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

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disaccharides sucrose and lactose consist of 50% glu-

cose and 50% fructose or galactose, respectively, both

have a lower glycemic index than maltose, the disac-

charide formed from two glucose molecules.

Amylose and Amylopectin

0006 The starches in cereal grains, rice, potatoes, and all

green plants are composed of repeating glucose units

arranged in straight, amylose, and branched-chain,

amylopectin polysaccharides. The absorption rate,

and hence the glycemic index, of these starches is

influenced by the ratio of amylose to amylopectin.

The more compact structure of amylose than amylo-

pectin results in a smaller surface area being available

for amylase digestion. Amylose-enriched starches

have lower glycemic indexes than those enriched in

amylopectin.

Relationship of Insoluble and Soluble Nonstarch

Polysaccharides (Fiber) to Glycemic Index

0007 Nondigestible complex carbohydrates are commonly

known as dietary fiber. The more correct terminology

is nonstarch polysaccharides (NSP). Dietary fiber/

NSP is either soluble or insoluble. Clinical studies

have shown that diets rich in soluble fiber/NSP, such

as guar gum, pectin, and sugar beet fibers lower post-

prandial blood glucose and insulin levels. Guar gum,

a beta-glactomannan from the Indian locust bean,

also reduces postprandial lipemia. Nonsoluble NSP

has no effect on dietary glycemic index.

0008 Soluble NSP such as pulses, vegetables, whole

fruits, oats, and barley form gelatinous gels within

the stomach which delay gastric emptying and en-

zymic digestion, the latter by forming a physical bar-

rier around the carbohydrate. Insoluble NSPs have

little effect on gastric emptying and no effect on glu-

cose absorption. High-fiber/high-NSP diets are there-

fore not necessarily synonymous with low-glycemic

foods. Cellulose is the most widely used NSP in

household cereals, wholemeal bread, and brown

rice, and as cellulose is insoluble, these foods have

the same glycemic index whether replete or deplete of

their dietary fiber/NSP. For unknown reasons, All

Bran is an exception and, despite its high insoluble

fiber content, has a low glycemic index.

0009The solubility of dietary fiber/NSP needs to be

specified if benefits on postprandial glycemia and

hyperinsulinemia are to be expected. Nonmetabolic

benefits are derived from increasing nonsoluble fiber/

NSP, and are notable on bowel function and bowel

pathology. However, protection against colon cancer

has recently been questioned.

Cell Structure, Food Preparation, and Processing

0010Cooking and food preparation can modify the glyce-

mic index. Highly processed convenience foods tend

to have high glycemic indexes. When cooking and

processing disrupt the cell wall of starch granules,

the glycemic index is increased. Cooked pulse vege-

tables have low glycemic indexes as their cell walls

are resistant to cooking. The intact cereal grains of

pumpernickel rye bread, granary bread, and burgule

wheat all have low glycemic indexes. However, when

granary bread is processed to wholemeal bread these

grains are disrupted and the glycemic index rises.

Cooling can paradoxically lower the glycemic index

of certain starches, such as potatoes, due to the for-

mation of retrograde starches which are resistant to

amylase digestion. Food particle size as well as the

constituency of the food bolus on swallowing also

influence absorption rates, as seen with boiled and

mashed potatoes.

Concerns Relating to the Glycemic Index

0011Despite the 1998 World Health Organization/Food

and Agriculture Organization nutritional report

advocating greater use of the glycemic index, no

American or British dietetic guidelines promote its

use. In fact, the only guidelines that do support the

use of the glycemic index are the 1998 European

guidelines.

tbl0002 Table 2 Comparison of common carbohydrate-containing foods and their glycemic index

Food type Carbohydrate type Glycemic index

Refined Unrefined Starch Sugar Soluble Nonsoluble

White bread [[ 100

Wholemeal bread [[ [ 100

Weetabix [[ [ 100

Cornflakes [[ 121

Porridge [[ [ 89

Baked beans [[[ 70

Digestive biscuits [[ [ 82

Apple [[ [ 49

Sucrose [[ 83

2918 GLUCOSE/Glucose Tolerance and the Glycemic (Glycaemic) Index

0012 Carbohydrates are still broadly grouped on their

biochemical basis into refined and complex carbohy-

drates; starch and NSPs and soluble and insoluble

fiber. Part of the failure of the glycemic index not

being more widely used is due to concerns relating

to its reproducibility, accuracy when part of a mixed

meal, and its overall validity. In terms of reproduci-

bility, the glycemic index, like all in vivo measure-

ments, such as the oral glucose tolerance test, is

dependent on physiological variables and, if these

are not controlled for, some variability in its measure-

ment is inevitable. Concerns relating to the glycemic

index when part of a mixed meal are theoretical

rather than actual, such as the glycemic response of

a mixed meal, can be reliably calculated knowing the

glycemic index of each constituent carbohydrates and

their proportional contribution to that meal. The

concerns on validity are centered on the way the

glycemic index is calculated, emphasizing the 2-h

integrated area rather than the pattern of response.

