Journal of the Science of Food and Agriculture J Sci Food Agric 87:437–446 (2007)

Effect of genotype, maturity stage

and post-harvest storage on phenolic
compounds, carotenoid content and
antioxidant capacity, of Andean mashua tubers
(Tropaeolum tuberosum Ruiz & Pavón)
Rosana Chirinos,1,4 David Campos,1∗ Carlos Arbizu,2 Hervé Rogez,3
Jean-François Rees,4 Yvan Larondelle,4 Giuliana Noratto5 and
Luis Cisneros-Zevallos5∗
1 Instituto de Biotecnologı́a (IBT), Universidad Nacional Agraria-La Molina, Lima 12, Peru
2 Centro Internacional de la Papa (CIP), La Molina, Lima 12, Peru
3 Departamento de Engenharia Quı́mica e de Alimentos, Universidade Federal do Pará, Belém-PA, Brazil
4 Unité de Biologie Animale & 4 Unité de Biochimie de la Nutrition, Institut des Sciences de la Vie, Université Catholique de Louvain, B-1348

Louvain-La-Neuve, Belgium
5 Department of Horticultural Sciences, Texas A&M University, College Station, Texas 77843-2133, USA

Abstract: The total phenolics, anthocyanins, flavan 3-ols, carotenoids and antioxidant capacity of mashua
(Tropaeolum tuberosum Ruiz & Pavón) tubers from 10 yellow and purple cultivars were determined at different
maturity stages (5–7.5 months after planting) and sunning post-harvest storage periods (7–35 days). Both the
antioxidant capacity (ORAC and ABTS assays) and the content of the bioactive compounds tested varied
markedly between cultivars. Purple varieties reached the highest antioxidant capacity during tuber development
(271– 446 µmol Trolox equiv g−1 DM, ORAC assay). The kinetics of accumulation or disappearance of the bioactive
compounds tested during maturation was dependent both on the cultivar and on the compound considered. For
anthocyanins, there was a marked increase during maturation in all the purple cultivars. During the sunning post-
harvest storage, the changes in antioxidant capacity (ABTS assay) and content of the bioactive compounds tested
also varied between cultivars. A marked decrease in anthocyanins was observed for the anthocyanin-containing
cultivars. In general, the correlation between antioxidant capacity and the content of bioactive compounds varied
markedly between cultivars. Antioxidant capacity in purple varieties correlated with total phenolics or flavan 3-ols
while only in some yellow varieties antioxidant capacity correlated with total phenolics.
 2007 Society of Chemical Industry

Keywords: mashua; Tropaeolum tuberosum; antioxidants; phenolic compounds; carotenoid; maturity stage; post-
harvest

INTRODUCTION under low levels of input.3,4 Mashua is also a valuable

There is growing interest in the search for edible plants
as novel sources of health-promoting phytochemicals.
Mashua (Tropaeolum tuberosum Ruiz & Pavon) is
a tuber crop indigenous to the Andean highlands
and is of economic value as a food and medicinal
crop. It is cultivated in the Andes of Bolivia, Peru,
Ecuador, Colombia and Venezuela1 and is currently
being grown experimentally in New Zealand and
the Pacific northwest to evaluate its potential for
worldwide cultivation.2 Mashua is adapted to high
Andean altitudes, low minimum temperatures, large
ranges of diurnal temperatures and is highly productive
crop because it is resistant to many insects, nematodes,
fungi and other pathogens that attack potatoes and
other tuber crops.5 The nutritional value of mashua
is high. For example, 100 g dry tubers may contain
14–16 g protein, almost 80 g carbohydrate and about
77.5 mg vitamin C.6,7 It has been traditionally and
extensively used in the folk medicine of the Andean
region, and its domestication may have been related
to its importance as a medicinal agent.8 Recent
studies indicate that mashua has a high content
of phenolic compounds (0.92–3.37 mg chlorogenic
acid equivalent g−1 fresh weight), anthocyanins

