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Kale: An excellent source of vitamin C, pro-vitamin A, lutein and glucosinolates


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Kale (Brassica oleracea Acephala group) is a green leafy vegetable with high content of nutraceuticals. However, the content of bioactive compounds of kale is affected by the cultivar, and thus it is necessary to identify kale cultivars that have the highest nutritional content, to promote their consumption. The present project’s objective was to characterize the bioactive compounds (phenolic compounds, vitamin C, glucosinolates and individual carotenoids) and the antioxidant capacity of two kale cultivars (Winterbor and Maribor). The Winterbor cultivar presented higher content of phenolic compounds, carotenoids and higher antioxidant capacity than the Maribor, whereas the Maribor cultivar showed higher levels of vitamin C and glucosinolates. One serving size of kale provides more than 100% of the recommended daily intake (RDI) of vitamin A and more than 40% of the RDI of vitamin C. Therefore, kale can be considered an excellent source of antioxidants.
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CyTA - Journal of Food
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Kale: An excellent source of vitamin C, pro-vitamin
A, lutein and glucosinolates
Alejandro Becerra-Moreno, Pedro A. Alanís-Garza, José Luis Mora-Nieves,
Juan Pablo Mora-Mora & Daniel A. Jacobo-Velázquez
To cite this article: Alejandro Becerra-Moreno, Pedro A. Alanís-Garza, José Luis Mora-Nieves,
Juan Pablo Mora-Mora & Daniel A. Jacobo-Velázquez (2014) Kale: An excellent source of
vitamin C, pro-vitamin A, lutein and glucosinolates, CyTA - Journal of Food, 12:3, 298-303, DOI:
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© 2013 Taylor & Francis
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Kale: An excellent source of vitamin C, pro-vitamin A, lutein and glucosinolates
Kale: Una fuente excelente de vitamina C, pro-vitamina A, luteína y glucosinolatos
Alejandro Becerra-Moreno
, Pedro A. Alanís-Garza
, José Luis Mora-Nieves
, Juan Pablo Mora-Mora
Daniel A. Jacobo-Velázquez
Centro de Biotecnología FEMSA, Department of Biotechnology and Food Engineering, School of Biotechnology and Food,
Tecnológico de Monterrey Campus Monterrey, E. Garza Sada 2501 Sur, C.P. 64849, Monterrey, NL, México;
Frigorizados La Huerta,
S.A. de C.V. Rancho Medio Kilo. Calle 1 número 140, Colonia Medio Kilo, C.P. 20350, San Francisco de los Romo, Aguascalientes,
(Received 8 August 2013; nal version received 29 September 2013)
Kale (Brassica oleracea Acephala group) is a green leafy vegetable with high content of nutraceuticals. However, the content of bioactive
compounds of kale is affected by the cultivar, and thus it is necessary to identify kale cultivars that have the highest nutritional content, to
promote their consumption. The present projects objective was to characterize the bioactive compounds (phenolic compounds, vitamin C,
glucosinolates and individual carotenoids) and the antioxidant capacity of two kale cultivars (Winterbor and Maribor). The Winterbor
cultivar presented higher content of phenolic compounds, carotenoids and higher antioxidant capacity than the Maribor, whereas the
Maribor cultivar showed higher levels of vitamin C and glucosinolates. One serving size of kale provides more than 100% of the
recommended daily intake (RDI) of vitamin A and more than 40% of the RDI of vitamin C. Therefore, kale can be considered an excellent
source of antioxidants.
Keywords: kale; Winterbor and Maribor cultivars; antioxidants; phenolic compounds; glucosinolates; lutein; pro-vitamin A, vitamin C
El kale (Brassica oleracea Acephala group) es un vegetal de hoja verde con alto contenido de nutracéuticos. Sin embargo, el contenido de
compuestos bioactivos del kale se ve afectado por el cultivar, por lo tanto es necesario determinar cuál cultivar tiene el más alto contenido
nutricional para promover su consumo. El objetivo del presente proyecto fue caracterizar los compuestos bioactivos (compuestos fenólicos,
vitamina C, glucosinolatos, y carotenoides individuales) y la capacidad antioxidante de dos cultivares de kale (Winterbor and Maribor). El
cultivar Winterbor presentó mayor contenido de compuestos fenólicos, carotenoides y mayor capacidad antioxidante que el Maribor,
mientras que el cultivar Maribor presentó mayor cotenido de vitamina C y glucosinolatos. Un tamaño de porción de kale brinda 100% de la
ingesta diaria recomendada (IDR) de vitamina A y más del 40% de la IDR de vitamina C. Por lo tanto, el kale puede ser considerado una
excelente fuente de antioxidantes.
Palabras claves: kale; cultivares Winterbor y Maribor; antioxidantes; compuestos fenólicos; glucosinolatos; luteína; pro-vitamina A,
vitamina C
Kale (Brassica oleracea Acephala group) is a leafy green vege-
table belonging to the Brassicaceae family. In recent years, kale
has gained the attention of the scientic community due to its
high content of bioactive compounds such as vitamin C, pro-
vitamin A, glucosinolates, phenolic antioxidants, dietary ber,
micronutrients (iron, zinc and manganese) and macronutrients
(calcium and magnesium) (Ayaz et al., 2006; Cartea, Velasco,
Obregón, Padilla, & de Haro, 2008; Khachik, Beecher, & Goli,
1991; Olsen, Aaby, & Borge, 2009). Likewise, in vitro and in
vivo studies suggest that kale have a positive impact on the
prevention of chronic diseases such as cardiovascular diseases
(Kahlon, Chapman, & Smith, 2007; Kim, Yoon, Kwon, Park, &
Lee-Kim, 2008; Kural, Küçük, Yücesan, & Örem, 2011) and
cancer (Chung, Lee, & Sung, 2002).