The way the glycemic index is calculated can result in

two obviously metabolically different carbohydrates,

which would evoke different patterns of glucose

and insulin release having similar glycemic indexes

(Table 2), for example, a rapidly absorbed and me-

tabolized carbohydrate, such as sucrose, and a com-

plex carbohydrate with poor intestinal absorption

and subsequent colonic fermentation. However, in

practice the glycemic index of most carbohydrates

provides a good indicator of the 2-h plasma glucose

and insulin exposure.

Clinical Significance of Postprandial

Versus Fasting Hyperglycemia in Diabetic

and Nondiabetic Populations

0013 Postprandial hyperglycemia in nondiabetic popula-

tions is a stronger predictor of insulin resistance and

CVD than fasting glucose. The combined 20-year

mortality data on men from the Whitehall, Paris Pro-

spective, and Helsinki Policemen studies showed that

the upper quintile compared with the lower for the

2-h postplasma load glucose was associated with a

2.7 times increased risk of CVD mortality. The fasting

glucose values were less discriminatory for CVD,

with only the upper 2.5% values conferring a 1.8-

fold increased mortality risk. Over a 7-year period,

elderly women in a study with isolated postprandial

hyperglycemia, 2-h value > 11.1 mmol l

1

and fasting

value < 7.0 mmol l

1

on a 75-g oral glucose tolerance

test had an approximately threefold increased risk of

heart disease when compared with women whose 2-h

values were below 11.1 mmol l

1

0014 In established diabetes postprandial glycemia

appears to have a stronger relationship with

microvascular and macrovascular disease than fasting

blood glucose. Similarly, in gestational diabetes

adverse pregnancy outcome is more closely related

to postprandial glycemia than fasting and premeal

glycemic values.

The Benefits of Low-Glycemic-Index

Carbohydrates on Diabetic Control

0015Three-month clinical studies have shown that low-

glycemic-index carbohydrates improve glycemic

control in both type 1 and type 2 diabetes and reduce

postprandial glucose and insulin values. Good

glycemic control and favorable lipid and fibrinolytic

profiles have also been reported in individuals

with type 1 and 2 diabetes who habitually consume

low-glycemic-index dietary carbohydrates. It remains

to be seen whether these diets bestow long-term bene-

fits on micro- or macrovascular complications.

The Benefits of Low-Glycemic-Index

Carbohydrates on CVD Risk Factors

0016High-glycemic-index foods induce postprandial

hyperinsulinemia, which is a powerful predictor for

metabolic risk factors and CVD in epidemiological

studies. Both cross-sectional and prospective popula-

tion studies have shown favorable lipid profiles in

association with high-carbohydrate diets. Initially,

these benefits were attributed to a high fiber content.

However, when the glycemic index is controlled for,

it is the low-glycemic-index diets rather than high

fiber content that have the greatest influence on

high-density lipoprotein (HDL)-cholesterol, insulin

sensitivity, and fibrinolytic parameters. In one large

cross-sectional study on middle-aged ambulatory

subjects, glycemic index was a stronger determinant

of HDL cholesterol than any other dietary carbo-

hydrate or fat. Low-glycemic-index diets also lower

serum cholesterol and triglyceride levels in hyper-

lipidemic subjects.

Potentially Detrimental Effects of a High-

Carbohydrate Diet

0017High-carbohydrate diets (> 60% of total dietary

energy) that consist predominantly of high glycemic

carbohydrates have detrimental metabolic effects.

These diets increase serum triglyceride and insulin

resistance, having the greatest adverse effect in

insulin-resistant states, such as type 2 diabetes or

pregnancy. As a consequence of this, the American

Diabetic Association recommends limiting dietary

carbohydrates to 45% in diabetic pregnancies and

to 80% of total energy derived from carbohydrate

GLUCOSE/Glucose Tolerance and the Glycemic (Glycaemic) Index 2919

and fat outside pregnancy. By contrast, the British

Diabetic Association’s guidelines advocate that 55%

of total dietary energy should come from dietary

carbohydrates. The European guidelines are the only

ones that specify the use of low-glycemic carbohy-

drates and the current American Dietetic Association

recommendations for the USA give glycemic index an

evidence-based rating of B, suggesting more long-

term studies are needed.

0018 Several short-term clinical studies have shown that

high-carbohydrate/low-glycemic-index diets reduce

insulin sensitivity and adversely influence lipid pro-

files in both normoglycemic and type 2 diabetic sub-

jects. Similar results have been seen in healthy

pregnancy and in individuals at risk of cardiovascular

disease. In all these studies, substitution of the dietary

carbohydrate with a low glycemic index reverses

these trends. Interestingly, the very-low-density-lipo-

protein (VLDL)-rich triglyceride particles generated

from high-carbohydrate diets are thought to be less

atherogenic than those associated with the insulin

resistance syndrome.