∗
Correspondence to: Luis Cisneros-Zevallos, Department of Horticultural Sciences, Texas A&M University, College Station, Texas 77843-2133, USA
E-mail: lcisnero@tamu.edu
David Campos, Instituto de Biotecnologia (IBT), Universidad Nacional Agraria-La Molina, Lama 12, Peru
E-mail: dcampos@lamolina.edu.pe
Contract/grant sponsor: USAID, through the Association Liaison Office for University Cooperation in Development (ALO).; contract/grant number: HNE-A-00-
97-00059-00
Contract/grant sponsor: Consejo Nacional de Ciencia y Tecnologı́a del Perú (Concytec)
Contract/grant sponsor: Belgian Coopération Universitaire au Développement
(Received 13 April 2005; revised version received 18 January 2006; accepted 5 September 2006)
Published online 10 January 2007; DOI: 10.1002/jsfa.2719

 2007 Society of Chemical Industry. J Sci Food Agric 0022–5142/2007/$30.00

R Chirinos et al.
(0.5–2.05 mg cyanindin 3-glucoside equivalent g−1
fresh weight), as well as an important hydrophilic
antioxidant capacity (955–9800 µg Trolox equivalents
g−1 fresh weight, ABTS assay).9 It was also found that
extracts of mashua induced concentration-dependent
suppression of the proliferation of human DU145
prostate carcinoma cells, human Caco-2 colon
adenocarcinoma cells and murine B16F10 melanoma
cells.10 This Andean crop is ready for harvest in
6–8 months and farmers often harvest it progressively
according to their needs, sometimes leaving tubers in
the ground for several weeks after the normal harvest
time. It is folklore knowledge that after sunning storage
the tubers become more suitable for eating,3 mainly
because of a lowering of their glucosinolate content.11
Interest in the role of antioxidants from fruit and
vegetables in human health has prompted research
within horticulture and food science to determine how
their content and activity can be maintained or even
improved through cultivar development, production
practices and food processing.12,13 The effects of
harvest time (age of plant) and post-harvest storage
on the retention of these dietary antioxidants is also
of major importance, together with differences in
the stability of these compounds between different
cultivars. Although there is a large germplasm of
mashua available at the International Potato Center
(Lima, Peru), very little research has been carried out
on the phytochemicals and antioxidant capacity of
different mashua cultivars. Virtually no information
is available on the changes of these constituents
during tuber development and post-harvest storage.
The purpose of this investigation was to compare 10
morphologically well identified cultivars of mashua
in terms of tuber antioxidant capacity and contents
in bioactive compounds such as phenolics and
carotenoids and to determine the effects of harvest
time (age of plant) and sunning post-harvest storage
on these parameters.
MATERIALS AND METHODS
Chemicals and reagents
All solvents (methanol, ethanol, acetone and hex-
ane) and other chemicals of analytical grade were
obtained from Merck (Darmstadt, Germany) and
Riedel de Haën (Seelze, Germany). 2,2 -azinobis (3-
ethylbenzothiazoline-6-sulfonic acid) (ABTS), fluo-
rescein, 2,2 -azobis (2-amidinopropane) dihydrochlo-
ride (AAPH), 4-(dymethylamino)-cinnamaldehyde
