Variation of glucosinolates and nutritional value in nabicol(Brassica napus pabularia group)

Marıa Elena Cartea Æ Vıctor Manuel Rodrıguez ÆAntonio de Haro Æ Pablo Velasco Æ Amando Ordas

Received: 30 January 2007 / Accepted: 14 May 2007 / Published online: 8 June 2007

� Springer Science+Business Media B.V. 2007

Abstract Glucosinolate levels in leaves were deter-

mined in a collection of 36 varieties of nabicol

(Brassica napus pabularia group) from northwestern

Spain grown at two locations. Crude protein, acid

detergent fibre, and sensory traits were also assessed

by a consumer panel. The objectives were to

determine the diversity among varieties in total

glucosinolate content and glucosinolate profile and

to evaluate their sensory attributes in relation to

glucosinolate content for breeding purposes. Eight

glucosinolates were identified, being the aliphatic

glucosinolates, glucobrassicanapin, progoitrin, and

gluconapin the most abundant. Glucosinolate com-

position varied between locations although the gluc-

osinolate pattern was not significantly influenced.

Differences in total glucosinolate content, glucosin-

olate profile, protein, acid detergent fibre, and flavour

were found among varieties. The total glucosinolate

content ranged from 1.4 mmol g�1 to 41.0 mmol g�1

dw at one location and from 1.2 mmol g�1 to

7.6 mmol g�1 dw at the other location. Sensory

analysis comparing bitterness and flavour with var-

iation in glucosinolate, gluconapin, progoitrin, and

glucobrassicanapin concentrations suggested that

other phytochemicals are probably involved on the

characteristic flavour. The variety MBG-BRS0035

had high total glucosinolate, glucobrassicanapin, and

gluconapin contents at both locations and could be

included in breeding programs to improve the

nutritional value of this vegetable crop.

Keywords Brassica napus � Bitterness � Flavour �Glucosinolates � Nabicol � Taste panel

Introduction

Brassica napus L. includes several crops grown as

fodder, oilseed, and vegetables. The most important

crops are oilseed rape (B. napus L. oleifera group

DC.), swede (B. napus napobrassica group L.

Reichenb.), and leaf rape (B. napus L. pabularia

group (DC.) Reichenb). Crops of the last group

take the common name of ‘nabicol’ in Spain or

‘couve-nabica’ in Portugal, where are grown as fresh

green vegetables during the winter season (Cartea

et al. 2005). In these areas, crops of B. napus

pabularia group are highly appreciated for human

nutrition.

M. E. Cartea (&) � V. M. Rodrıguez �P. Velasco � A. Ordas

Department of Plant Genetics, Mision Biologica de

Galicia, Spanish Council for Scientific Research (CSIC),

Apartado 28, 36080 Pontevedra, Spain

e-mail: ecartea@mbg.cesga.es

A. de Haro

Department of Agronomy and Plant Breeding, Instituto de

Agricultura Sostenible, Spanish Council for Scientific

Research (CSIC), Alameda del Obispo s/n, 14080

Cordoba, Spain

123

Euphytica (2008) 159:111–122

DOI 10.1007/s10681-007-9463-x

Most research on the chemical composition of

Brassica species has been focussed on both the

negative and positive effects of glucosinolates, a

group of compounds derived from amino acids,

which are largely responsible for the specific flavour

of Brassica crops (Rosa 1999). The relation between

some glucosinolates with bitter taste has been already

studied in Brussels sprouts (Van Doorn et al. 1998)

and turnip greens (Jones and Sanders 2002; Padilla

et al. 2007). Some glucosinolates and their derived

products have been reported to have beneficial

properties on human health. Among their breakdown

products, isothiocyanates have been found to have

anticarcinogenic and anti-mutagenic effects (Rosa

et al. 1997; Farnham et al. 2004; Smith et al. 2005).

Besides these beneficial effects, other glucosinolates

have been associated to goitrogenic effects in animals

but the evidence for goitrogenic effects in humans is

scarce (Mithen et al. 2000). Detailed reviews of

glucosinolate content occurring in the major Brassica

crops have been reported by several authors (Fenwick

et al. 1983b; Rosa et al. 1997; Rosa 1999). Concern-

ing B. napus crops, the glucosinolate composition in

oilseed rape and swedes has been reported (Carlson

et al. 1981; Griffiths et al. 1991; Rosa et al. 1997).

However, little information is available on the

nutritional quality and the glucosinolate composition

of the leaves of B. napus vegetable crops, such as

nabicol or couve-nabica, widely used for human

consumption.

Brassica vegetables are relatively high in protein

and fibre contents compared to other vegetables of

high water content (Rosa 1999). There is renewed

interest in the use of Brassica as crops for forage

because of their high protein content and high dry

matter digestibility. In several European countries,

such as Great Britain, France or Germany, they are

used in autumn as grazing either for sheep or,

occasionally, for dairy cows (Rosa 1999). In this

regard, the vegetable types of nabicol grown in

Galicia might be also used for fodder with a mixed

use of the crop. However, it would be necessary to

know other quality attributes, such as fibre and

protein content for this eventual use.

This study has two aims: (i) to determine the

composition of glucosinolates, acid detergent fibre,

and crude protein of nabicol varieties and (ii) to study

the relation of some glucosinolates with flavour and

bitterness.

Material and methods

Plant material

The trials comprised 35 local populations from

Galicia (northwestern Spain) and one commercial

variety. The morphological description of these

nabicol varieties have been recently described (Rod-

rıguez et al. 2005). Trials were performed during two

years (2002 and 2003) in two locations in northwest-

ern Spain: Pontevedra (42824’ N, 8838’ W) and

Fornelos de Montes (42820’ N, 8826’ W). Both

locations have a humid climate with an annual

rainfall of about 1600 mm in Pontevedra and about

3500 mm in Fornelos de Montes. The soil type is acid

sandy loam. Varieties were evaluated in a 6 · 6

lattice design with three replications. Each experi-

mental plot consisted of two rows with 10 plants per

row. Rows were spaced 0.9 m apart and plants

between rows 0.6 m apart. Cultural operations,

fertilization and weed control were made according

to local practices. Leaves were harvested four months

after transplanting from the experimental fields

described by Rodrıguez et al. (2005) for glucosino-

late, fibre and protein analyses and the consumer

panel test.