The extensive growth and commercialization of kale can be
a realistic approach to increase the dietary intake of antioxidants
in the population. However, research is needed to determine
variations in the content of bioactive compounds amoung kale
cultivars, in order to promote the production of those with the
highest nutraceutical content. Therefore, the present projects
objective was to evaluate the concentration of phenolic com-
pounds (PC), vitamin C, glucosinolates, individual carotenoids
and the antioxidant capacity (ORAC value) of two kale cultivars
(Winterbor and Maribor).
Materials and methods
Plant material and chemicals
Kale cultivars (Winterbor and Maribor) were produced by
Frigorizados La Huerta, S.A de C.V. (San Francisco de los
Romo, Aguascalientes, Mexico). Methanol (HPLC grade), tert-
Butyl methyl ether (tBME; HPLC grade), acetone (HPLC grade),
isopropyl alcohol (HPLC grade), perchloric acid (HClO
) and
phosphoric acid (H
) were purchased from Desarrollo de
Especialidades Químicas, S.A. de C.V. (San Nicolás de los
Garza, Nuevo León, México). Gallic acid (GA), L-ascorbic
acid (AA), 6-hydroxy-2,5,7,8-tetramethyl-2-carboxylic acid
(Trolox), trichloroacetic acid (TCA), triuoroacetic acid (TFA),
HEPES, thioglucosidase from Sinapis alba, sinigrin hydrate,
glucose oxidase/peroxidase (GOP) assay kit, DL-dithiothreitol
(DTT), N-ethylmaleimide (NEM), 2, 2´-bipyridyl and iron (III)
*Corresponding author. Email:
CyTA Journal of Food, 2014
Vol. 12, No. 3, 298303,
© 2013 Taylor & Francis
chloride (FeCl
) were obtained from Sigma-Aldrich Co. (St.
Louis, MO, USA). Butylated hydroxytoluene (BHT) was
obtained from Spectrum Quality Products (New Brunswick,
Extraction and quantication of total phenolic
compounds and determination of total antioxidant
Kale tissue (5 g) was homogenized with methanol (20 mL) using
a tissuemizer (Advanced homogenizing system, VWR, Radnor,
PA, USA). Subsequently, the homogenates were stored overnight
(~12 h at 4ºC) and centrifuged (10,000×g, 15 min, 4°C). The
clear supernatant (methanol extract) was used for the analyses of
total PC and antioxidant capacity.
The total PC was determined with the method described by
Singleton and Rossi (1965), adapted to 96-well micro-plate
format (Jacobo-Velázquez, Martínez-Hernández, Rodríguez,
Cao, & Cisneros-Zevallos, 2011). The concentration of total
PC was expressed as mg of GA equivalents per kg of kale
fresh weight (FW).
The antioxidant capacity of the methanol extract was
determined with the oxygen radical absorbance capacity
(ORAC) assay. The ORAC value was obtained using the
procedure described by Wu et al. (2004) for hydrophilic
ORAC. Results were expressed as mg of trolox equivalents
Extraction and quantication of total glucosinolates
The extraction and quantication of total glucosinolates was
performed as described by Miranda Rossetto, Shiga, Vianello,
and Pereira Lima (2013) with slight changes. Briey, kal e
tissue (3 g of mature leaves) was homogenized with 70%
methanol (5 mL) and TFA (1 mL) using a tissuemizer
(advanced homogenizing system, VWR, Radnor, PA, USA).
Subsequently, the homogenates were stored in an incubator
(30 min, 70ºC, 150 rpm) and centrifuged (10,000×g,30min,
4°C). The clear supernatant was microltered using nylon
membranes (0.45 μm, VWR, Radnor, PA, USA). Thereafter,
the extracts were evaporated to dryness using a continuous ow
of nitrogen. Finally, the extract was resuspended in HEPES-
KOH (1 mL, 0.2 M, pH 7). The re-suspended extract (further
referred as re-constituted extract) was used for the quantica-
tion of total glucosinolates.
For total glucosinolates determinations, an enzymatic assay
was performed. The method consisted of the enzymatic hydro-
lysis of glucosinolates by a thioglucosidase from Sinapis alba,
and the subsequent quantication of glucose by the GOP assay
kit. Briey, the re-constituted extracts (50 µL) were incubated
(37°C, 24 h) with thioglucosidase solution (25 µL, 0.12 U,
HEPES-KOH 0.2 M, pH 7). The enzymatic reaction was stopped
by the addition of HClO
(25 µL, 18 mM). Total glucosinolates
content was calculated according to stoichiometry, which states
that 1 mol of released glucose is equivalent to 1 mol of gluco-
sinolate. As blank, the levels of glucose in the re-constituted
extract non-treated with thioglucosidase were determined.
Finally, the glucose released by thioglucosidase was determined
with the GOP assay kit according to manufacturers instructions.
Sinigrin hydrate was used as a positive control. Results were
expressed as µmol equivalents of glucosinolates per kg of
kale FW.