solution (DMCA), Folin-Ciocalteu reagent (Catalog
# F9252), chlorogenic acid, (+)-catechin hydrate,
Trolox (6-hydroxy-2,5,7,8-tetramethyl chromane-2-
carboxylic acid) and β-carotene were purchased from
Sigma Chemicals Co. (St Louis, MO).
Mashua sampling
Tubers of 10 cultivars of mashua (Tropaeolum
tuberosum Ruiz & Pavón) were supplied by the
International Potato Center (CIP, Lima, Peru). The
438
tubers were grown in the rural community of La
Libertad, Huancayo (Junin, Peru) at approximately
3700 m above sea level, at approx. 70% relative
humidity and 10 ◦ C (average temperature). They were
planted at the end of September 2002 and harvested
at 5 (beginning of tuberization process), 5.5, 6, 6.5,
7 and 7.5 months (usual harvest time) after being
planted. Sets of tubers harvested at 7.5 months were
stored on a rooftop at CIP’s experimental station
Santa Ana (Huancayo, Peru) at 3280 m above sea
level and exposed 10 h a day to bright sunlight (average
temperature 15 ◦ C) during 7, 14, 21, 28 and 35 days.
During the night period (12 h) the tubers were covered
with Ichu (Andean grass) to protect them from the
low temperatures (−2 to −15 ◦ C). Harvested tubers
were immediately packed in paper bags and sent to
the Universidad Agraria La Molina (Lima, Peru)
where they were lyophilized and stored at −20 ◦ C.
Three samples made of 10 tubers of each cultivar
were randomly collected for analysis. The lyophilized
samples were rehydrated prior to the analyses to
improve the recoveries of the extraction procedures.
All analyses were conducted in triplicate.
DETERMINATION OF ANTIOXIDANT CAPACITY
ABTS assay
Antioxidant capacity was determined by the ABTS
method adapted from Arnao et al.14 For hydrophilic
antioxidant capacity, a sample of 0.3 g was homog-
enized with 20 mL of methanol for 2 min using an
Ultra-Turrax homogenizer (IKA works, Inc., Wilm-
ington, NC) and left at 4 ◦ C for 24 h. The extract was
centrifuged at 20 000 × g for 15 min and the super-
natant was collected. A 150 µL of supernatant was
mixed with 2.85 mL of ABTS solution prepared as
described by Awika et al.15 with an absorbance of
approx. 1.1 measured at 734 nm. This mixture was
allowed to react at 20 ◦ C until a steady absorbance
was reached. For measurements of lipophilic antioxi-
dant capacity (LAC), the pellet was mixed with 20 mL
of dichloromethane and shaken for 15 min before fil-
tration. An aliquot of the yellow–orange supernatant
was taken and treated as described above. Simultane-
ously, a 150 µL aliquot of either solvent was treated
in the same way as the sample and used as a con-
trol. The Genesys-5 UV–visible spectrophotometer
(Milton Roy, New York, NY) was blanked with the
respective solvent, and the decrease in absorbance due
to antioxidants was recorded at 734 nm. Both antiox-
idant capacities were calculated as µmol of Trolox
equivalents (TE) g−1 dry matter (DM) from a standard
curve developed with Trolox.
Oxygen radical absorbance capacity
The oxygen radical absorbance capacity (ORAC)
assay was carried out on different mashua genotypes
during the maturity stage studies with the purpose
of having a reference comparison with the ABTS
assay used and with information published in the
J Sci Food Agric 87:437–446 (2007)
DOI: 10.1002/jsfa