Glucosinolate analysis

A sample of five healthy and fresh leaves was

collected from five plants from each variety at each

location. The five upper leaves per plant (the two next

to the apical leaf along with the adjacent three leaves)

were sampled because they are the tender leaves used

for human consumption. Leaf samples were frozen

in situ and were taken immediately into the labora-

tory where they were stored at �808C. Then, they

were ground in liquid N2, freeze-dried and milled to a

fine powder for the glucosinolate extractions. Gluco-

sinolate composition was determined by HPLC

according to Font et al. (2005). For each leaf sample,

100 mg dry wt was weighed and ground in a Janke

and Kunkel, Model A10 mill (IKA-Labortechnik) for

about 20 s and a two-step glucosinolate extraction

was carried out in a water bath at 758C to inactivate

myrosinase. In the first step, the sample was heated

for 15 min in 2.5 ml 70% aqueous methanol and

200 ml 10 mM sinigrin (2-propenyl glucosinolate) as

an internal standard. A second extraction was

112 Euphytica (2008) 159:111–122

123

conducted after centrifugation (5 min, 5000 g) by

using 2 ml of 70% aqueous methanol. One ml of the

combined glucosinolate extracts was pipetted onto

the top of an ion-exchange column containing 1 ml

Sephadex DEAE-A25 in the formate form. Desulph-

ation was carried out by the addition of 75 ml of

purified sulphatase (E.C. 3.1.6.1, type H-1 from Helix

pomatia) (Sigma) solution. Desulphated glucosino-

lates were eluted with 2.5 ml (0.5 ml · 5) Milli-Q

(Millipore) ultra-pure water and analysed with a

Model 600 HPLC instrument (Waters) equipped with

a Model 486 UV tunable absorbance detector

(Waters) at a wavelength of 229 nm. Separation

was carried out by using a Lichrospher 100 RP-18 in

Lichrocart 125-4 column, 5 mm particle size (Merck).

HPLC solvents and gradient followed the ISO

protocol (ISO Norm 1992). The HPLC chromatogram

was compared to the desulpho-glucosinolate profile

of three certified reference materials recommended

by U.E. and ISO (CRMs 366, 190 and 367). The

amount of each individual glucosinolate present in

the sample was calculated with an internal standard,

and expressed as mmol g�1 of dry wt. The total

glucosinolate content was computed as the sum of all

the individual glucosinolates present in the sample.

Data were corrected for UV response factors for

different types of glucosinolates (ISO Norm 1992).

Protein and fibre analysis

For the protein and fibre analysis, a sample of five

fresh leaves was collected from five plants from each

variety at each location. Leaves (lamina and petioles)

were detached from the stems and both were oven-

dried at 608C for at least 48 h to determine dry matter

concentration. Dried samples were ground (1 mm

screen) for subsequent laboratory analysis. The ADF

content was analysed by Van-Soest method and leaf

protein concentration by Kjedhal method following

the protocols of AOAC (2000).

Consumer panel

A sample of healthy and fresh leaves was collected

from five plants from each plot at each location.

Twenty five to thirty tender leaves from each plot

were harvested according to the maturity cycle of

each variety at the optimum time for consumption

and boiled for 2 min in 1 dm3 of water with 5 g salt.

The taste panel consisted of 12 members, all of them

regular consumers of vegetable brassica crops.

Hardness, bitterness, fibrousness and flavour were

scored on a continuous scale from ‘1’ to ‘5’. For

bitterness and fibrousness, a rating of 1 was consid-

ered ‘slight’ and 5 ‘high’; for hardness a rating of 1

was considered ‘tender’ and 5 ‘hard’. Finally, for

flavour, a rating 1 was ‘very bad’ and 5 ‘very good’.

The consumer panel evaluation lasted 36 days, as no

more than 6 samples per day could be tasted.

Statistical analyses

The experiment was set up in a completely random

design and glucosinolate content per plant analyzed

by individual analysis of variance. Varieties were

considered as fixed effects. Comparisons of means

among varieties were performed for each trait using

the Fisher’s protected least significant difference

(LSD) at P = 0.05 (Steel et al. 1997). A combined

analysis of variance across locations was carried in a

randomized block design with each location as a

block and using the location · variety interaction as

the experimental error. Varieties were considered as

fixed effects and locations as random effects.

For sensory characteristics, analyses of variance

were performed for each trait and combined over

locations according to a completely randomized

block design and using the GLM procedure of SAS

(SAS Institute 2000). Replications and locations were

considered as random factors while varieties were

considered as fixed factors. Comparison of means

among varieties were made by Fisher’s protected

significant difference (LSD) at P = 0.05 (Steel et al.

1997). All analyses were performed with the SAS

statistical package (SAS Institute 2000). Phenotypic

correlation coefficients were estimated among some

glucosinolates with bitterness and flavour (Johnson

et al. 1955).

Results and discussion

Variability of glucosinolates in B. napus

collection

Eight glucosinolates belonging to the three chemical

classes were identified in the leaves of nabicol

Euphytica (2008) 159:111–122 113

123

varieties: four aliphatic, three indolyl, and one

aromatic (Table 1) Three glucosinolates were de-

tected in all varieties: glucobrassicanapin, progoitrin,

and glucobrassicin. Aliphatic glucosinolates were

predominant, representing the 90% of total glucosin-

olate content. Glucobrassicanapin was the most

abundant (41% of total glucosinolate content in

location 1 and 54% in location 2) followed by

progoitrin and gluconapin (Fig. 1). Smaller amounts

of glucobrassicin, glucoalyssin, gluconasturtiin,

neoglucobrassicin, and 4-methoxyglucobrassicin

were also detected. Gluconapoleiferin and 4-hydrox-

yglucobrassicin were found in some varieties analy-

sed in location 2 although in low concentrations.

Despite differences on glucosinolate content between

locations, the glucosinolate pattern found at each

location was very similar. Previous studies have

reported a similar glucosinolate pattern in leaves of

fodder rapes (Rosa 1999) and vegetable types (Rosa

et al. 1996) and in seeds of oilseed rape (Fenwick

et al. 1983b, Li et al. 1999) and nabicol varieties

(De Haro et al. 1995).

The combined analysis of variance showed signif-

icant differences for total glucosinolate content

among locations, varieties, and the location · variety

interaction (Table 2). The average glucosinolate

content detected in leaves collected from location 1

was higher (17.1 mmol g�1 dw) than those collected

from location 2 (3.4 mmol g�1 dw) (Table 3).