0019 High-carbohydrate diets have been associated

with both the development of and the prevention of

type 2 diabetes and CVD. These conflicting studies

are likely to be explained by differences in dietary

glycemic index. Previously many clinical and epi-

demiological dietary studies have not specified

the glycemic index. High-carbohydrate diets in non-

industrialized rural populations, such as Africa,

are typified by low-glycemic starches and soluble

NSP, carbohydrates associated with reduced insulin

resistance, and metabolic CVD risk factors. By

contrast, high-carbohydrate diets in westernized

industrial societies such as North America, with

high background incidences of obesity, insulin resist-

ance, and type 2 diabetes, contain high-glycemic

carbohydrates due to food processing and soft drinks,

and it is these diets which are metabolically disadvan-

tageous.

Obesity and Glycemic Index

0020 Obesity contributes to the pathogenesis and morbid-

ity of type 2 diabetes. Obesity is associated with

changes in carbohydrate and fat metabolism that are

central to the development of insulin resistance.

While low-glycemic-index diets reduce insulin sensi-

tivity and improve metabolic cardiovascular risk

factors, they will not reduce weight unless part of an

energy-deficient diet. However, in obese subjects,

when low-glycemic carbohydrates are incorporated

into a hypocaloric diet, there is a greater fall in insulin

resistance than can be accounted for by weight loss

alone.

Glycemic Index and the Prevention of

Type 2 Diabetes

0021Changes in diet and physical activity levels, both

alone and in combination, reduce the progression of

impaired glucose tolerance to diabetes. Two large

American prospective population studies have dem-

onstrated protection against type 2 diabetes for both

men and women when the habitual diet is character-

ized by having a low glycemic index and low fat

content. A similar protective effect against diabetes

has been reported in populations consuming high-

fiber foods and high quantities of fruit and vegetables.

Although it was not calculated, one would predict

that these diets would also have a low glycemic index.

Pregnancy and Glycemic Index

0022Throughout pregnancy in well-nourished urbanized

women consuming typical westernized diets, there is

a deterioration in glucose tolerance. African women

during pregnancy, living in traditional rural popula-

tions and consuming high-carbohydrate/low-glycemic-

index diets, do not invariably experience deterioration

in their glucose tolerance. Clinical studies in the west

show that women consuming similar high-carbohy-

drate/low-glycemic-index diets throughout pregnancy

also have no deterioration of glucose tolerance despite

the physiological increase in insulin resistance that

occurs secondary to maternal and placental hormones.

When the proportion of dietary carbohydrate increases

above 50% in women with gestational diabetes, if no

emphasis no low-glycemic-index carbohydrates is

given, glucose tolerance will deteriorate.

Proposed Mechanism by which Dietary

Carbohydrates Influence Insulin

Resistance

0023Adipocyte metabolism is central to the pathogenesis

of insulin resistance and dietary carbohydrates influ-

ence adipocyte function. The previous simplistic view,

that insulin resistance resulted from the downregula-

tion of the insulin receptors in response to hyperinsu-

linemia, is being replaced by the hypothesis that high

circulating nonesterified fatty acid (NEFA) levels

both impair insulin action and reduce pancreatic

b-cell secretion. A plausible explanation of how

low-glycemic-index carbohydrates can reduce insulin

sensitivity is by their ability to reduce adipocyte

NEFA release.

0024Adipocyte release of NEFA is increased in insulin

resistance as a consequence of suppressed hormone-

sensitive lipase activity. An inverse relationship be-

tween adipocyte NEFA release and insulin sensitivity

2920 GLUCOSE/Glucose Tolerance and the Glycemic (Glycaemic) Index

has been demonstrated in patients with coronary

heart disease. High circulating NEFA levels reduce

muscle uptake of glucose and downregulate the

insulin receptor. High circulating NEFA levels reduce

hepatic insulin clearance; this exaggerates the pre-

existing hyperinsulinemia and insulin resistance.

Reduced lipoprotein lipase activity increases NEFA

levels and hepatic VLDL production; however, VLDL

clearance is reduced, and this results in a lower high-

density lipoprotein concentration and increased TG-

enriched VLDL particles. The latter are catabolized

into smaller atherogenic forms of LDL. The meta-

bolic consequences of increased adipocyte NEFA re-

lease therefore fuel lipoprotein changes associated

with coronary heart disease.

0025 High circulating NEFA levels also adversely affect

insulin secretion. High NEFA levels have been postu-

lated to have a lipotoxic effect on the b-cells due to

their ability to increase nitric oxide production which

is cytotoxic to islets. In addition, when triglyceride

accumulates within the b-cell, glucose-stimulated

insulin secretion is blunted.

0026 Many of the metabolic benefits associated with

low-glycemic-index carbohydrates can be attributed

to their ability to reduce adipocyte NEFA release.

Low-glycemic-index foods have been consistently

shown to improve insulin sensitivity and animal stud-

ies have shown that improvements in fat and muscle

insulin sensitivity are accompanied by decreases in

fatty acid synthatase activity, adipocyte size, and

lipid storage. Although human studies have shown

that low-glycemic-index diets consumed over 3 weeks

increase adipocyte insulin sensitivity, no direct effect

on adipocyte metabolism has been identified so far.