Extraction and quantication of vitamin C
To extract vitamin C from kale, the tissue (2 g) was rst homo-
genized under liquid nitrogen using a chilled mortar and pestle.
Thereafter, the tissue was homogenized with TCA (6%, 15 mL),
transferred to a 50 mL tube, centrifuged (15,000×g, 25 min,
4 ºC) and maintained on ice until needed for the assay (approxi-
mately 1 h). Total vitamin C content was determined in the
supernatant by using the 2, 2´-bipyridyl method (Gillespie &
Ainsworth, 2007; Okamura, 1980). Briey, the extract (100 μL)
was placed in a 2-mL tube and mixed with DTT solution
(20 mM, 100 μL). The mixture was incubated for 10 min at
room temperature in the dark. Then, NEM solution (0.5%, 100
μL) was added to the mixture and incubated for 30 s.
Subsequently, TCA (10%, 500 μL), H
(43%, 400 μL), 2,
2-bipyridyl (4%, 400 μL) and FeCl
(3%, 200 μL) solutions
were added to the assay tubes. The assay tubes were incubated at
37 ºC for 1 h. Then, 200 μL of the reaction solutions from the
assay tubes were placed in a well of a clear 96-well microplate
and absorbance readings were collected at 525 nm. Absorbance
values were compared against an AA standard curve (0.15
10 mM) prepared in TCA (6%). Results were expressed as mg
of vitamin C per kg of kale FW.
Extraction, identication and quantication of kale
carotenoids by HPLC-PDA
The extraction of carotenoids from kale tissue was performed as
described by Jacobo-Velázquez and Hernández-Brenes (2012)
with slight modications. Briey, kale tissue (1 g) was homo-
genized with 0.1% BHT acetone solution (10 mL) using a
tissuemizer. Subsequently, the homogenates were ltered under
vacuum through Whatman No. 1 lter papers (Piscataway, NJ,
USA) to obtain the acetone extracts. This procedure was
repeated twice to ensure the complete extraction of carotenoids.
The acetone extracts were pooled and concentrated in a rotary
evaporator (BÜCHI Labortechnik, AG, Flawil, Switzerland)
operating at 35°C, 60 rpm and 250 mbar until acetone was
completely evaporated. The remaining water in the ask was
removed by evaporating the extracts to dryness using a contin-
uous ow of nitrogen. The dried samples were re-dissolved in
isopropyl alcohol (1 mL) and ltered through nylon membranes
(0.45 μm) prior to injection to the HPLC system.
Carotenoids were identied and quantied by HPLC with
photodiode array detection (PDA). The HPLC system used was
composed of two 515 binary pumps, a 717-plus autosampler and
a 996-PDA (Waters Corp, Mildford, MA, USA). Carotenoids
were separated on a 4.6 mm × 150 mm, 3 μm, C30 reverse
phase column (YMC Carotenoid, Waters Corp, Mildford, MA,
USA). The mobile phases were methanol/water (96:4, v/v, phase
A) and tBME (phase B). The gradient solvent system was 0/95,
10/90, 40/55, 45/25, 50/0, 55/0 and 57/95 (min/% phase A) at a
constant ow rate of 0.75 mL/min. Chromatographic data was
processed with the Millenium software V3.1 (Waters Corp,
Mildford, MA, USA).
The tentative identication of each chromatographic peak
was achieved by comparing their retention time and UV/
Visible (UV/Vis) absorption spectra characteristics with those
of commercial carotenoid standards. In addition, the order of
elution and the UV/Vis spectra characteristic from carotenoids
reported in previous studies were used as additional parameters
of identication. For the quantication of individual carotenoids,
standard curves of lutein and all-trans-β-carotene were prepared
CyTA Journal of Food 299
at a range of 0.46.0 ppm (mg/L). The concentration of carote-
noids was expressed as mg of each individual compound per kg
of kale FW.
Statistical analysis
Statistical analyses were performed using three replicates. Data
represent the mean values of the three replicates and bars indi-
cate their standard error of the mean. Analyses of variance
(ANOVA) were conducted using JMP software version 5.0
(SAS Institute Inc. Cary, NC, USA) and mean separations per-
formed using LSD test (p< 0.05).
Results and discussion
Total phenolics, vitamin C, glucosinolates and
antioxidant capacity content of kale cultivars
The total PC, vitamin C, glucosinolates and antioxidant capa-
city values of the Winterbor and Maribor kale cultivars are
shown in Table 1. The Winterbor cultivar showed ~42% higher
concentration of PC and ~94% higher antioxidant capacity
(ORAC value) than the Maribor, whereas the Maribor cultivar
had ~141% higher vitamin C and ~159% higher total glucosi-
nolates content than the Winterbor. It is interesting to observe
that although the Maribor cultivar had higher glucosinolates
and vitamin C content than the Winterbor, the antioxidant
capacity of the Winterbor cultivar was higher than the
Maribor. Jacobo-Velázquez and Cisneros-Zevallos (2009) pro-
posed that the antioxidant capacity of plant foods is mainly
attributed to their PC content. In addition, the authors suggested
that the antioxidant activity value is not only affected by the
total PC content but also by the type of individual PC present
in the plant food (phenolic proles). Therefore, Jacobo-
Velázquez and Cisneros-Zevallos (2009) applied the concept
of specic antioxidant capacity (AOX
) value to establish the
effectiveness of a mixture of phenolic compounds to neutralize
free radicals. The AOX
value is calculated by dividing the
total antioxidant activity of a plant food by its total PC content.