Environmental factors, such as temperature, water

stress and soil type are known to exert a significant

effect on glucosinolate content (Fenwick et al. 1983b;

Rosa et al. 1996; Rosa et al. 1997). Soil differences

across locations could be the cause of the significant

differences between locations and location · variety

interaction for all glucosinolates. Differences in the

soil parameters were proved by edaphic analyses

showing that the soil pH was strongly acid in location

2. The highest glucosinolate content occurred in loca-

tion 1, with the highest soil pH, suggesting some type of

relation between glucosinolate content and soil effect.

Besides the influence of soil composition, Brassica

crops grown under cool temperatures and abundant

rainfall seem to have lower glucosinolate content (Rosa

et al. 1997). In location 2, the temperature was low

during the early development stages of growth and total

rainfall was more abundant than in location 1.

Differences between locations were found for

aliphatic, indolyl, and aromatic glucosinolates

(Table 2). In our case, and under stress conditions

for nabicol crop such as previously described in

location 2, the pathway for glucosinolate biosynthesis

Table 1 List of glucosinolates found in leaves of 36 varieties

of nabicol evaluated at two locations in northwestern Spain

Glucosinolate Common name Abbreviation

Aliphatic glucosinolates (derived from methionine)

4-Pentenyl Glucobrassicanapin GBN

2-Hydroxy-3-butenyl Progoitrin PRO

3-Butenyl Gluconapin GNA

5-Methylsulfinylpentyl Glucoalyssin GAL

Indolyl glucosinolates (derived from tryptophan)

Indol-3-ylmethyl Glucobrassicin GBS

1-Methoxyindol-3-

ylmethyl

Neoglucobrassicin NGBS

4-Methoxyindol-3-

ylmethyl

4-Methoxygluco

brassicin

MeGBS

Aromatic glucosinolates (derived from phenylalanine)

2-Phenylethyl Gluconasturtiin GST

0

10

20

30

40

50

60

GBN PRO GNA GAL GBS NGBS MeGBS GST

location 1 location 2Fig. 1 Percentage of

glucosinolates in leaves of

nabicol varieties sampled at

two locations. GBN:

Glucobrassicanapin; PRO:

Progoitrin, GNA:

Gluconapin, GAL:

Glucoalyssin, GBS:

Glucobrassicin, NGBS:

Neoglucobrassicin,

MeGBS: 4-

Methoxyglucobrassicin,

GST: Gluconasturtiin

114 Euphytica (2008) 159:111–122

123

could have been modified. The biosynthesis of the

individual glucosinolates in Brassica is under genetic

control by means of enzymes via different side chain

elongation. These enzymes could be modified

depending on specific environmental conditions,

increasing or decreasing some glucosinolates

(Giamoustaris and Mithen 1996).

The total glucosinolate content ranged from

1.4 mmol g�1 to 41.0 mmol g�1 dw in location 1

and from 1.2 mmol g�1 to 7.6 mmol g�1 dw in

location 2 (Table 3). These contents are lower than

those found in other B. napus crops (Carlson et al.

1981; Griffiths et al. 1991; Rosa et al. 1997).

Glucobrassicanapin, the main glucosinolate, showed

an average of 7.08 and 1.87 mmol g�1 dw in locations

1 and 2, respectively while the other glucosinolates

had values lower than 1 mmol g�1 dw in the last

location (Table 3).

Varieties were significantly different for the total

glucosinolate content and for all individual glucosin-

olates, except for gluconasturtiin and 4-methoxyg-

lucobrassicin at location 2 (Table 2). Due to

beneficial effects of isothiocyanates derived from

glucobrassicanapin and gluconapin on human health

(Mithen et al. 2003; Rose et al. 2005), the most

promising varieties for future breeding purposes

would be those with the highest total glucosinolate

content. The variety MBG-BRS0029 had the highest

total glucosinolate content (41 mmol g�1 dw) in

location 1 (Fig. 2A, Table 4) and the highest

p r o g o i t r i n a n d g l u c o n a p i n c o n t e n t s

(16.5 mmol g�1 dw and 8.8 mmol g�1 dw, respec-

tively) at this location (Table 4). Other varieties with

high total glucosinolate content at location 1 were

MBG-BRS0035 and MBG-BRS0374 (Fig. 2A). This

last variety also had the highest glucobrassicanapin

content (16.3 mmol g�1 dw). In location 2, varieties

showed a low glucosinolate content. The variety

MBG-BRS0329 had the highest total glucosinolate

content (7.6 mmol g�1 dw) (Fig. 2A, Table 3) and the

highest glucobrassicanapin and progoitrin contents

(4.1 mmol g�1 dw and 2.3 mmol g�1 dw, respectively)

(Table 4). Other varieties with high total glucosino-

late content at this location were MBG-BRS0035 and

MBG-BRS0337 (Fig. 2A). Regarding both locations

MBG-BRS0035 had the highest total glucosinolate

content and MBG-BRS0092 the lowest glucosinolate

content. The commercial variety MBG-BRS0373 had

Table 2 Mean squares of the analysis of variance within each site and mean squares of the combined analysis of variance for the

total glucosinolate content in the nabicol varieties evaluated at two locations in northwestern Spain

Traits Location 1 Location 2 Combined analysis

Variety Error Variety Error Location Variety Location · Variety

Total glucosinolate 548.8** 39.3 8.5** 3.1 14452.8** 243.0** 181.7**

Aliphatic

GBN 103.61** 11.92 2.44** 0.64 2206.32** 46.48** 33.87**

PRO 57.01** 5.51 0.66* 0.35 1067.61* 24.26** 19.58*

GNA 14.99** 2.06 0.24** 0.05 280.05** 5.81** 5.21*

GAL 7.37** 1.15 0.20* 0.04 53.47* 3.04** 2.27**

GNL – – 0.06** 0.02 – – –

Indolyl

GBS 3.09* 0.37 0.10* 0.05 30.62* 1.38** 1.35**

NGBS 0.24** 0.04 0.06** 0.01 1.68** 0.12* 0.17**

MeGBS 0.04* 0.01 & 0 & 0 1.08* 0.02** 0.02*

Aromatic

GST 0.30** 0.03 0.04 0.03 0.45** 0.15** 0.15**

Degrees of freedom 35 134 32 79 1 35 32

*, ** significant at P � 0.05 or 0.01, respectively

GBN: Glucobrassicanapin; PRO: Progoitrin; GNA: Gluconapin; GAL: Glucoalyssin; GNL: Gluconapoleiferin; GBS: Glucobrassicin;

NGBS: Neoglucobrassicin; MeGBS: 4-Methoxyglucobrassicin; GST: Gluconasturtiin

Euphytica (2008) 159:111–122 115

123

low total glucosinolate content in both locations.