literature for other crops. A fluorimeter Ascent F.L.
(Fluoroscan Labsystems, Helsinki, Finland) was used.
The ORAC values were determined by the method
adapted from Ou et al.,16 Huang et al.17 and Cao
and Prior18 using microplates (96-well, opaque white;
Greiner Bio-One,Wemmel, Belgium). The change of
fluorescence intensity is an index of the degree of
damage caused to the fluorescein protein due to free
radicals. Fluorescein and the peroxyl radical generator
(AAPH) were prepared in a 75 mmol L−1 phosphate
buffer at pH 7.4. The samples were treated as
described for the ABTS assay. Twenty-five microliters
of methanolic extract and 250 µL of 55 nmol L−1
fluorescein were placed in a 96-well microplate and
preincubated at 37 ◦ C for 10 min before automatic
injection of an AAPH solution (25 µL, 153 nmol
L−1 ) and measurement of the initial fluorescence. The
phosphate buffer was used as a blank and Trolox as a
standard during each run. Fluorescence readings were
performed every minute until a value lower than 5%
of the initial fluorescence was reached (about 50 min).
Fluorescence conditions were: excitation at 485 nm,
emission at 520 nm. The relative ORAC values were
calculated using the differences of areas under the
decay curves and were expressed as µmol TE g−1 DM.
Determination of total phenolics
Total phenolics were determined using Folin–
Ciocalteau reagent.19 A sample of 0.3 g was homog-
enized with 95% aqueous ethanol for 2 min and
left at 4 ◦ C for 24 h. The extract was centrifuged
at 20 000 × g for 15 min. A 0.5 mL of supernatant
(0.5 mL of 95% aqueous ethanol for the blank) was
diluted with 8 mL of distilled water. The Folin-
Ciocalteu reagent was diluted with water (dilution
factor 8) and 0.5 mL were added to the diluted
extracts, vortex mixed and allowed to react for 3
min. Then, 1 mL of 0.5 mol L−1 Na2 CO3 was added
and allowed to react for 10 min. The absorbance was
measured at 725 nm when a steady state was reached.
Chlorogenic acid was used as standard for the calibra-
tion curve, and the total phenolics were expressed as
mg of chlorogenic acid equivalents g−1 DM.
Determination of total anthocyanins
Anthocyanin quantitation was performed by the pH
differential method.20 A 0.2 g of sample and 15 mL of
extraction solvent (95% aqueous ethanol:HCl 1.5 mol
L−1 , 85:15, v/v) were homogenized for 2 min and
left at 4 ◦ C for 24 h. The extract was centrifuged
at 20 000 × g for 15 min. The supernatant was
diluted ten times in a pH 1.0 solution (0.1 mol
L−1 HCl, 25 mmol L−1 KCl) and in a pH 4.5
solution (0.4 mol L−1 CH3 COONa). The absorbance
of the mixtures was then measured at 535 and
700 nm against distilled water. The value (Abs535 −
Abs700 )pH1.0 − (Abs535 − Abs700 )pH4.5 corresponds to
the absorbance due to the anthocyanins. Calculation
of the anthocyanin concentrations was based on a
cyanidin 3-glucoside molar extinction coefficient of
J Sci Food Agric 87:437–446 (2007)
DOI: 10.1002/jsfa
Antioxidants during maturity and storage of Mashua tubers
25 96521 and a molecular weight of 449 g mol−1 .
Results were expressed as mg of cyanidin 3-glucoside
equivalents g−1 DM.
Determination of flavan 3-ols
The method for the determination of flavan 3-ols
was adapted from Delcour and de Varebeke22 and
McMurrough and McDowell.23 A 0.2 g of sample
was homogenized with 15 mL of aqueous acetone
solvent (70% v/v) for 2 min. Then the mixtures
were shaken at 500 rpm in a orbital shaker (IKA
works, Inc., Wilmington, NC) for 1 h, and centrifuged
at 20 000 × g for 15 min. The supernatants were
collected and concentrated in a rotary evaporator at
<35 ◦ C until acetone was eliminated. One milliliter of
DMCA solution (250 mg in 500 mL of HCl:MeOH,
30:70, v/v) was added to 200 µL of concentrated
extract to react at 20 ◦ C. Simultaneously, a blank was
prepared with distilled water and treated in the same
way as the samples. The absorbance was measured at
640 nm, when a steady value was reached. Catechin
was used as standard for the calibration curve and the
flavan 3-ols content was expressed as mg of catechin
equivalents g−1 DM.
Determination of total carotenoids
The total carotenoids were estimated by the method
reported by Talcott and Howard.24 A 0.2 g of
sample was homogenized for 2 min with 20 mL of
acetone/ethanol solution (1:1, v/v) containing 200 mg
L−1 2,6-di-tert-butyl-4-methylphenol (BHT). The
extract was centrifuged at 20 000 × g for 15 min.
The supernatant was transferred into a graduated
cylinder and the acetone/ethanol solvent was added
to a final volume of 100 mL. Fifty milliliters of hexane
and 25 mL of water were then added. The mixture
was shaken vigorously before standing for 30 min to
allow the phases to separate. The spectrophotometer
was blanked with hexane and the absorbance of the
hydrophobic phase (hexane) was measured at 470 nm.
β-Carotene was used as standard for the calibration
curve and the total carotenoids content was expressed
as µg of β-carotene equivalents g−1 DM.
Statistical analysis
SPSS for Windows 11.5 was used. Two-way ANOVA
tests were performed with cultivars and maturity
stages or post-harvest storage as fixed effects and
content of bioactive compounds or antioxidant activity
as dependent variable. Means were compared with
Tukey’s multiple-range comparison test (α = 0.05).
Pearson correlations at the level 0.01 and 0.05 were
performed.
RESULTS AND DISCUSSION
Effect of genotype in terms of antioxidant
capacity, total phenolics, flavan 3-ols, total
anthocyanins and total carotenoids
The changes of antioxidant capacity, total pheno-
lics flavan 3-ols, anthocyanins, and carotenoids, for
439
R Chirinos et al.

