

Finot PA, Aeschbacher HU, Hurrell RF and Liardon R (eds)

(1990) The Maillard Reaction in Food Processing,

Human Nutrition and Physiology. Basel: Birkha

user.

Friedman M (1996) Food browning and its prevention: an

overview. Journal of Agricultural and Food Chemistry

44: 631–653.

Fujimaki M, Kato H and Namiki M (eds) (1986) Amino-

Carbonyl Reactions in Food and Biological Systems.

Amsterdam: Elsevier.

Hodge JE (1953) Dehydrated foods: chemistry of browning

reactions in model systems. Journal of Agricultural and

Food Chemistry 1: 928–943.

Ikan R (ed.) (1996) The Maillard Reaction. Consequences

for the Chemical and Life Sciences. Chichester: John

Wiley.

Ledl F and Schleicher E (1990) New aspects of the Maillard

reaction in foods and in the human body. Angewandte

Chemie International Edition in English 29: 565–706.

Leong LP and Wedzicha BL (2000) A critical appraisal of

the kinetic model for the Maillard browning of glucose

and glycine. Food Chemistry 68: 21–28.

Namiki M (1988) Chemistry of Maillard reactions: recent

studies on the browning reaction mechanism and the

development of antioxidants and mutagens. Advances

in Food Research 32: 115–184.

O’Brien J and Morrisey PA (1989) Nutritional and toxico-

logical aspects of the Maillard browning reaction in

foods. Critical Reviews in Food Science and Nutrition

28: 211–248.

O’Brien J Nursten HE, Crabbe MJC and Ames JM (eds)

(1998) Maillard Reaction in Foods and Medicine. Cam-

bridge: Royal Society of Chemistry.

Waller GR and Feather MS (eds) (1983) The Maillard Re-

action in Foods and Nutrition. ACS Symposium Series

215. Washington, DC: ACS.

672 BROWNING/Nonenzymatic

BROWNING

Toxicology of Nonenzymatic

Browning

P A Finot, Nestec, Nestle

Research Centre, Lausanne,

Switzerland

This article is reproduced from Encyclopaedia of Food Science,

Introduction

0001 Reactions between reducing sugars and free amino

groups in foods, without any catalytic involvement of

enzymes, lead to nonenzymatic browning (Maillard

reaction) causing a reduction in nutritive value and

certain physiological and/or toxicological effects.

The browning reaction develops during both home

cooking and industrial processing of foods. Whilst

contributing to an improvement of the organoleptic

properties of foods, through aroma development,

browning is often an undesirable side-effect of obliga-

tory heat treatments applied for microbiological

(sterilization and drying) or nutritional (cooking)

reasons and for convenience (storage). Since the

Maillard reaction occurs so frequently and Maillard

reaction products are present in practically all meals,

an understanding of its biological implications is of

importance.

0002 Because of the multiplicity of the food systems, the

complexity of the chemical reactions, and the large

variety of heat treatments involved, any generaliza-

tion on the biological outcome of nonenzymatic

browning of foods is not easy. Nevertheless, a number

of studies of model amino acid/protein and sugar

reaction systems permits a classification of the bio-

logical outcome as: (1) nutritional; (2) physiological;

and (3) toxic (including genotoxic) effects.

Nutritional Effects

0003 The nutritional effects of the Maillard reaction in

foods are due both to the chemical modification of

essential nutrients which thereby become unavailable

(direct effects) and to the presence of Maillard prod-

ucts which reduce the bioavailability and disturb the

metabolism of other nutrients (indirect effect).

0004 Since it proceeds through many chemical routes

and produces a large number of chemical species, the

Maillard reaction has been divided for clarity and

convenience into two distinct stages, early and ad-

vanced, which are associated with different nutri-

tional and physiological consequences (Figure 1).

The Early Maillard Reaction

0005This first stage involves the reaction between a free

amino group and a reducing sugar to form, through

the Amadori rearrangement, a stable deoxyketose

addition compound, also called the Amadori com-

pound. This is the obligatory step for the continu-

ation of the reaction to the advanced stage.