Seeds of commercial variety came from Portugal

where they were bought as ‘couve-nabica’ but they

presumably correspond to a rapeseed variety.

Because of the adverse effects of glucosinolates on

animal nutrition, current commercial varieties of

oilseed rape have very low glucosinolate levels in

seeds, less than 30 mmol g�1. However, no recom-

mended values for glucosinolates have been de-

scribed in vegetable Brassica crops.

Varieties displayed the standard glucosinolate

profile, except for MBG-BRS0029, MBG-BRS0037,

MBG-BRS0041, MBG-BRS0048, MBG-BRS0061,

MBG-BRS0113, and MBG-BRS0356 in location 1,

which had relatively moderate progoitrin levels

(Fig. 2B, Table 4). High levels of progoitrin have

been implicated on goitrogenic effects in animals

causing thyroid hypertrophy (Rosa et al. 1997)

although no evidence on its possible involvement in

goitre have been demonstrated in humans (Mithen

et al. 2000). The use of varieties with high progoitrin

content is considered inconvenient for food purposes.

However, nabicol varieties included in this study

displayed low progoitrin contents.

Among the glucosinolates surveyed in Brassica

crops, of particular interest is glucoraphanin, which

has beneficial effects for human diet because of its

role as a cancer protecting agent (Rosa et al. 1997;

Farnham et al. 2004; Smith et al. 2005). The

glucoraphanin was absent in the collection evaluated

whereas progoitrin and gluconapin were abundant

which could offer future perspectives to modify the

glucosinolate composition because glucoraphanin,

progoitrin and gluconapin are in the same pathway

of the biosynthesis of the aliphatic glucosinolates

(Giamoustaris and Mithen 1996).

Biosynthesis of gluconapin requires a functional

allele at the Gsl-alk locus that converts glucoraphanin

to gluconapin. Down-regulation of Gsl-Alk could

produce Brassica varieties lacking the antinutrient

progoitrin and would simultaneously produce plants

accumulating glucoraphanin as a source of anticar-

cinogens. Li and Quiros (2003) obtained transformed

Arabidopsis plants with reduced concentration of

glucoraphanin, which was converted into gluconapin.

Nutritional and sensory traits

Regarding the nutritional value, crude protein and

acid detergent fibre are important parameters in

traditional farming systems, where residual post-

harvest leaves are used as feeding. Significant

differences (P � 0.01) were found between locations

for both traits. In location 1, varieties had more

protein (28.6% dw) and less ADF (16.7% dw) than in

location 2 (13% dw of protein and 19.6% dw of ADF)

(Table 5). Previous works agree with these results,

since high temperatures and low soil humidity

increase protein content and reduce fibre content in

Brassica fodder crops (Wiedenhoeft and Barton

1994; Rosa and Heaney 1996). Since the loca-

tion · variety interaction was not significant for

crude protein and ADF and varieties differed signif-

icantly for both traits, means for each variety are

shown across locations. The commercial nabicol

variety showed intermediate values for both ADF

and crude protein. The average crude protein found in

the nabicol leaves ranged from 16.5% to 25.5% dw

(Table 5) and this content was similar to the values

Table 3 Mean (mmol g�1 dw) and range for total and indi-

vidual glucosinolate content in leaves of 36 varieties of nabicol

evaluated at two locations in northwestern Spain

Location 1 Location 2 LSD (5%)

Mean Range Mean Range

t-GSL 17.1 1.4–41.0 3.4 1.23–7.55 1.22

Aliphatic

GBN 7.08 0.62–16.26 1.87 1.01–4.09 0.67

PRO 4.94 0.38–16.48 0.74 0.01–2.32 0.46

GNA 2.45 0.05–8.79 0.26 0–1.39 0.28

GAL 1.09 0–5.12 0.10 0–1.41 0.21

GNLa &0 – 0.08 0–0.67 –

Indolyl

GBS 0.94 0.26–1.09 0.23 0.03–0.98 0.12

NGBS 0.24 0.04–1.09 0.06 0–0.61 0.04

MeGBSa 0.14 0–0.38 & 0 – –

Aromatic

GST 0.23 0–0.86 0.16 0–0.36 0.04

t-GSL: total glucosinolate content, GBN: Glucobrassicanapin,

PRO: Progoitrin, GNA: Gluconapin, GAL: Glucoalyssin,

GNL: Gluconapoleiferin, GBS: Glucobrassicin, NGBS:

Neoglucobrassicin, MeGBS: 4-Methoxyglucobrassicin, and

GST: Gluconasturtiina Values were negligible for Gluconapoleiferin at location 1

and for 4-methoxyglucobrassicin at location 2

116 Euphytica (2008) 159:111–122

123

reported on other Brassica forage crops such as kales

(Rosa 1999).

Brassica crops are relatively low in fibre and are

readily digested, providing good concentrations of

energy for ruminants. The combined mean over

locations was 18.1% dw with a range between 14.9%

and 21.31% dw (Table 5). The average ADF content

was similar to the values found in different

B. oleracea crops (Rosa 1999) and it would be

consider appropriate for fodder use. Additionally,

dietary fibre and protein quality are some of the

arguments used to increase Brassica consumption. In

this context, values obtained in this work for ADF

and crude protein concentration would allow us to use

this crop for edible leaves either as food and/or feed.

For sensory traits, varieties grown in location 1

had a best acceptability by taste panel (best flavour,

less fibrousness, and less hardness) than those grown

in location 2. Average variety values were 4.1, 2.1,

and 2.5 in location 1 and 3.2, 3.6, and 3.1 in location

2 for flavour, hardness, and fibrousness, respectively.