500 500

(µmol of TE g−1 DM) by ORAC

(µmol of TE g−1 DM) by ABTS (a) (b)
450 450
400 400

Antioxidant capacity
Antioxidant capacity

350 350
300 300
250 250
200 200
150 150
100 100
50 50
0 0
5 5.5 6 6.5 7 7.5 5 5.5 6 6.5 7 7.5
30 8
(c) (d)
Total phenolics (mg g−1 DM)

Flavan-3-ol (mg g−1 DM)

25
6
20
5
15 4
3
10
2
5
1
0 0
5 5.5 6 6.5 7 7.5 5 5.5 6 6.5 7 7.5
10 140
9 (e) (f)
Anthocyanin (mg g−1 DM)

120
Carotenoids (µg g−1 DM)

8
7 100
6
80
5
4 60

3 40
2
20
1
0 0
5 5.5 6 6.5 7 7.5 5 5.5 6 6.5 7 7.5
Maturity stage (month) Maturity stage (month)

Figure 1. Changes in antioxidant capacity (ABTS assay (a) and ORAC (b)), and in total phenolics (c), flavan 3-ols (d), anthocyanins (e), and
carotenoids (f) in mashua tubers harvested at different maturity stages. (-♦-) DP 0224; (--) ARB 5241; (--) AGM 5109; (--) ARB 5576; (--)
AVM 5562; (- -) M6COL2C; (- -- -) DP 0223; (- -- -) DP 0203; (- -) DP0207; (- -♦- -) DP 0215, (mean values, n = 3).
ž
each cultivar at different maturity stages and post- for ARB 5241, DP 0224 and AGM 5109 (446, 259
harvest storage are shown in Figs 1 and 2, respectively. and 271 µmol of TE g−1 DM, respectively), were
Mean antioxidant capacity values ranged from 80 to higher than the values reported for various berry
378 µmol TE g−1 DM, and from 59 to 389 µmol TE species,25 (35–162.1 µmol TE g−1 DM). Accordingly,
g−1 DM for the ABTS and ORAC tests, respectively. the three dark-purple mashua cultivars had high phe-
The values of total phenolics, flavan 3-ols, antho- nolics contents (14–24 mg g−1 DM, at 7.5 months),
cyanins and carotenoids (mean values for all maturity comparable to raspberry cultivars25 (12.0–15.3 mg g−1
stages evaluated) for the 10 cultivars studied ranged DM), blackberry cultivars (12.1–15.6 mg g−1 DM)
from 9 to 21 mg chlorogenic acid equivalents g−1 DM, and purple corn26 (19 mg g−1 DM), but lower than
0.2 to 5.3 mg catechin equivalents g−1 DM, 2.4 to red sweet potato26 (32.2 mg g−1 DM). Anthocyanins
5.7 mg cyanidin 3-glucoside equivalents g−1 DM, and were only present in the purple mashua cultivars DP
70 to 132 µg β-carotene equivalents g−1 DM, respec- 0224, ARB 5241 and AGM 5109 and, at 7.5 months,
tively. ARB 5241, DP 0224 and AGM 5109, the reached values of 8.7, 4.7 and 3.7 mg g−1 DM, respec-
three purple cultivars investigated, presented the high- tively. These values were lower than those reported
est rankings for antioxidant capacity, and flavan 3-ols, for purple corn26 (17.7 mg g−1 DM) and blackberry
but the lowest ones for carotenoids. At the last matu- cultivars25 (7.9–10.2 mg g−1 DM) but were higher
rity stage evaluated (7.5 months), the ORAC values than in various strawberry cultivars25 (2.3–3.8 mg g−1

440 J Sci Food Agric 87:437–446 (2007)

DOI: 10.1002/jsfa
 Antioxidants during maturity and storage of Mashua tubers

500 30
(a) (b)
450

Total phenolics (mg g−1 DM)

(µmol TE g−1 DM) by ABTS 25
400
Antioxidant capacity

350
20
300
250 15
200
10
150
100
5
50
0 0
0 7 14 21 28 35 0 7 14 21 28 35

8 10
(c) (d)
9

Anthocyanins (mg g−1 DM)

7
Flavan-3-ol (mg g−1 DM)

8
6
7
5 6
4 5
4
3
3
2 2
1 1
0
0 0 5 10 15 20 25 30 35
0 7 14 21 28 35
Sunning post-harvest (day)
180
(e)
160
Carotenoids (µg g−1 DM)