Taking this into consideration, the AOX
value in the Winterbor
cultivar was ~17 mg TE/mg of PC, whereas the AOX
of the
Maribor cultivar was ~13 mg TE/mg PC. These results indicate
that the phenolic prole in the Winterbor cultivar is more
effective to neutralize free radicals than the PC present in the
Maribor cultivar. The values obtained for total PC and ORAC
are similar to those previously reported for kale (Zhou & Yu,
2006) when the values reported herein are calculated in dry
weight basis (considering 85% of moisture). Although the total
PC values in kale are relatively low compared to other
commonly consumed vegetable, the ORAC value of the
Winterbor cultivar is high and comparable to the antioxidant
capacity reported for berries such as raspberry (11920 mg TE/
kg) and strawberry (8860 mg TE/kg) (Wu et al., 2004).
The concentration of vitamin C observed in the kale varieties
evaluated herein (Table 1) are similar to values previously
reported (Sikora & Bodziarczyk, 2012). Vitamin C content in
kale is high, especially in the Maribor cultivar (Table 1). Indeed,
based on the denition of serving size for raw leafy vegetables
established by the USDA (1 cup = 67 g), the Maribor cultivar
can be considered an excellent source of vitamin C since one
serving size provides ~40% of the RDI (recommended daily
intake) for men (RDI = 90 mg/day) and ~50% of the RDI for
women (RDI = 75 mg/day). On the other hand, the Winterbor
cultivar is a good source of vitamin C, since one serving size
provides more than 10% of the RDI for men and women.
Therefore, nutrient claims such as good source of vitamin C
antioxidantand high in vitamin C antioxidantcould be used
in the label of fresh Winterbor and Maribor kale cultivars,
respectively. Compared to other green leafy vegetables, the con-
centration of vitamin C in kale is higher than lettuce (92 mg/kg),
and similar to those reported for Swiss chard (300 mg/kg) and
spinach (750 mg/kg) (USDA, 2013).
The values obtained for total glucosinolates in the Maribor
cultivar are higher than those earlier reported for Brassica vege-
tables such as broccoli (624 µmol/kg), and both kale cultivars
showed higher concentration of glucosinolates than those pre-
viously reported for Brussels sprouts (172 µmol/kg) and cauli-
ower (135 µmol/kg) (Song & Thornalley, 2007).
Glucosinolates are converted into isothiocyanates in the human
body by the action of the enzyme myrosinase (Fahey, Zhang, &
Talalay, 1997). It is well known that isothiocyanates are potent
phase II enzymes inducers that protects against chemical carci-
nogens (Aires, Carvalho, Rosa, & Saavedra, 2013; Fahey et al.,
1997). Therefore, the consumption of kale would be benecial
for the prevention of cancer, as it has been suggested for prostate
cancer (Steinbrecher, Nimptsch, Hüsing, Rohrmann, &
Linseisen, 2009). However, it would be interesting to determine
the glucosinolates prole of kale since individual compounds
differ with respect to impact on health (Dinkova-Kostova &
Kostov, 2012).
Carotenoid analysis
The typical HPLC-PDA kale carotenoid chromatograms (shown
at 450 nm) obtained from Winterbor and Maribor cultivars is
shown in Figure 1. No difference was observed between the
Table 1. Total phenolic compounds, vitamin C, glucosinolates and antioxidant capacity (ORAC value) of Winterbor and Maribor kale cultivars.
Tabla 1. Compuestos fenólicos totales, vitamina C, glucosinolatos y capacidad antioxidante (valor ORAC) de los cultivares Winterbor y Maribor de kale.
Total phenolic compounds (mg/kg) 610.3 ± 18.4 a 419.8 ± 27.8 b
Vitamin C (mg/kg) 237.8 ± 25.6 b 572.6 ± 38.4 a
Total glucosinolates (µmol/kg) 318.2 ± 41.7 b 824.5 ± 139.7 a
Total antioxidant capacity (mg trolox equivalents/kg) 10513.0 ± 373.2 a 5407.7 ± 441.6 b
Results are expressed in fresh weight (FW).
Values represent the mean of tree replications ± standard error of the mean.
Different letters in the same row indicates
statistical difference by the LSD test (p< 0.05).
Los resultados están expresados en base húmeda (FW).
Los valores representan el promedio de tres repeticiones ± el error estándar del promedio.
Diferentes letras en
la misma la indican diferencia estadísticamente signicativa por la prueba LSD (p< 0.05).