Location 1 represents the environmental conditions

where crop is well adapted while location 2 is an

inland location where climate is more severe and

environmental conditions are not suitable for nabicol

crop. Bitterness was the only sensory trait for which

locations showed no significant differences (data not

shown), suggesting that this trait is more related to

other factors such as glucosinolate content than to the

climatic conditions (Van Doorn et al. 1998). Varieties

were significantly different for flavour (P < 0.05),

being MBG-BRS0056, MBG-BRS0041, MBG-

BRS0337, and MBG-BRS0333 the best varieties

with a value near to 4 in a scale from 1 to 5. The

A

0

5

10

15

20

25

30

35

40

45

BR

S00

14

BR

S00

28

BR

S00

29

BR

S00

34

BR

S00

35

BR

S00

37

BR

S00

39

BR

S00

41

BR

S00

44

BR

S00

48

BR

S00

54

BR

S00

56

BR

S00

61

BR

S00

63

BR

S00

65

BR

S00

68

BR

S00

73

BR

S00

79

BR

S00

85

BR

S00

87

BR

S00

90

BR

S00

92

BR

S01

05

BR

S01

07

BR

S01

10

BR

S01

13

BR

S01

31

BR

S01

34

BR

S03

29

BR

S03

33

BR

S03

37

BR

S03

46

BR

S03

56

BR

S03

73

BR

S03

74

BR

S03

78

0

1

2

3

4

5

1coL 2coL ruovalf ssenrettib

g/lomµ

B

0

2

4

6

8

10

12

14

16

18

BR

S00

14

BR

S00

28

BR

S00

29

BR

S00

34

BR

S00

35

BR

S00

37

BR

S00

39

BR

S00

41

BR

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44

BR

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48

BR

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54

BR

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61

BR

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63

BR

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65

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68

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73

BR

S00

79

BR

S00

85

BR

S00

87

BR

S00

90

BR

S00

92

BR

S01

05

BR

S01

07

BR

S01

10

BR

S01

13

BR

S01

31

BR

S01

34

BR

S03

29

BR

S03

33

BR

S03

37

BR

S03

46

BR

S03

56

BR

S03

73

BR

S03

74

BR

S03

78

0

1

2

3

4

5

g/lomµ

1coL 2coL ruovalf ssenrettib

Fig. 2 (A) Bitterness, flavour and total glucosinolate content

expressed as (mmol/g dw) in 36 varieties of Brassica napusevaluated by a consumer panel and sampled at both locations

(B) Bitterness, flavour and progoitrin content expressed as

(mmol/g dw) in 36 varieties of Brassica napus evaluated by a

consumer panel and sampled at both locations. Bitterness

expressed as rating scale from 1 = slight to 5 = high. Flavour

expressed as rating scale from 1 = very bad to 5 = very good.

Varieties MBG-BRS0056, MBG-BRS0061, and MBG-

BRS0378 were not sampled at location 2

Euphytica (2008) 159:111–122 117

123

Table 4 Total and individual leaf glucosinolate content (mmol g�1 dw) of 36 varieties of nabicol grown at two locations in

northwestern Spain

Variety (MBG-) loc t-GSL GBN PRO GNA GAL GBS GST NGBS MeGBS

BRS0014 1 20.26 7.88 5.39 3.16 2.27 0.94 0.46 0.18 0

2 1.48 1.06 0.14 0 0 0.18 0.10 0 0

BRS0028 1 16.28 8.71 3.83 2.62 0.34 0.35 0.28 0.15 0

2 3.84 1.75 1.02 0.19 0.08 0.43 0.30 0.07 0

BRS0029 1 40.99 11.92 16.48 8.79 1.91 1.17 0.23 0.42 0.07

2 2.04 1.24 0.30 0.08 0 0.16 0.13 0.12 0.01

BRS0034 1 7.83 3.49 2.70 1.26 0.09 0.19 0 0.04 0.06

2 2.99 1.29 0.52 0 0 0.28 0.29 0.61 0

BRS0035 1 34.55 16.13 11.23 4.78 0.36 1.69 0 0.20 0.20

2 6.18 3.71 1.20 0.67 0 0.13 0.20 0.09 0

BRS0037 1 8.04 2.12 3.68 1.49 0.34 0.30 0 0.04 0.07

2 5.84 2.97 1.83 0.77 0 0.05 0.17 0.05 0

BRS0039 1 22.58 8.74 6.69 3.57 1.77 1.29 0 0.31 0.21

2 2.59 1.46 0.56 0.10 0 0.16 0.22 0.08 0.01

BRS0041 1 4.56 1.27 1.93 0.85 0 0.25 0.08 0.07 0.10

2 3.63 2.03 0.48 0.09 0.24 0.44 0.32 0.01 0.01

BRS0044 1 22.61 8.63 5.95 4.01 1.75 1.63 0 0.37 0.26

2 3.28 2.00 0.52 0.31 0.08 0.37 0.11 0 0

BRS0048 1 11.37 4.40 4.45 1.80 0.00 0.15 0 0.45 0.12

2 3.29 1.50 1.30 0.09 0.07 0.20 0.22 0 0

BRS0054 1 26.72 9.41 6.26 4.12 5.12 1.09 0.14 0.41 0.16

2 1.23 1.01 0.01 0 0 0.06 0.15 0 0

BRS0056 1 4.02 1.92 0.80 0.96 0 0.15 0.05 0.08 0.08

2 -a - - - - - - - -

BRS0061 1 3.21 0.83 1.78 0.05 0.03 0.18 0.05 0.06 0.23

2 -a - - - - - - - -

BRS0063 1 24.05 9.69 5.89 3.39 2.74 1.80 0.09 0.32 0.12

2 2.97 1.95 0.60 0.21 0 0.08 0.11 0.02 0

BRS0065 1 4.62 2.11 1.29 0.91 0 0.14 0 0.10 0.08

2 3.25 2.26 0.50 0.25 0 0.08 0.11 0.05 0

BRS0068 1 18.00 9.81 4.64 2.66 0.09 0.56 0 0.13 0.11

2 1.59 1.15 0.19 0 0 0.04 0.17 0.05 0

BRS0073 1 7.53 2.84 2.56 1.56 0.25 0.20 0 0.06 0.06

2 1.51 1.28 0.11 0.07 0 0.04 0 0.01 0

BRS0079 1 7.42 6.62 0.38 0.16 0 0.13 0 0.04 0.09

2 4.15 1.95 1.23 0.17 0 0.61 0.16 0.03 0

BRS0085 1 11.99 5.12 3.54 1.48 1.07 0.53 0.02 0.11 0.13

2 2.59 1.17 0.32 0 0 0.29 0.36 0.45 0.01

BRS0087 1 20.00 9.85 5.49 2.81 0.69 0.56 0.40 0.10 0.10

2 2.75 1.78 0.28 0.25 0 0.19 0.23 0.03 0

BRS0090 1 15.55 8.07 3.43 2.58 0.11 0.33 0.73 0.20 0.10

2 4.26 2.43 1.15 0.53 0 0.07 0.07 0 0.01

118 Euphytica (2008) 159:111–122

123

location · variety interaction was significant

(P < 0.01) for bitterness and fibrousness. Accessions

were significantly different for these traits in the

location 1, with MBG-BRS0028, MBG-BRS0037,

MBG-BRS0374, and MBG-BRS0356 significantly

less bitter.