140
120
100
80
60
40
20
0
0 5 10 15 20 25 30 35
Sunning post-harvest (day)

Figure 2. Changes in antioxidant capacity ABTS assay (a), anthocyanins (b), flavan 3-ols (c), total phenolics (d) and carotenoids (e) in mashua
tubers at different sunning post-harvest storage periods. (-♦-) DP 0224; (--) ARB 5241; (--) AGM 5109; (--) ARB 5576; (--) AVM 5562; (- -) ž
M6COL2C; (- -- -) DP 0223; (- -- -) DP 0203; (- -) DP0207; (- -♦- -) DP 0215, (Mean values, n = 3).

DM). Carotenoids compounds were present in all cul- to 37 g (5 months) and increased to a range of approx.
tivars. At normal harvest time yellow mashua cultivars 28 to 77 g (7.5 months) (data not shown). The changes
DP 0207, AVM 5562 and ARB 5576 showed the in antioxidant capacity (ABTS and ORAC assay), total
highest values. When expressed on fresh weight, the phenolics, flavan 3-ols, anthocyanins and carotenoids
carotenoid values obtained for these three cultivars with the maturity stage of tubers are shown in Fig. 1.
at 7.5 months after planting (8.9–14.4 µg β-carotene Significant differences in antioxidant capacity were
g−1 fresh weight) were higher than those for potatoes found between the 10 mashua cultivars for each
reported by Breithaupt and Bamedi27 (0.6–1.8 µg maturity stage (one-way ANOVA test, P < 0.05,
β-carotene g−1 fresh weight) and Iwanzik et al.28 analyses not presented). The patterns of changes
(1.7–3.4 µg β-carotene g−1 fresh weight). These high were also different between cultivars (Fig. 1a and
carotenoid-ranking cultivars ranked lower for the b). Regarding the purple cultivars, the antioxidant
other four traits studied, especially for the hydrophilic capacity of ARB 5241 tubers increased regularly from
antioxidant capacity. 5.0 to 7.5 months after planting, while it reached a
maximum at 6.0 months for DP 0224 and remained
The effect of maturity stage relatively constant during the whole maturation
During the development of mashua tubers, initial process for AGM 5109. The patterns of changes along
weight along different cultivars ranged from approx. 4 maturation were less variable in the case of the yellow