0027 Low-glycemic-index diets, by attenuating the insu-

lin response, ensure a more prolonged suppression of

postprandial NEFA levels than those following high-

glycemic foods. Low-glycemic meals taken in the

evening can effectively suppress circulating NEFA

concentrations and hepatic glucose output through-

out the night. These metabolic effects would be pre-

dicted to promote insulin sensitivity.

0028 Our own work has shown that insulin-resistant

adults with a history of, or who are at risk of CHD

improve their adipocyte insulin sensitivity when con-

suming a low-glycemic-index diet and in addition

their circulating NEFA levels decline. These human

studies complement the animal work that show low-

glycemic-index diets improve insulin sensitivity by

modulating adipocyte metabolism.

Conclusion

0029 The glycemic index of a diet provides a measure of

postprandial metabolism. Dietary carbohydrates are

absorbed and metabolized differently and therefore

influence postprandial glucose, insulin, and NEFA

concentrations differently. In western society the pro-

portion of the day that we spend in the postprandial

state is increasing as the tendency to snack throughout

the day replaces sit-down main meals. The known

detrimental consequences of high-glycemic foods on

postprandial metabolism should encourage us to

advocate low-glycemic diets to counteract the present

epidemic of insulin resistance-related diseases,

notably CVD and diabetes. The relevance of the gly-

cemic index to these two major preventable diseases of

the western world argues strongly for its greater ac-

ceptance into our current nutritional guidelines.

See also: Carbohydrates: Metabolism of Sugars;

Coronary Heart Disease: Antioxidant Status; Diabetes

Mellitus: Etiology; Chemical Pathology; Obesity: Etiology

and Diagnosis; Pregnancy: Nutrition in Diabetic

Pregnancy

Further Reading

Alberts DS, Ritenbaugh C, Story JA, Aickin M, Rees A,

McGee S, Buller MK, Atwood J, Phelps J, Ramanujam

PS et al. (1996) Randomized, double-blinded, placebo-

controlled study of effect of wheat bran fiber and

calcium on fecal bile acids in patients with resected ade-

nomatous colon polyps. J Natl Cancer Inst 88: 81–92.

American diabetes association (1999) Nutritional recom-

mendations and principles for people with diabetes mel-

litus. Diabetes Care 22 Suppl 1: S42–S46

Balkau B, Shipley M, Jarrett RJ, Pyorala K, Pyorala M,

Forhan A and Eschwege E (1998) High blood glucose

concentration is a risk factor for mortality in middle-

aged nondiabetic men. 20-year follow-up in the White-

hall Study, the Paris Prospective Study, and the Helsinki

Policemen Study. Diabetes Care 21: 360–367.

Englyst HN, Trowell H, Southgate DAT and Cummings JH

(1987) Dietary fiber and resistant starch. Am J Clin Nutr

46: 873–881.

FAO/WHO Report of a joint FAO/WHO report Rome

(1998) 14–18 April 1997, editor. Carbohydrates in

human nutrition. Paper 66. FAO Food and Nutrition.

Foster-Powell K and Miller JB (1995) International tables of

glycemic index. Am J Clin Nutr 628: 71S–93S.

Frost G, Leeds A, Trew G, Margara R and Dornhorst A

(1998) Insulin sensitivity in women at risk of coronary

heart disease and the effect of a low glycaemic index

diet. Metabolism 47: 1245–1251.

Frost GS, Keogh B, Smith D, Leeds AR and Dornhorst A

(1998) Reduced Adipocyte Insulin sensitity in Caucasian

and Asian Subjects with Coronary Heart Disease. Dia-

betes Medicine 15: 1003–1009.

Frost G, Leeds AR, Dore CJ, Madeiros S, Brading S and

Dornhorst A (1999) Glycaemic index as a determinant

of serum HDL-cholesterol concentration. Lancet 353:

1045–1048.

GLUCOSE/Glucose Tolerance and the Glycemic (Glycaemic) Index 2921

Fuchs CS, Giovannucci EL, Colditz GA, Hunter DJ, Stamp-

fer MJ, Rosner B, Speizer FE and Willett WC (1999)

Dietary fiber and the risk of colorectal cancer and aden-

oma in women. New Eng J Med 340: 169–176.

Ha KK and Lean MEJ (1998) Recommendations for the

nutritional management of patients with diabetes

mellitus. Eur J Clin Nutr 52: 467–481.

Hanefield M, Temelkova-Kurktschiev T, Schaper F, Henkel

E, Siegert G and Koehler C (1999) Impaired fasting

glucose is not a risk factor for atherosclerosis. Diabet

Med 16: 212–218.

Holt SHA, Miller JCB and Petocz P (1997) An insulin index

of foods: the insulin demand generated by 1000-kJ por-

tions of common foods. American Journal Of Clinical

Nutrition 66: 1264–1276.

Jarvi AE, Karstrom BE, Granfeldt Y, Bjorck I, Asp NG and

Vessby B (1999) Improved glycemic control and lipid

profile and normalized fibrinolytic activity on a low-

glycaemic index diet in type 2 diabetes. Diabetes Care

22: 10–18.