Bitter taste foods are frequently disliked and this is

one reason for the low acceptability of some Brassica

crops. Most nabicol varieties showed less bitterness

(less than mean value of 3.0 in a scale from 1 to 5)

than other B. brassica crops as Brussels sprouts, turnip

greens or turnip tops (Padilla et al. 2007) along with a

Table 4 continued

Variety (MBG-) loc t-GSL GBN PRO GNA GAL GBS GST NGBS MeGBS

BRS0092 1 3.36 1.37 1.10 0.41 0 0.20 0.13 0.08 0.08

2 1.59 1.21 0.20 0.04 0 0.07 0.07 0 0

BRS0105 1 28.48 13.70 7.70 2.79 1.25 2.35 0.58 0.05 0.06

2 3.84 2.28 0.62 0.21 0 0.23 0.35 0.15 0

BRS0107 1 26.81 14.75 5.55 3.87 0.97 0.92 0.36 0.28 0.12

2 3.22 1.69 1.10 0.16 0 0.03 0.21 0.02 0

BRS0110 1 26.88 9.03 7.97 4.16 3.04 1.60 0.60 0.10 0.38

2 3.47 1.59 1.20 0.13 0 0.29 0.17 0.08 0.01

BRS0113 1 10.66 2.91 4.07 1.68 1.15 0.45 0.20 0.14 0.07

2 3.23 1.18 0.95 0.09 0.39 0.37 0.18 0.04 0

BRS0131 1 22.80 7.60 6.89 3.00 0.95 3.26 0.61 0.22 0.27

2 1.67 1.19 0.38 0.07 0 0.03 0 0 0

BRS0134 1 25.90 8.76 8.31 4.49 1.85 1.62 0.43 0.14 0.30

2 2.52 1.76 0.42 0.30 0 0.06 0 0.02 0

BRS0329 1 24.33 9.45 6.67 1.92 1.88 2.18 0.86 1.09 0.32

2 7.55 4.09 2.32 0.55 0 0.24 0.29 0.06 0

BRS0333 1 2.21 0.62 0.54 0.20 0 0.29 0.13 0.22 0.21

2 3.78 1.98 0.26 1.40 0 0.04 0.10 0 0

BRS0337 1 6.49 2.17 1.44 0.63 0.11 0.80 0.27 1.00 0.08

2 6.24 3.26 1.14 0.80 0.71 0.35 0 0 0

BRS0346 1 26.31 12.34 6.69 2.32 2.86 1.13 0.63 0.15 0.19

2 5.91 2.12 1.09 0.35 1.41 0.77 0.19 0 0

BRS0356 1 22.08 6.40 9.75 1.84 0.54 2.81 0.45 0.14 0.15

2 4.79 2.58 1.00 0.45 0.45 0.19 0.13 0 0

BRS0373b 1 4.16 2.40 0.62 0.40 0 0.26 0.03 0.24 0.21

2 1.92 1.36 0.37 0 0 0.12 0.06 0.01 0

BRS0374 1 32.34 16.26 7.31 4.11 2.41 1.33 0.56 0.24 0.12

2 3.89 1.31 1.20 0.25 0 0.98 0.13 0.02 0

BRS0378 1 19.97 7.54 4.79 3.31 2.98 0.83 0.19 0.21 0.12

2 -a - - - - - - -

LSD (5%) loc 1 8.14 4.49 3.05 1.87 1.39 0.79 0.21 0.25 0.12

LSD (5%) loc 2 3.12 1.42 1.04 0.42 0.31 0.38 - 0.16 -

t-GSL: total glucosinolate content, PRO: Progoitrin, GAL: Glucoalyssin, GNA: Gluconapin, GBN: Glucobrassicanapin, GBS:

Glucobrassicin, GST: gluconasturtiin, NGBS: Neoglucobrassicin, MeGBS: 4-Methoxyglucobrassicina Varieties no evaluated in location 2b Commercial nabicol variety

Euphytica (2008) 159:111–122 119

123

good flavour. Isothiocyanate compounds derived

from sinigrin, glucobrassicanapin, and gluconapin

contribute to the bitter flavour of cruciferous

vegetables while the bitterness associated with pro-

goitrin is due to its decomposition product, the 5-

vinyl-oxazolidine-2-thione (OZT) (Fenwick et al.

1983a). Descriptive sensory analysis comparing fla-

vour attributes with variation in glucosinolate con-

centration have been performed in turnip greens

(Padilla et al. 2007), broccoli and cauliflower

(Schonhof et al. 2004), and in Brussels sprouts

(Van Doorn et al. 1998) but not in nabicol varieties.

The content of sinigrin and progoitrin was positively

correlated with bitterness and negatively correlated

with taste preference in Brussels sprouts (Van Doorn

et al. 1998).

The relation between flavour and bitterness with

the total glucosinolate and progoitrin contents are

shown in Figs. 2A and B, respectively. Total

glucosinolate content and progoitrin content appears

to be related to flavour. MBG-BRS0029 with the

highest glucosinolate and progoitrin contents in

location 1 had a bad flavour in a scale from 1 to 5

whereas MBG-BRS0333 with the lowest levels of

glucosinolates in location 1 had a good flavour.

However, no relation was found between bitterness

and the levels of glucosinolates and progoitrin;

varieties with high glucosinolate and progoitrin

contents were as bitter as varieties with low gluco-

sinolate and progoitrin contents (Figs. 2A and 2B).