J Sci Food Agric 87:437–446 (2007) 441

DOI: 10.1002/jsfa
R Chirinos et al.
cultivars and showed a trend to remain constant or
slightly decrease. Studies performed on blackberries
and strawberries reported that the highest ORAC
values were found during the green stages, whereas
red raspberries had the highest ORAC activity at the
ripe stage.25 Similarly, Prior et al.29 have reported
that ORAC values for Vaccinium species increased
with increasing maturity. All these results indicate
that many processes of synthesis and degradation of
compounds with antioxidant properties may take place
during maturation, resulting in different balances in
the overall antioxidant capacity of the specific crop.
When the lypophilic antioxidant capacity (LAC) was
evaluated, values ranged between 1.1 and 9.7 µmol
TE g−1 DM. However there was no defined trend
or correlation between LAC and carotenoid content
during the tuber development for any of the genotypes
studied (data not shown). Our previous study9 has
shown that the antioxidant capacity of these crops is
mainly attributed to the phenolic compounds which
are present in higher amount than carotenoids.
Also, significant differences in the total phenolic
contents were found between the 10 mashua cultivars
for each maturity stage (P < 0.05). Among the
purple cultivars, ARB 5241 showed a progressive
phenolics increase during the whole maturity process
whereas AGM 5109 presented almost constant values
and DP 0224 showed a phenolics increase until
6 months followed by a decrease up to 7.5 months
(Fig. 1c). For the yellow cultivars, the phenolic
compounds showed a pattern of slight decrease
during maturation. Genotypic differences could affect
the patterns of phenolics changes in the course of
maturation, between mashua cultivars. Increases of
phenolic compounds30 and decreases31 have been
reported during maturation in different cultivars of
coloured potato tubers. Expressing the antioxidant
capacity (ABTS assay) on a phenolic basis gives
comparative information on the effectiveness of the
phenolics present. A range from 4.6 to 19 µmol TE
mg−1 chlorogenic acid equivalent was obtained in
the present study for purple mashua cultivars which
is higher than those values reported for ripe berry
species25 (7.7 to 14.3 µmol TE mg−1 ), and various ripe
strawberry cultivars25 (11.1 to 12.7 µmol TE mg−1 ).
Significant differences were observed between the
ten mashua cultivars regarding the flavan 3-ol contents
for each maturity stage (P < 0.05) (Fig. 1d). For the
purple cultivars, the changes in flavan 3-ols followed
a similar trend observed for the total phenolics.
The flavan 3-ol values increased progressively for
ARB 5241, leading to a 1.74-fold increase for the
whole maturation period, whereas for DP 0224,
it increased up to 6.5 months (1.48-fold increase),
before declining, and for AGM 5109, it remained
almost constant. On the other hand, increases and
decreases in flavan 3-ols appeared throughout the
tuber development process for the yellow cultivars,
but the values were extremely low. In relative terms,
the ratio of flavan 3-ols to total phenolics represented
442
between 0.26 and 0.34 for purple cultivars and only
from 0.01 to 0.1 for yellow cultivars. Flavan 3-ols
have been investigated in other plant systems because
of their biological properties and pharmacological
potential.32,33 As monomers, they have a very high
potential in quenching reactive oxygen species.34 In
contrast to our results, studies have shown changes
of epicatechin content (flavan 3-ol family) in the flesh
and peel of three apple cultivars to sharply decrease
during the early stage of development followed by
relatively constant values during maturation.35
We found significant differences in the anthocyanin
content between the three purple cultivars at each
maturity stage (P < 0.05). The anthocyanin contents
were consistently higher at 7 and 7.5 months than
at the other maturity stages (Fig. 1e), indicating an
increase in anthocyanin synthesis (∼2.2- to 3.7-fold) at
the later stages of tuber maturation. Similar pattern has
been reported for anthocyanin synthesis in cranberry
varieties,36 bilberry37 and colored potato tubers.30 In
our study the ratio of anthocyanins to total phenolics,
represented between 0.19 and 0.61, at full maturity
(7.5 months), with DP 0224 having the greatest ratio.
As for the other evaluated traits, carotenoid contents
were significantly different between the 10 mashua
cultivars at each maturity stage (P < 0.05) (Fig. 1f).
High values of total carotenoids were always observed
in cultivars with low phenolic contents. A gradual
increase in the carotenoid content was observed during
the tuber development among the yellow cultivars
and the AGM 5109 purple cultivar, the highest
values being obtained at full maturity. Increases in
carotenoids along maturation of the yellow cultivars
ranged from 1.25- to 3.80-fold for ARB 5576 and DP
0215, respectively. The purple cultivars DP 0224 and
ARB 5241 presented almost constant values during
the tuber development.
The correlation analyses between the traits eval-
uated along the tuber development are shown in
Table 1. We found a high correlation between ABTS
and ORAC methods for eight cultivars (0.790 < r <
0.953, P < 0.01), indicating that the bioactive com-
pounds of these cultivars have comparable activities
in the two systems. The flavan 3-ol content was
significantly associated with the antioxidant capac-
ity for the three purple cultivars (DP 0224, ARB
5241 and AGM 5109, r = 0.840, 0.897 and 0.673
respectively, P < 0.01) and for two yellow culti-
vars (DP 0223 and DP 0215, r = 0.624 and 0.884,
P < 0.01). Among the purple cultivars, ARB 5241
was the only one that presented a high correla-
tion between anthocyanins and antioxidant capacity
(r = 0.89, P < 0.01). The poor relationship found
for the other purple cultivars (DP 0224 and AGM
5109), indicate that other phenolic compounds may
have predominant antioxidant effects. A significant
correlation was observed between antioxidant capac-
ity and the total phenolics for cultivars DP 0224,
ARB 5241, AGM 5109, M6COL2C, DP 0215 and
J Sci Food Agric 87:437–446 (2007)
DOI: 10.1002/jsfa