300 A. Becerra-Moreno et al.
compounds identied in both cultivars. The tentative identica-
tions of the individual carotenoids present in both cultivars are
reported in Table 2. The identity of each chromatographic peak
was assigned based on the absorption maxima of each chromato-
graphic peak by comparison with the absorption maxima of
commercial standards and prior literature data (Chandrika,
Jansz, & Warnasuriya, 2005; de Azevedo & Rodriguez-Amaya,
2004; Lee, Castle, & Coates, 2001; Mendes-Pinto, Ferreira,
Oliveira, & De Pinho, 2004; Taylor, Brackenridge, Vivier, &
Oberholster, 2006). Furthermore, the order of elution of the
individual carotenoids was considered as an additional criterion
for identity when prior publications used the same chromato-
graphic conditions (Chandrika et al., 2005; de Azevedo &
Rodriguez-Amaya, 2004; Khachik, Beecher, & Whittaker,
1986; Lee et al., 2001; Mendes-Pinto et al., 2004; Taylor et al.,
The carotenoids identied in the Winterbor and Maribor
kale cultivars included neoxanthin, violaxanthin, lutein, all-
trans-β-carotene and 9-cis-β-carotene (Figure 1,Tabl e 2). In
general, the Winterbor cultivar had higher concentration of total
carotenoids as compared to the Maribor (Table 3 ). For both
cultivars, the carotenoid that showed highest concentration was
the lutein (~44% of total). These results are consistent with
previous reports on the identication of the main carotenoids
present in kale (Khachik et al., 1986). The concentration of
lutein in the Winterbor cultivar was ~28% higher than the
Maribor. Compared to other plant foods rich in lutein, the
Winterbor cultivar contains similar amounts as those present
in the main sources of lutein (i.e. spinach, turnip greens and
collards) of the human diet (Holden, Eldridge, & Beecher,
1999). Although currently there is no RDI for lutein, the
American Macular Degeneration Foundation (AMDF) recom-
mends daily dosages of 630 mg. One serving size of the kale
cultivars evaluated herein provides 5.07.0 mg of lutein per
serving size, and thus kale could be considered an excellence
source of lutein.
5 10152025303540455055
15 20 25 30
35 40 45 50 55
Figure 1. Typical HPLC-PDA kale carotenoid and chlorophyll chromatograms (shown at 450 nm) obtained from Winterbor (A) and Maribor (B)
cultivars. Tentative identication of the chromatographic peaks was performed as indicated in Table 2. Peak assignments: (1) neoxanthin, (2) violaxanthin,
(3) chlorophyll b, (4) lutein, (5) chlorophyll a, (6) all-trans-β-carotene and (7) 9-cis-β-carotene.
Figura 1. Perl cromatográco típico obtenido mediante HPLC-PDA de carotenoides y clorolas de kale (mostrado a 450 nm) presentes en los cultivares
Winterbor (A) y Maribor (B). La identicación tentativa de los picos cromatográcos se realizó como se indica en al Tabla 2. Asignación de picos: (1)
neoxantina, (2) violaxantina, (3) clorola b, (4) luteína, (5) clorola a, (6) all-trans-β-caroteno y (7) 9-cis-β-caroteno.
CyTA Journal of Food 301
Regarding pro-vitamin A content (all-trans-β-carotene and
9-cis-β-carotene) of kale, the cultivars evaluated contain similar
amounts (Table 3). The concentration of β-carotene in kale is
similar to the content in sweet potato (92 mg/kg), carrot (88 mg/
kg), pumpkin (69 mg/kg), spinach (56 mg/kg) and collard
(33 mg/kg), which are considered the main sources of β-carotene
in the human diet (Holden et al., 1999). Converting the all-trans-
β-carotene of kale to retinol (Vitamin A) concentration (1 μgof
retinol = 6 μg of all-trans-β-carotene) (Haskell et al., 2004), each
serving size of the Winterbor and Maribor kale cultivars provides
845 μg and 763 μg of retinol, respectively. Therefore, kale can
also be considered an excellent souce of vitamin A because it
provides more than 100% of its RDI (600 μg). Nutrient claims
such as high in pro-vitamin A antioxidantcould be used in the
label of fresh kale. However, although kale is an excellent source
of antioxidants, it should be eaten in moderation to prevent
hypercarotenenameia (yellow-orange pigmentation in the skin),
which is associated with the excessive intake of vegetables rich
in pro-vitamin A carotenoids (Nagai et al., 1999).
The kale cultivars evaluated in the present study contain high levels
of antioxidants. Their content of vitamin C is high (particularly in
the Maribor cultivar), and the levels of pro-vitamin A, lutein and
glucosinolates are comparables to those present in the main sources
of these antioxidants. Therefore, kale can be considered as a func-
tional food since it may be useful for the prevention of different
chronic degenerative diseases. The extensive production and
comercialization of kale can be a realistic approach to increase
the daily intake of antioxidants in the population.
This study is based upon research supported by research funds
from the Tecnológico de Monterrey Research Chair Initiative
(CAT 161). We would also like to thank Frigorizados La Huerta,
S.A. de C.V., for providing kale cultivars. The author, A.B.-M.,
also acknowledges the scholarship (285802) from the Consejo
Nacional de Ciencia y Tecnología (CONACYT, México).
Aires, A., Carvalho, R., Rosa, E., & Saavedra, M. J. In press.
Phytochemical characterization and antioxidant properties of baby-
leaf watercress produced under organic production system. CyTA
Journal of Food, doi:10.1080/19476337.2013.769025
Sanz, C., & Hayirlioglu-Ayaz, S. (2006). Nutrient contents of kale
(Brassica oleraceae L. var. acephala DC). Food Chemistry,96(4),
Table 2. Tentative identication of kale (Brassica oleracea Acephala group) carotenoid and chlorophyll chromatographic proles obtained by HPLC-
Tabla 2. Identicación tentativa de los perles cromatográcos de carotenoides y clorolas de kale (Brassica oleracea Acephala group) obtenidos
mediante HPLC-PDA.