Because gluconapin, glucobrassicanapin, and pro-

goitrin were the major glucosinolates found in this

work, simple correlation coefficients between these

three glucosinolates with flavour and bitterness were

calculated (Table 6). Flavour was related to high

glucosinolate content, but no to high gluconapin,

glucobrassicanapin or progoitrin contents. Bitterness

was not related to high glucosinolate contents,

suggesting that these compounds and their break-

down products are not the primary determinants of

the bitter taste and aroma of this vegetable and

Table 5 Means and standard deviation for acid detergent fibre

(ADF) and crude protein of 36 varieties of nabicol grown at

two locations in northwestern Spain

Varieties ADF Crude protein

(% of dry weight)

MBG-BRS0014 19.50 ± 1.85 20.41 ± 9.61

MBG-BRS0028 17.56 ± 1.90 24.73 ± 8.58

MBG-BRS0029 15.97 ± 2.60 20.75 ± 15.34

MBG-BRS0034 19.01 ± 3.56 20.94 ± 12.25

MBG-BRS0035 16.43 ± 2.86 22.98 ± 15.59

MBG-BRS0037 19.22 ± 2.57 21.54 ± 10.97

MBG-BRS0039 16.97 ± 0.61 23.02 ± 12.13

MBG-BRS0041 18.57 ± 3.92 20.40 ± 8.06

MBG-BRS0044 18.69 ± 2.24 24.22 ± 9.74

MBG-BRS0048 21.28 ± 0.50 18.24 ± 8.58

MBG-BRS0054 16.01 ± 1.70 18.18 ± 13.47

MBG-BRS0056 17.47 ± 0.75 21.12 ± 7.89

MBG-BRS0061 16.89 ± 2.10 19.19 ± 9.77

MBG-BRS0063 16.08 ± 2.01 18.91 ± 14.56

MBG-BRS0065 18.12 ± 1.72 18.22 ± 12.13

MBG-BRS0068 16.84 ± 2.78 21.99 ± 12.88

MBG-BRS0073 17.19 ± 2.53 20.50 ± 8.07

MBG-BRS0079 16.97 ± 3.63 17.28 ± 9.08

MBG-BRS0085 16.49 ± 0.86 22.11 ± 13.42

MBG-BRS0087 14.94 ± 0.76 19.80 ± 12.45

MBG-BRS0090 19.33 ± 4.43 20.77 ± 9.09

MBG-BRS0092 17.95 ± 0.79 20.74 ± 13.10

MBG-BRS0105 19.52 ± 0.87 19.78 ± 12.91

MBG-BRS0107 17.12 ± 4.13 21.81 ± 14.13

MBG-BRS0110 16.95 ± 0.92 21.44 ± 14.66

MBG-BRS0113 16.81 ± 0.30 20.29 ± 10.91

MBG-BRS0131 16.57 ± 0.52 22.03 ± 15.24

MBG-BRS0134 16.99 ± 0.01 25.53 ± 7.74

MBG-BRS0329 17.18 ± 0.03 17.92 ± 14.83

MBG-BRS0333 16.98 ± 2.23 21.11 ± 8.20

MBG-BRS0337 17.10 ± 0.70 18.39 ± 9.35

MBG-BRS0346 17.99 ± 0.16 21.35 ± 13.22

MBG-BRS0356 15.41 ± 1.85 23.63 ± 3.35

MBG-BRS0373 16.21 ± 1.42 20.29 ± 2.56

MBG-BRS0374 17.83 ± 0.24 22.14 ± 11.40

MBG-BRS0378 17.16 ± 2.07 16.53 ± 10.14

LSD (5%) 4.025 6.382

Table 6 Simple correlation coefficients among total and some

specific glucosinolates and two quality traits on 36 varieties of

nabicol grown in two locations in northwestern Spain

t-GSL PRO GBN GNA

Flavour 0.47 ** 0.35 * 0.36 * 0.21

Bitterness 0.11 0.13 0.10 0.13

*, ** significant at P � 0.05 or 0.01, respectively

t-GSL = Total glucosinolate content; PRO = Progoitrin;

GBN = Glucobrassicanapin; GNA = Gluconapin

120 Euphytica (2008) 159:111–122

123

therefore, other glucosinolates or other phytochemi-

cals are involved in the characteristic bitterness of

this Brassica species. Because both low palatability

and bitterness are not such critical in nabicol crops

compared with other Brassica vegetables, it is

probably difficult to detect relations between bitter-

ness and high glucosinolate contents.

Although nabicol varieties look quite uniform on

the basis of molecular data (Cartea et al. 2005;

Soengas et al. 2006) and agronomical characteristics

(Rodrıguez et al. 2005), variation on glucosinolate

levels have been observed. With the application of

appropriate breeding procedures, new nabicol varie-

ties of acceptable agronomic type could be developed

with higher glucosinolate contents than those found

in varieties currently grown as food. The effect on

other plant characters such as disease and pest

resistance should be monitored to ensure that they

are not adversely affected because leaf glucosinolate

content can influence feeding behaviour of insect pest

(Giamoustaris and Mithen 1996). Because the attack

by insect pests to Brassica crops is important in

northwestern Spain (Picoaga et al. 2003), it would be

interesting to manipulate the glucosinolate levels to

improve pest and disease resistance. The interaction

between plant glucosinolates and disease and pest

resistance is complex and need considerable further

study.

As conclusion, nabicol varieties from northwestern

Spain differed greatly in glucosinolate contents,

ADF, crude protein and flavour which enable us to

select some interesting varieties to obtain improved

nabicol varieties with different glucosinolate content

that could be used for different purposes. The

glucosinolate composition was significantly influ-

enced by the environment although the glucosinolate

pattern did not vary. The variety MBG-BRS0035 had

high total glucosinolate content at both locations and

could be included in breeding programs to improve

the nutritional value of this vegetable crop. This is the

first study about the nutritional value of nabicol crops

and it provides an interesting and valuable material

for further breeding programs.

Acknowledgements We thank G. Fernandez, E. Santiago

and R. Abilleira for work laboratory. This work has been

supported by the Projects AGL 2003-01366 and AGL 2006-

04055 of the Spanish Government and Excma. Diputacion

Provincial De Pontevedra. V.M. Rodrıguez acknowledges a

fellowship from the Ministry of Science and Technology from

Spain.

References

AOAC (2000) Association of official analytical chemists.