Jenkins DJA, Kendall CWC and Ransom TPP (1998) Diet-

ary fiber, the evolution of the human diet and coronary

heart disease. Nutrition Research 18: 633–652.

Kabir M, Rizkalla SW, QuignardBoulange A, GuerreMillo

M, Boillot J, Ardouin B, Luo J and Slama G (1998) A

high glycemic index starch diet affects lipid storage-

related enzymes in normal and to a lesser extent in

diabetic rats. Journal Of Nutrition 128: 1878–1883.

McGarry JD and Dobbins RL (1999) Fatty acids, lipotoxi-

city and insulin sectretion. Diabetologia 42: 128–138.

GLUCOSINOLATES

B Holst, Nestle

Research Center, Lausanne,

Switzerland

G R Fenwick, Institute of Food Research,

Norwich, UK

Introduction

0001 Glucosinolates are secondary plant metabolites that

are characteristic of the Cruciferae and related

families within the order Capparales. Glucosinolate-

containing plants have always made major contribu-

tions to the diets of humans and farm animals, and

include condiments and relishes (mustard and

wasabi), traditional leaf vegetables (cabbage, swede),

and crops used as animal feedstuffs (rapeseed, kale,

turnip).

0002 Glucosinolates are thioglycosides that differ in the

structure of the aglycone side chain; they are rela-

tively biologically inactive per se, but following tissue

disruption they undergo hydrolysis to form a broad

range of structurally diverse hydrolysis products pos-

sessing varying biological activities. These products

are the basis of the sensory characteristics typical of

brassicas but also, through their biological effects,

affect their nutritional value. Some of the hydrolysis

products (goitrin, thiocyanate ion, several isothiocya-

nates, and nitriles) may have antinutritional or toxic

effects; others, especially isothiocyanates (e.g., sulfor-

aphane), are considered to be responsible for the

protective, anticarcinogenic effects of a cruciferous-

rich diet. These adverse and beneficial effects are

highly dose-dependent and the physiological range is

relatively narrow. Glucosinolate hydrolysis products

also exert antifungal, antimicrobial, and insecticidal

properties, and so contribute to the plant’s overall

defense mechanism.

0003The large number of glucosinolates, the plethora of

hydrolysis products possessing differing biological

activities, and the dose dependence of the effects ob-

served make research in this area both challenging

and complex. Thus, plant-breeding strategies that

concentrated solely on reducing the glucosinolate

content of rapeseed for nutritional benefits succeeded

in producing very low (‘zero’) glucosinolate varieties

that were more nutritious but these required a signifi-

cantly increased expenditure for crop protection. On

the other hand, pest-resistant crop cultivars that con-

tain high amounts of individual glucosinolates may

have unacceptable sensory characteristics and may,

possibly, also exhibit antinutritional properties.

Chemical Structure, Properties, and

Nomenclature

0004Naturally occurring glucosinolates are (Z)-cis-N-

hydroximinosulfate esters, possessing a sulfur-linked

b-d-glucopyranose moiety and an amino acid-derived

side chain (R

: Figure 1). The side chain and sulfate

group have an antistereochemical configuration,

the structure of the former being highly variable;

the glucosinolates may posses aliphatic (alkyl, alke-

nyl, hydroxyalkenyl, o-methylthioalkyl), aromatic

(benzyl, substituted benzyl), or heterocyclic (indolyl)

side chains. Aliphatic- and aromatic glucosinolates

share the same degradation pathways but glucosino-

lates with side chains possessing a terminal double

bond or a b-OH group form different products.

The major glucosinolates may be subdivided into

2922 GLUCOSINOLATES

aliphatic and aromatic (R

1.1

), terminally unsaturated

1.2

), b-hydroxy-(R

1.3

), and indolyl (R

1.4

) groups

according to their breakdown. The first group in-

cludes a wide range of homologs (n ¼3ton ¼11)

and structural classes. The most common side chains

include o-methylthio (3), o-sulfinyl (4), o-sulfonyl-

alkyl (5), and aromatic (6) groups.

0005 The presence of the sulfate group confers strongly

acidic properties on glucosinolates, irrespective of the

chemical structure of the side chain, and they occur as

salts with the glucosinolate anion usually counter-

balanced by potassium, less frequently by sinapinium

cation. The sulfate group and thioglucose moiety

make glucosinolates nonvolatile and hydrophilic,

and the naturally occurring forms exhibit levrotation

in solution. Trivial nomenclature was used for many

years, but latterly a more precise system has been

developed that uses prefixes characteristic of the

side chain followed by the word ‘glucosinolate.’ Al-

though this nomenclature is more informative and

prevents confusion, the trivial names are still widely

used.

Occurrence

0006 Glucosinolates are limited to a restricted number

of dicotyledonous angiosperms and are prevalent

throughout 15 botanical families of the order

Capparales, e.g., the Brassicaceae, Capparaceae,

Resedaceae. As more than 80% of glucosinolates so

far identified occur in the Brassicaceae, their presence

has been used as a chemotaxonomic criterion for

classification within this family. Species of the Brassi-

caceae are of great economic importance as vege-

tables (e.g., cabbage, broccoli, cauliflower, Brussels

sprouts; root vegetables, e.g., radish, turnip, swede,

and leaf vegetables, e.g., rocket salad), relishes (e.g.,

wasabi, mustard), oilseeds (canola, rapeseed), and

forage crops (kale, forage rape).