Peak number
(retention time) λmax
(nm) Tentative identication Previously reported in kale
Method of identication
1 (11.1) (415), 439.8, 468.8 Neoxanthin ii, iii B, C
2 (12.3) (412), 436.2, 455.2 Violaxanthin ii, iii B, C
3 (19.6) (315.4), 344, 466.4 Chlorophyll b i B, C
4 (20.9) (422), 444.7, 473.7 Lutein ii, iii A, B, C
5 (25.9) (338), (384.4), 408 Chlorophyll a i B, C
6 (40.5) (430), 451.1, 478.5 All-trans-β-carotene ii, iii A, B, C
7 (42.1) (420), 447.1, 473.7 9-cis-β-carotene iii B, C
Number of peak assigned according to the order of elution from the C30 stationary phase.
Wavelengths of maximum absorption in the UV/Vis spectra of each
chromatographic peak, values in parentheses indicate a shoulder in the peak.
Previously reported by (i) Khachik et al. (1986), (ii) Khachik et al. (1991) and (iii) de Azevedo &
Rodriguez-Amaya (2004).
Method applied for the identication of the peak: (A) identication by comparison with the retention time and wavelengths of maximum absorption
in the UV/Vis spectra of commercial standards; (B) identication by spectral interpretation of the wavelengths of maximum absorption in the UV/Vis spectra and comparison
with wavelengths of maximum absorption reported in the literature (Chandrika et al., 2005; de Azevedo & Rodriguez-Amaya, 2004; Lee et al., 2001; Mendes-Pinto et al., 2004;
Taylor et al., 2006); (C) identication by the order of chromatographic elution reported in the literature (Chandrika et al., 2005; de Azevedo & Rodriguez-Amaya, 2004;
Khachik et al., 1986; Lee et al., 2001; Mendes-Pinto et al., 2004; Taylor et al., 2006).
Número de pico asignado de acuerdo al orden de elución de la fase estacionaria C30.
Longitud de onda máxima de absorción en el espectro UV/Vis de cada pico
cromatográco, los valores en paréntesis indican un hombro en el pico.
Previamente reportado por, (i) Khachik et al. (1986), (ii) Khachik et al. (1991) y (iii) de Azevedo &
Rodriguez-Amaya (2004).
Método aplicado para la identicación del pico: (A) identicación por comparación con el tiempo de retención y la longitud de onda máxima de
absorción en el espectro UV/Vis de estándares comerciales; (B) identicación por la interpretación del espectros de absorción máxima en el espectroUV/Vis de acuerdo a
reportes previos de la literatura (Chandrika et al., 2005; de Azevedo & Rodriguez-Amaya, 2004; Lee et al., 2001; Mendes-Pinto et al., 2004; Taylor et al., 2006); (C)
identicación por el orden de elución reportado en la literatura (Chandrika et al., 2005; de Azevedo & Rodriguez-Amaya, 2004; Khachik et al., 1986; Lee et al., 2001; Mendes-
Pinto et al., 2004; Taylor et al., 2006).
Table 3. Individual carotenoids and chlorophylls content in Winterbor
and Maribor kale cultivars.
Tabla 3. Contenido de carotenoides y clorolas individuales en los
cultivares de Winterbor y Mirabor de kale.
Individual carotenoids
content (mg/kg FW)
Winterbor Maribor
Neoxanthin 20.88 ± 1.26 a 5.91 ± 1.40 b
Violaxanthin 14.64 ± 0.28 a 11.26 ± 0.61 b
Chlorophyll b 100.48 ± 4.35 a 83.67 ± 2.94 b
Lutein 100.05 ± 3.83 a 71.58 ± 2.28 b
Chlorophyll a 87.85 ± 4.02 a 66.21 ± 2.47 b
All-trans-β-carotene 75.68 ± 3.57 a 68.31 ± 2.17 a
9-cis-β-carotene 12.88 ± 0.40 a 7.91 ± 0.62 b
Concentrations are reported as lutein equivalents for peaks 1, 2, 3, 4 and 5;
and as all-trans-β-carotene equivalents for peaks 6 and 7. All compounds were
quantied at 450 nm.
Values represent the mean of 3 replications ± standard error
of the mean.
Different letters in the same row indicate statistical difference by the
LSD test (p< 0.05).
Las concentraciones para los picos 1, 2, 3, 4 y 5 están reportadas como
equivalentes de luteína; y para los picos 6 y 7 como equivalentes de all-trans-β-
caroteno. Todos los compuestos se cuanticaron a 450 nm.
Los valores representan
el promedio de 3 repeticiones ± el error estándar del promedio.
Diferentes letras en
la misma la indican diferencia estadísticamente signicativa por la prueba LSD
(p< 0.05).
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The sedentary lifestyle coupled with continuously changing food habits and search for nutrient-dense enriched protective foods has resulted in increased demand and consumption of natural foods. Among natural dietary ingredients, vegetables are the daily consumed dietary ingredients packed with vitamins, minerals, antioxidants, and an array of bioactive phytochemicals. Amongst vegetable crops, cruciferous vegetables like broccoli, cabbage, cauliflower, arugula, horseradish, mustard green, bok choy, brussels sprouts, etc. are the crops which are perceived far important than the mere table items for daily consumption owing to their rich functional bioactive profile. Presence of the sulfur-rich compounds (methyl cysteine sulfoxide and glucosinolates), coloring pigments (carotenoids, anthocyanins), minerals (Se, Fe, K, Ca), vitamins (B complex and C), dietary fiber, and other bioactive compounds (phytoalexins, terpenes, tocopherols, hydroxycinnamic acid, chlorogenic acid, ferulic acid, synapic acid and flavonols) give them the distinctive nutraceutical status with well documented therapeutic benefits. Most of the research effort in the last decade has been directed to effectively find out the exact mode of action of these bioactive compounds on health with their minimum effective concentration, means to ensure their effective delivery to the target organs, and increased bioavailability of these compounds. Though, there is substantial evidence based on in vivo and in vitro findings that scientifically demonstrate their benefits more research needs to be conducted with an exploration of the unknown beneficial activities as well as the unwanted effects. Future research should be directed towards the functional enrichment either through genetic modifications or through regulation of pathways for ensuring the national and health security to the general population and the health-conscious people.