Official Methods of Analysis. AOAC International 17th

edition, Arlington, VA

Carlson DG, Daxenbichler ME, Van Etten CH, Tookey HL,

Williams PH (1981) Glucosinolates in crucifer vegetables:

turnips and rutabagas. J Agric Food Chem 29:1235–1239

Cartea ME, Soengas P, Picoaga A, Ordas A (2005) Relation-

ships among Brassica napus (L.) germplasm from Spain

and Great Britain as determined by RAPD markers. Genet

Resour Crop Evol 52:655–662

De Haro A, Fernandez G, Baladron JJ, Ordas A (1995) Estudio

de la variabilidad respecto a componentes nutritivos en

brassicas gallegas. Proceedings of VI Congreso de la

Sociedad Espanola de Ciencias Hortıcolas, Barcelona,

Spain, p. 123

Farnham MW, Wilson PE, Stephenson KK, Fahey JW (2004)

Genetic and environmental effects on glucosinolate con-

tent and chemoprotective potency of broccoli. Plant Breed

123:60–65

Fenwick RG, Griffiths NM, Heaney RK (1983a) Bitterness in

Brussels sprouts (Brassica oleracea L. var. gemmifera):

the role of glucosinolates and their breakdown products.

J Sci Food Agric 34:73–80

Fenwick RG, Heaney RK, Mullin WJ (1983b) Glucosinolates

and their breakdown products in food plants. CRC Crit

Rev Food Sci Nutr 18:123–201

Font R, del Rıo M, Cartea ME, de Haro A (2005) Application

of Near-Infrared Spectroscopy to the analysis of total and

individual glucosinolates in Brassica napus leaves. Phy-

tochemistry 66:175–185

Giamoustaris A, Mithen R (1996) Genetic of aliphatic gluco-

sinolates. Side-chain modification in Brassica oleracea.

Theor Appl Genet 93:1006–1010

Griffiths DH, Bradshaw JE, Taylor J, Gemmell DJ (1991) Ef-

fect of cultivar and harvest date on the glucosinolates and

S-methylcystein sulphoxide content on sweedes (Brassicanapus spp. rapifera). J Sci Food Agric 56:539–549

ISO Norm (1992) Rapeseed–determination of glucosinolates-

part 1: method using high-performance liquid chromato-

graphy. ISO 9167-1, pp 1–9

Johnson HW, Robinson HF, Comstock RE (1955) Genotypic

and phenotypic correlations in soybeans and their impli-

cations in selection. Agron J 47:477–483

Jones G, Sanders OG (2002) A sensory profile of turnip greens as

affected by variety and maturity. J Food Sci 67:3126–3129

Li G, Quiros CF (2003) In planta side-chain glucosinolate

modification in Arabidopsis by introduction of dioxy-

genase Brassica homolog BoGSL-ALK. Theor Appl

Genet 106:1116–1121

Li Y, Kiddle G, Bennet R, Doughty K, Wallsgrove R (1999)

Variation in the glucosinolate content of vegetative tissues

of Chinese lines of Brassica napus L. Ann Appl Biol

134:131–136

Euphytica (2008) 159:111–122 121

123

Mithen R, Faulkner K, Magrath R, Rose P, Williamson G,

Marquez J (2003) Development of isothiocyanate-en-

riched broccoli and its enhanced ability to induce phase 2

detoxification enzymes in mammalian cells. Theor Appl

Genet 106:727–734

Mithen RF, Dekker M, Verkerk R, Rabot S, Johnson IT (2000)

The nutritional significance, biosynthesis and bioavail-

ability of glucosinolates in human foods. J Sci Food Agric

80:967–984

Padilla G, Cartea ME, Velasco P, de Haw A, Ordas A (2007)

Variation of glucosinolates in vegetable crops of Brassicarapa. Phytochemistry 68:536–545

Picoaga A, Cartea ME, Soengas P, Monetti L, Ordas A (2003)

Resistance of kale populations to Lepidopterous pests in

northwestern Spain. J Econ Entomol 96:143–147

Rodrıguez VM, Cartea ME, Padilla G, Velasco P, Ordas A

(2005) The nabicol: a horticultural crop in northwestern

Spain. Euphytica 142:237–246

Rosa EAS (1999) Chemical composition. In: Gomez-Campo C

(ed) Biology of Brassica coenospecies. Elsevier Science

BV, Amsterdam, pp 315–357

Rosa EAS, Heaney RK (1996) Seasonal variation in pro-

tein, mineral, and glucosinolates composition of Por-

tuguese Brassica crops. Anim Feed Sci Technol 57:

111–127

Rosa EAS, Heaney RK, Fenwick GR, Portas CAM (1997)

Glucosinolates in crop plants. Hort Rev 19:99–215

Rosa EAS, Heaney RK, Portas CAM, Fenwick GR (1996)

Changes in glucosinolate concentrations in Brassica crops

(B. oleracea and B. napus) throughout growing seasons. J

Sci Food Agric 71:237–244

Rose P, Huang Q, Ong CN, Whiteman M (2005) Broccoli and

watercress suppress matrix metalloproteinase-9 activity

and invasiveness of human MDA-MB-231 breast cancer

cells. Toxicol Appl Pharmacol 209:105–113

SAS Institute Inc. (2000) SAS OnlineDoc, version 8. SAS

Institute. Inc., Cary, North Carolina, USA

Schonhof I, Krumbein A, Bruckner B (2004) Genotypic effects

on glucosinolates and sensory properties of broccoli and

cauliflower. Nahrung/Food 48:25–33

Smith TK, Lund EK, Clarke RG, Bennet RN, Johnson IT

(2005) Effects of Brussels sprout juice on the cell cycle

and adhesion of human colorectal carcinoma cells (HT29)

in vitro. J Agric Food Chem 53:3895–3901

Soengas P, Velasco P, Padilla G, Ordas A, Cartea ME (2006)

Genetic relationships among Brassica napus crops based

on SSRs markers. HortScience 41:1195–1199

Steel RDG, Torrie JH, Dickey DA (1997) Principles and

procedures in statistics: a biometrical approach, 3rd ed.

Mc Graw Hill, New York, USA

Van Doorn HE, Van der Kruk GC, Van Holst GJ, Raaijmakers-

Ruijs NC, Potsma E, Groeneweg B, Jongen WHF (1998)

The glucosinolates sinigrin and progoitrin are important

determinants for taste preference and bitterness of Brus-

sels sprouts. J Sci Food Agric 78:30–38

Wiedenhoeft MH, Barton BA (1994) Management and envi-

ronmental effects on Brassica forage quality. Agron J

86:227–232

122 Euphytica (2008) 159:111–122

123