0006At this Amadori stage, nutritional damage has

already been done. No enzyme in animal tissues

can split these complexes to regenerate the amino

compounds, which are nutritionally unavailable. In

the case of amino acids, rat growth and metabolic

studies have shown that the Amadori compounds are

biologically unavailable. Cecal and large intestinal

microorganisms in experimental animals can liberate

the complexed amino acids, but this occurs too late

in the digestive/absorptive process for a significant

in vivo utilization. The nutritional loss includes

those free amino acids and peptides that have reactive

a-amino groups, protein-bound lysine which has a

reactive e-amino group, and vitamins (thiamin,

pyridoxine, and folic acid).

0007In a model system representative of the early stage,

comprising milk powder stored at 60



C or lower for

several weeks, the most quantitatively important nu-

trient which is damaged is lysine. This is due to its high

level in milk protein compared to the other amino

compounds and of the high reactivity of its e-amino

group (Table 1). Milk is one of the most sensitive foods

to this reaction because of its high content of the redu-

cing sugar lactose in addition to lysine. The most im-

portant negative consequence of the early Maillard

reaction is therefore the ‘blockage’ of lysine in milk-

based products (infant formulae and weaning foods).

Lysine is essential for growth and its requirement is

high (103 mg kg

1

day

1

for babies as compared to

12 mg kg

1

day

1

for adults). The high, recommended

lysine level in baby formulae (minimum of 6.7 g per

100 g of protein, equal to the level in mother’s milk) is

reached using cows’ milk which contains an excess of

lysine of at least 20% compared to mother’s milk.

0008Industrially treated milks contain a certain amount

of such blocked lysine as its Amadori compound, the

amount varying between 0 and 15% depending on

the treatments applied (Table 2). Higher values are

reached with the roller drying process, though this

process is no longer used in the industrial-scale pro-

duction of milk formulae.

0009Lysine bioavailability is alsoaffectedby the Maillard

reaction in otherheat-treated foods likebread, biscuits,

and pastas, but the negative nutritional consequences

BROWNING/Toxicology of Nonenzymatic Browning 673

are much lower than for milk-based infant formulae

as these cereal foods are poor sources of lysine.

Quantification of Lysine Damage

0010 Many biological and chemical methods have been

developed to quantify available and reactive lysine

respectively. Available lysine (¼ reactive lysine  di-

gestibility) may be evaluated after enzymic hydro-

lysis, by microbiological tests and, in animals, by

growth tests. Reactive lysine may be measured as

e-N-dinitrophenyl-lysine after derivatization with

fluorodinitrobenzene, as homoarginine after derivati-

zation with O-methylisourea, and as lysine after re-

duction of the Amadori compound with sodium

Reactions

Amadori compound

Advanced 1, 2-Dicarbonyl

Dehydration

Fission

Polymerization

Heterocyclic

amines

Melanoidins

Strecker

degradation

Creatinine

Short-chain

carbonyls and

dicarbonyls

Premelanoidins

N-heterocyclics

Aldehydes

+ NH

Hydroxymethylfurfural

2, 3-Dicarbonyl +

Reducing sugar

+ amino acid

Amino

acid

Biological effects

Amino acid loss

(lysine)

Vitamin loss

Mutagens

Antimutagens

Physiological

effects

Mutagens/

carcinogens

Metal

chelation

Amino acid loss

(lysine, methionine,

tryptophan)

Stages

Early

fig0001 Figure 1 Browning reaction. Chemical pathways and biological effects. Reproduced from Browning: Toxicology of Nonenzymatic

Browning, Encyclopaedia of Food Science, Food Technology and Nutrition, Macrae R, Robinson RK and Sadler MJ (eds), 1993, Academic

Press.

tbl0001 Table 1 Nutritional effects of early and advanced Maillard reactions. Model system: dried milk powder

Maillard

status treatments

Percentage of original values of untreated sample

Reactive

lysine

Lysine as

Amadori compound

Lysine in

advanced Maillard

products

Vitamin B

Pantothenic

acid

Vitamin B

Folic acid Vitamin B

Early



C, 4 weeks 74 26 0 81 90 84 29 100

Advanced



C, 4 weeks 17 14 69 5 7 18 3 31

Tryptophan

Reactive lysine Nitrogendigestibility Chemical analysis Bioavailability Methionine

Early



C, 9 weeks 79 98 100 100 100

Advanced



C, 4 weeks 20 75 100 92 92

674 BROWNING/Toxicology of Nonenzymatic Browning