This chapter will explore the main techniques and commercialized methodologies for the extraction and formulation of valuable components from tomato processing by-products, predominantly carotenoids and phenolic compounds, along with vitamins, dietary fibers, proteins, and essential oils. Following a brief outline regarding the salient properties of the valuable ingredients present in tomato processing waste, a survey will be conducted on their potential applications in relation to health and well-being, whereas market existing products and their characteristics will be described, too.
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Carotenuids of the fruit pulp of Chrysophyllum roxburghii (Sinhala: lavalu) amounted to about 180 mgkg-1 by fresh weight. The carotenoids were isolated by open column chromatography (MgO:Celite 1:1) using mixtures of petroleum ether 40-60°C and acetone and identified by UV/visible spectra, chemical tests, and High Performance Liquid Chromatography (HPLC) using authentic standards and a photodiode array detector (PAD). The major carotenoid was trons-violaxanthin (113 mgkg-1). Also present was cisviolaxanthin, neoxanthin, β-cryptoxanthin monoepoxide, lutein, β-cryptoxanthin, ζ-carotene and β-carotene. The retinol equivalent of the pulp was only 68 RE/100 g. The study shows that Chrysophyllum roxburghii is not a good source of provitamin A. Further as violaxanthin is reported to be not absorbed by humans, it is of no use as a dietary antioxidant. However, as trans-violaxanthin can be obtained in quantity in the pure crystalline state, directly from the column and has the benefit of eluting at 20% acetone away from most carotenoids. the compound will be useful as a standard for HPLC analysis of carotenoids from other fruits and leaves.
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The aim of this study was to evaluate the phytochemical composition and the antioxidant properties of watercress (Nasturtium officinale R. Br., Brassicaceae) produced under organic production system. Fresh baby-leaf watercress samples were collected from local organic farms. High performance liquid chromatography with diode array detection (HPLC-DAD) and high performance liquid chromatography with diode array detection mass spectrometry (HPLC-MS) were used to assess the phytochemical composition, and spectrophotometric methods were used to assess the antioxidant capacity. Two major classes of healthier secondary plant metabolites were identified: phenolics and glucosinolates. The major phenolics determined were chlorogenic acid, quercetin-3-O-rutinoside, dicaffeoyltartaric acid, and isorhamnetin. The glucosinolates were composed exclusively of gluconasturtiin, the precursor of the anticarcinogenic, and antimicrobial compound 2-phenylethyl isothiocyanate. The extracts of organic young baby-leaf watercress presented high antioxidant capacity and this property was highly related with caffeic acid, quercetin-3-O-rutinoside, isorhamnetin, and glucosnastrutiin. The results achieved showed that baby-leaf watercress can provide high contents of antioxidant compounds at levels even superior to similar adult plant material.
The limited availability of foods that are free of pesticides has led Brazil to search for alternative production methods to meet the desires of consumers. Currently, organic cultivation represents a production system that complies with general expectations of producers and consumers. Organic cultivation is particularly interesting mainly because of its effect on plant secondary metabolite content, which may help plants to naturally combat pests; in humans, these substances can also contribute to the prevention of chronic diseases. We report on the extraction of glucosinolates (both as total glucosinolates and as benzylglucosinolate) with trifluoroacetic acid addition in a 70:30 MeOH:water (v/v). Total glucosinolates, determined by a thioglucosidase coupled assay, were measured in different Brassicaceae species and were similar to values reported in the literature. For broccoli, analyses were carried out separately on inflorescences, leaves and stalks; analyses were also conducted on thermally processed samples to simulate cooking. Furthermore, when the analysis was conducted on conventional and organic products, the highest concentrations of these substances were most often found in organically cultivated Brassicaceae. The benzylglucosinolate concentrations were evaluated on the same samples using HPLC. The concentration of benzylglucosinolate was significantly higher in organically cultivated vegetables, as well.
Major constituents of the extracts from five green vegetables (broccoli, cabbage, spinach, brussels sprouts, kale), several of which are members of the genus Brassica (Cruciferous), have been separated by high-performance liquid chromatography (HPLC) on a C-18 reversed-phase column. Three classes of compounds were shown to be present. In the order of chromatographic elution, these were xanthophylls, chlorophylls and their derivatives, and the hydrocarbon carotenoids (carotenes). The xanthophylls were identified as neoxanthin, violaxanthin, lutein epoxide, and lutein. Several mono cis isomers of xanthophylls were also shown to be present in the extracts from these vegetables. The chlorophylls were identified as chlorophylls b and a and their decomposition products pheophytins b and a. The only hydrocarbon carotenoids present in these vegetables were all-trans-β-carotene and its 15,15′-cis isomer. β-Apo-8′-carotenal and decapreno-β-carotene have been employed respectively as internal standards for quantification of xanthophylls and carotenes. The effect of cooking on the qualitative and quantitative distribution of carotenoids in some of the vegetables has been discussed.