0007Glucosinolates have been found in all parts of these

plants and, whilst as many as 15 different glucosino-

lates have been found in the same plant, three to four

usually predominate. Occurrence and concentrations

vary according to species and cultivar, tissue type,

physiological age, plant health, agronomic practice,

and nutrition. Levels in reproductive tissues (flowers

and seeds) are much higher (up to 10% dry weight)

than in vegetative tissues (1–5000 ppm). Glucosino-

lates are synthesized in the silique walls from rela-

tively few protein amino acids and chain-elongated

homologs and translocated to the seeds, where they

accumulate inside the vacuole.

0008The common glucosinolate biosynthetic pathway

involves aglycone synthesis via initial N-hydroxyla-

tion, oxidative decarboxylation to form an aldoxime,

incorporation of sulfur to yield a thiohydroximic acid

General structure of glucosinolates

Common structural part

Aliphatic and aromatic

1.1

regular un- and

substituted

(6)

(CH

1.2

unsaturated,

terminally unsaturated

(CH

1.3

β-hydroxy R

1.4

indolyl

Classification of the major glucosinolates according to the formation of breakdown product

= Side chain

−

(3)

(CH

(4)SO

(CH

(7)

(5)

(1)

(9)

(8)

(CH

(2)

(CH

fig0001 Figure 1 General and individual glucosinolate structures.

GLUCOSINOLATES 2923

that undergoes S-glucosylation to the desulfogluco-

sinolate and, finally, sulfation. Modifications of the

side chain, including oxidation, hydroxylation, and

reduction, are responsible for the observed structural

diversity of glucosinolates.

0009 Although several individual steps of the overall

biosynthetic pathway remain to be elucidated and

several enzymes require purification and character-

ization, existing knowledge may be exploited for

breeding or biotechnological production of new

plant varieties with enhanced flavor and functional-

ity, including putative health-promoting activity.

Thus breeding has produced rapeseed varieties with

very low glucosinolate concentrations, e.g., the

‘double zero’ rapeseed with a very low content of

2-hydroxy-3-butenylglucosinolate, the precursor of

5-vinyloxazolidine-2-thione (goitrin). Recently there

have been attempts to increase levels of certain

glucosinolates in crops used as human foods, e.g.

enhancing 4-methylsulfinylbutylglucosinolate, the

precursor of the anticarcinogenic isothiocyanate, sul-

foraphene.

0010 The role of the complex myrosinase (b-thiogluco-

sidase, EC 3.2.3.1.)-glucosinolate system in plants is

considered to be diverse; glucosinolates may repre-

sent a sink for nutrients like nitrogen, sulfur, or

growth factors (e.g., indole-3-acetic acid, IAA) and,

hence, may be involved in growth regulation, while

their hydrolysis products contribute to nonspecific

plant defense mechanism against insect, fungi, and

microorganism infections (Table 1).

0011 Whilst disruption of cellular tissue leads to hydro-

lyses of glucosinolates, under some conditions (after

plant–fungi, plant–insect, and plant–microorganism

interactions, or mechanical wounding) de novo syn-

thesis may be induced. Thus, under conditions of low

stress and minor cell damage, de novo synthesis of

glucosinolates may balance – or indeed exceed –

losses due to their breakdown. More work is required

to establish the underlying biochemical mechanisms

controlling these processes.

Hydrolysis

0012As emphasized above, hydrolysis products – rather

than intact glucosinolates – are responsible for the

biological activities and flavor characteristic of cru-

ciferous vegetables. Depending on reaction condi-

tions, glucosinolate breakdown may be enzymatic

and/or nonenzymatic; since myrosinase co-occurs

with glucosinolates in plant tissue, the former pre-

dominates, with chemical hydrolysis only taking

place where myrosinase is inactivated, e.g., after

cooking, or under highly acidic/basic conditions.

Myrosinase isoenzymes and glucosinolates are local-

ized in all cells but are compartmentalized; glucosi-

nolates are stored in the vacuole and myrosinases are

localized in cytoplasm.

0013Following tissue damage, enzyme and substrate

come into contact, causing hydrolysis of the S-

glucose bond and, thereby, yielding glucose and an

unstable aglycone (10, Figure 2) that undergoes spon-

taneous rearrangement to an isothiocyanate, or nitrile

and elemental sulfur. Only allyl-, benzyl-, and 4-

methylthiobutyl glucosinolates appear to undergo

enzymatic degradation to thiocyanates, although the

mechanism remains obscure. Other products, such as

epithionitriles and oxazolidine-2-thiones, can also be

formed; the nature of the resulting product depending

on side-chain structure and reaction conditions (pH,

temperature, presence of protein cofactors modifying

the action of the enzyme).