A total of 38 commonly consumed vegetable samples, including 3 kale, 1 rhubarb, 3 spinach, 3 broccoli, 2 green bean, 5 carrot, 10 tomato, and 11 potato samples, were investigated for their total phenolic contents (TPC) and antioxidant properties. The measured antioxidant properties included free radical scavenging activities against DPPH, superoxide anion radical (O2−), ABTS+ and peroxyl radical, and Fe2+ chelating capacity. The tested vegetables differed in their antioxidant properties and TPC, although all vegetables had significant antioxidant activities and contained significant levels of phenolics. The TPC and the measured antioxidant properties were correlated to each other. The Fe+2 chelating capacity and the scavenging capacities against ABTS+, DPPH and O2− were reported for these commonly consumed vegetables for the first time. Results from this study suggest that kale, spinach, broccoli, and rhubarb are the better dietary sources of natural antioxidant activities and phenolic compounds. Furthermore, kale, broccoli and spinach produced in Colorado may have greater antioxidant contents than those grown at other locations.
When compared to thermal and chemical alternatives, high hydrostatic pressure (HHP) processing is the most effective non-thermal technology to process avocados. Herein we report the effects of HHP-processing (600 MPa, 3 min) and storage (40 days, 4 °C) on the stability of avocado paste (Persea americana, cv.Hass) carotenoids. Likewise, the effects of HHP-processing and storage on hydrophilic and lipophilic oxygen radical ab- sorbance capacities (ORAC) of the product were studied. Pressurization induced a significant increase (approx. 56%) in concentrations of total extractable carotenoids. Highest increases for individual carotenoids were observed for neoxanthin-b (513%), followed by !-cryptoxanthin (312%), !-carotene (284%), à-cryptoxanthin (220%), à-carotene (107%), and lutein (40%). Carotenoid levels declined during storage, but at the end of the product's sensory shelf-life were higher than those initially present in unprocessed avocado paste. Interestingly, ORAC-values followed a different trend than carotenoids; they decreased immediately after HHP-processing and increased during storage, therefore indicating that carotenoids appear to be minor contributors to the total antioxidant capacity of the fruit.
Cabbage vegetables, like Brassica group, are perceived as very valuable food products. They have a very good nutritive value, high antioxidant activity and pro-healthy potential. Especially, kale (Brassica oleracea L. var. acephala) is characterized by good nutritional and pro-healthy properties, but this vegetable is not popular in Poland. The aim of this work was to assess the chemical composition and antioxidant activity of kale variety Winterbor F(1) and investigation of cooking process on selected characteristics. The chemical composition and antioxidant activity were determined in leaves of kale Winterbor F(1) variety after three subsequent years of growing. In one season, analyses were performed on raw and cooked leaves. The investigated kale was characterized by high average contents of: β-carotene (6.40 mg/100 g f.m.), vitamin C (62.27 mg/100 g f.m.), alimentary fiber (8.39 g/100 g f.m.) and ash (2.11 g/100 g f.m.). The average amounts of nitrites (III) and (V) were 3.36 mg NaNO(2)/kg f.m. and 1206.4 mg NaNO(3)/kg f.m., respectively. The investigated kale contained polyphenolic compounds at average level of 574.9 mg of chlorogenic acid/100 g f.m., and its antioxidant activity measured as ABTS radical scavenging ability was 33.22 μM Trolox/g of fresh vegetable. It was observed a significant lowering of antioxidant compounds as a result of cooking. The losses of vitamin C were at about 89%, polyphenols at the level of 56%, in calculation on dry mass of the product. The highest stability was shown in the case of beta-carotene, for which the losses were at about 5%. Antioxidant activity of cooked vegetable lowered and reached the level of 38%. There were also some losses observed in macro-components from 13% for zinc to 47% for sodium. The contents of harmful nitrites and nitrates in calculation on dry mass were significantly lower as a result of cooking, by 67% and 78%, respectively. Winterbor F(1) variety of kale has a great nutritive value and high antioxidant activity. The cooking process of kale resulted in lowering of the antioxidant activity of its antioxidants especially of vitamin C, polyphenols and to the lesser extent of β-carotene what confirms that vegetable should be eaten in raw form or just undergo little processing before consumption, for example blanching.
The principal carotenoids of kale were identified by chemical reactions, high-performance liquid chromatography/mass spectrometry and high-performance liquid chromatography/photodiode array detection and were quantified by the last technique. In kale taken from conventional farms, the β-carotene and lutein contents were significantly higher in the mature leaves, violaxanthin was at an unusually high level in the young leaves, and neoxanthin had practically the same concentration at both stages of maturity. In samples taken from an organic farm, the carotenoid contents were essentially the same in the young and the mature leaves. Except for β-carotene, which did not differ with season, the carotenoid concentrations of marketed minimally processed kale were found to be significantly higher in the summer than in the winter, reflecting seasonal rather than processing effects. In minimally processed kale monitored during 5 days of storage at 7–9 °C, β-carotene, lutein, violaxanthin and neoxanthin were reduced by 14, 27, 20 and 31% respectively. Thus minimal processing, seasonal and maturity factors were found to have an influence on the carotenoid content of kale. Copyright © 2004 Society of Chemical Industry