Structure of the Side Chain

0014At a pH of 6–7, most glucosinolates yield stable iso-

thiocyanates (11, 12). However, those possessing a

b-hydroxylated side chain form unstable hydroxy-

isothiocyanates that spontaneously cyclize to oxazo-

lidine-2-thiones (13), the best-known example being

5-vinyloxazolidine-2-thione (goitrin). Indole gluco-

sinolates also form unstable isothiocyanates; these

undergo lysis, initially forming the corresponding

alcohol, e.g., indole-3-carbinol (14) and subsequently

condensing to the dimer (15), trimer, or tetramer. In

the presence of ascorbic acid, ascorbigen (16) is the

major product (Figure 3).

0015Depending on the plant species, autolysis of fresh

material (pH * 6) can yield nitriles (17–21). Several

authors have discussed a ‘nitrile-forming factor’ but

this has neither been isolated nor characterized.

Under acidic conditions, nitriles are the major deg-

radation products (17, 18, 20, 21: Figure 4). In the

presence of active epithiospecifier protein (ESP) and

2þ

ions, glucosinolates with terminally unsaturated

side chains transfer sulfur from the S-glucose moiety

to the alkenyl moiety to form an epithionitrile (19).

ESP is a small protein, which co-occurs and interacts

tbl0001 Table 1 Factors affecting glucosinolate profiles and

concentration

Biotic factors Abiotic factors

Fungi Temperature

Insects Light (ultraviolet irradiation)

Microorganisms Water supply

Animals Fertilizer

Weeds and other

competing plants

Harvest conditions

Plant density Postharvest and storage conditions

Developmental stage Processing conditions

Part of the plant (organ)

2924 GLUCOSINOLATES

S-Glucose

NOSO

−

NOSO

−

β-Thioglucosidase

(Myrosinase)

Thiohydroxamate-O-sulfonate

+ Glucose(10)

−H

−S

pH 6−7 pH 3−4; pH 5−7 (tissue-dependent:

nitrile-forming factor? ESP, Fe

)

(2.2) Nitrile pathway

(Figure 4)

(2.1) Isothiocyanate pathway

(Figure 3)

(2.3) Thiocyanate pathway

fig0002 Figure 2 General and initial step of enzymatic glucosinolate hydrolysis.

1.1

1.2

1.3

1.4

1.1

CH (CH

)n N C S

(12)

(13)

(11)

OHCH

3-Diindolylmethanes (15) Ascorbigens (16)

Trimers, tetramers

−N

(14)

− SCN

−

Isothiocyanates

Alkenylisothiocyanates

Unstable

isothiocyanates

Spontaneous

cyclization

5-substituted oxazolidine-2-

thiones

Unstable

isothiocyanates

Indole-3-carbinols−CH

+ Ascorbic

acid

fig0003 Figure 3 Glucosinolate breakdown products – Isothiocyanates (2.1, Figure 2).

Refers to Figure 1.

GLUCOSINOLATES 2925

with myrosinase but is not present in all glucosino-

late-containing species. In the absence of ESP the

addition of ferrous ions to reaction mixtures pro-

motes nitrile formation (18).

Reaction Conditions

0016 Most factors modifying glucosinolate hydrolysis

affect either myrosinase activity and specificity, or

the activity of ESP, which is a very labile protein, in

marked contrast to myrosinase. Low concentrations

of ascorbic acid increase myrosinase activity in some

species, whilst high concentrations are inhibitory. The

broad range of possible degradation pathways and

their resulting complexity, as well as the diversity

of hydrolysis products, are most likely due to their

function in plant defense mechanisms. Hydrolysis

products, particularly thiocyanate ion and isothiocya-

nates, are toxic to animals, fungi, and microbes as

well as to other plants. Catabolic detoxification is

achieved by transformation to a variety of simple

compounds that are either recycled or excreted as

volatiles such as methylisothiocyanate, methylthio-

cyanate, methylthiol, dimethyldisulfide, and disulfide.

Plant cell damage, accompanied by competing pro-

cesses of hydrolysis and de novo biosynthesis of spe-

cific glucosinolates, especially indole glucosinolates,

occurs during storage and processing of cruciferous

vegetables. Chopping, cooking, freezing, and ferment-

ing lead to enzymatic and/or nonenzymatic glucosino-

late hydrolysis and de novo glucosinolate biosynthesis

as a wound response. Leaching into cooking liquor is a

major cause of glucosinolate loss during processing.

There are thus prominent changes in glucosinolate

profiles and in the quality and quantity of hydrolysis

products formed (and hence flavor characteristics)

during processing.

0017Predicting the degree of glucosinolate loss, and the

nature of hydrolysis products formed, is difficult

because of the high number of parameters which

affect the process; these include the final size of the

plant material after chopping, the amount of cooking

water, the use of cold or boiling water at the start of

the cooking process, the nature of the processing/

cooking, cooking time, and precooking/storage

conditions (frozen material will be more heat-sensi-

tive than fresh). There are attempts to investigate

systematically the effects of single and combined pro-

cessing parameters on glucosinolate hydrolysis; by

modulating these it may eventually be possible to

predict the outcome of a given process and identify

the major parameters affecting changes in gluco-

sinolates.

1.4

Indole-3-acetonitriles Indole-3-carboxylic acids

(17)

1.1

Aliphatic/aromatic nitriles

1.2

1.3

(20)

(21)

(18)

Epithioalkylnitriles

(CH

(19)

(22)

(CH

Alkenylnitriles

Epithiospecifier protein

β-Hydroxynitriles

fig0004 Figure 4 Glucosinolate breakdown products – nitriles (2.2, Figure 2).

Refers to Figure 1.

2926 GLUCOSINOLATES

Analysis

0018 The type of glucosinolate analysis depends on the

particular data required, i.e., overall (total) concen-

tration, amount of individual glucosinolates, quantity

and type of degradation products and/or metabolites.

For screening purposes in plant breeding and/or the

study of environmental influences, the easiest, fastest,

and cheapest method to determine total glucosinolate

concentrations is by glucose release. Myrosinase-me-

diated hydrolysis of glucosinolates releases glucose

and sulfate in equimolar amounts. Both can be used

for total glucosinolate determination but glucose,

which can be measured instrumentally with a high

sample throughput and low cost, is the analyte of

choice if large numbers of samples need to be

screened.

0019 The chemical and biological properties of individ-

ual glucosinolates and their products can be very

different, which is why determination of the levels

of individual glucosinolates is often the goal. Intact

glucosinolates can be separated by high-performance

liquid chromatography (HPLC) on reverse-phase

matrices using ion pair reagents to neutralize the

strongly ionic sulfate group. Desulfation has been

used to circumvent the need for ion pair reagents

and also represents a useful – indeed, frequently ne-

cessary – clean-up step. Such a method has been

declared as a European Union (EU) reference method.

Whilst lack of standards makes identification by

ultraviolet detection at 227 nm or using diode array

detectors difficult, novel coupling techniques such as

HPLC – mass spectrometry (MS) or HPLC–MS nu-

clear magnetic resonance (NMR) have facilitated glu-

cosinolate separation and identification without

derivatization.

0020 HPLC–MS is also the method of choice for analy-

zing glucosinolate hydrolysis products and metabo-

lites; these, together with the parent glucosinolates,

can be separated on reverse-phase columns. Since

high-performance MS instruments and techniques

allow the identification and quantification of com-

pounds incompletely separated, it is now possible to

analyze complex mixtures.

0021 Extracts of plant material, biological fluids, and

cell culture media or biological tissues are all very

complex; depending on the subsequent type of analy-

sis, time- and labor-intensive extraction and purifica-

tion procedures may be necessary. The most common

extraction procedure for glucosinolates uses hot

methanol followed by separation on strong ion ex-

change materials. Novel, liquid chromatography

(LC) integrated preparation methods save time and

can improve results through on-column analyte

enrichment; quantitative and matrix-independent

recovery; increased analytical capacity; and low

throughput cost per sample.

Bioactivity

0022While glucosinolates per se are biologically inert,

their hydrolysis products possess chemical properties

rendering them powerful allelochemical agents

against microbes, fungi, and plants. They also con-

tribute significantly to the typical flavor of crucifer-

ous vegetables; however, considerable current interest

centers on their biological effects on humans and

animals, which are highly dose- and structure-

dependent, and may elicit positive, protective effects

as well as antinutritional and potentially toxic re-

sponses.

Cancer-Preventive Potential

0023The inverse correlation between Brassica con-

sumption and cancer risk is well established through

many epidemiological studies. However, only limited

human intervention studies have been conducted

(e.g., with Brussels sprouts) to confirm these results.

Mechanistic investigations, largely based on cell

culture studies, have demonstrated glucosinolate

hydrolysis products to be a source of the anticarcino-

genic effects. Thus, indole-3-carbinol derived from 3-

indolylmethylglucosinolate modulates the estrogen

hydroxylation pathway to produce a less potent form

of estradiol and, thereby, might confer protection

against estrogen-related cancer.

0024Individual glucosinolate-derived isothiocyanates,

e.g., phenethyl- and 4-methylsulfinylbutyl isothiocya-

nate (this latter commonly termed sulforaphane),

have been shown to prevent formation of chemically

induced tumors of the esophagus, stomach, liver,

lung, and mammary gland in animals, most likely

through mechanisms involving inhibition of cyto-

chrome P-450 carcinogen activation and induction

of phase II enzymes such as glutathione-S-transferase

and NAD(P)H:quinone reductase. Glucosinolate-

derived isothiocyanates may alter the balance of

phase I/II enzyme metabolism such that phase I

oxidative activation of carcinogens is decreased as

compared with the increased rate of phase II detoxifi-

cation.

0025Several isothiocyanates may also induce cell cycle

arrest and apoptosis in colon cancer cells, which

represents an alternative mechanism of anticar-

cinogenic activity. Thus, individual isothiocyanates

belong to a small, but important group of compounds

exhibiting dual inhibitory action against carcino-

genesis. It is not surprising, therefore, that glucosino-

late-derived compounds have been identified as

targets for functional food or nutraceutical product

GLUCOSINOLATES 2927