Table 1. Pearson’s correlation coefficientsa of flavan 3-ols, total anthocyanins, total phenolics and antioxidant capacity (ABTS and ORAC) during
tuber development
Cultivar Color, peel/flesh
DP 0224 P/P
ARB 5241 P/Y
AGM 5109 P/Y
ARB 5576 Y/Y
AVM 5562 Y/Y
M6COL2C Y/Y
DP 0203 Y/Y
DP 0223 Y/Y
DP 0207 Y/Y
DP 0215 Y/Y
a, ∗, ∗∗ Significant at P ≤ 0.05 and 0.01, respectively. ACY = anthocyanins; TPH = total phenolics; FV = flavan 3-ols. P = purple, Y = yellow.
DP 0203 (0.691 < r < 0.911, P < 0.01). These differ-
ences in the correlation coefficients suggest important
genotypic differences between cultivars that could be
related to different profiles of antioxidant phenolic
compounds. Only identification and evaluation of
all phenolic components involved in the free radical
scavenging reactions may elucidate the relationship
between phenolic compounds and antioxidant capac-
ity in mashua cultivars and thereby lead to possible
selection based on breeding or molecular approaches.
The process of identification of these compounds is
currently under way and results will be published.
Effect of post-harvest storage
The five traits studied were affected by the sunning
post-harvest storage time (Fig. 2). Significant differ-
ences in antioxidant capacity (ABTS assay), total
phenolics, flavan 3-ols, anthocyanins and carotenoids
were found between the ten mashua cultivars for each
period of sunning post-harvest storage (P < 0.05).
Again, different trends among the 10 cultivars have
been observed. High mean values of antioxidant capac-
ity were always observed in purple cultivars ARB 5241,
DP 0224 and AGM 5109 along the post-harvest stor-
age time (Fig. 2a). Antioxidant capacity of DP 0224
increased in average 1.37-fold at 14 days, followed by a
gradual decrease up to 14% at 35 days while ARB 5241
decreased in 40% and AGM 5109 presented values
almost constant by the end of the storage period. On
the other hand, yellow cultivars with low antioxidant
capacity showed an increase ∼28% during the sunning
storage. The LAC for all the genotypes studied along
the sunning post-harvest storage ranged between 0.8
and 4.3 µmol TE g−1 DM, being these values very low
compared with the hydrophilic antioxidant capacity.
No defined trend and correlation were found between
LAC and carotenoid content (data not shown). The
different changes observed in hydrophilic antioxidant
capacity would be related to the balance between the
new synthesis of bioactive phenolic compounds and
the degradation of others.
For purple tubers DP 0224 and AGM 5109 the
total phenolic content increased between 0 and 14 days
(12% and 64%. respectively) while for ARB 5241 there
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DOI: 10.1002/jsfa
Antioxidants during maturity and storage of Mashua tubers
ORAC:ABTS FV:ABTS ACY:ABTS TPH:ABTS
0.938∗∗ 0.840∗∗ −0.035 0.879∗∗
0.856∗∗ 0.897∗∗ 0.891∗ 0.841∗∗
0.854∗∗ 0.673∗∗ −0.481 0.786∗∗
0.273 −0.155 −0.435
0.405 0.072 0.207
0.794∗∗ −0.584∗ 0.911∗∗
0.812∗∗ −0.191 0.691∗∗
0.894∗∗ 0.624∗∗ −0.3
0.953∗∗ 0.536∗ −0.447
0.860∗∗ 0.884∗∗ 0.797∗∗
was a decrease of 12%. All purple cultivars decreased
in phenolic content after 14 days (Fig. 2b). The total
phenolic content for yellow cultivars increased during
the 35-day storage period. In general, changes in total
phenolic content followed similar trends to that of
antioxidant activity.
With respect to flavan 3-ol contents (Fig. 2c),
increases were observed for purple tubers DP 0224
and AGM 5109 during the first 7 days of sunning
storage (41% and 12%, respectively) while it decreased
for ARB 5241 tubers (∼6%). It has been reported for
other plant systems that flavan 3-ols, such as catechin,
accumulate after pathogen infection or wounding
stress.38,39 In general, all mashua varieties showed
weight loss ranging from 37 to 77%, which can be
considered an additional stress to the tissue (data not
shown). After 7 days, flavan 3-ols decreased gradually
for all purple varieties up to 25% at 35 days. On the
other hand, yellow cultivars presented values almost
constants during the post-harvest storage.
The anthocyanin loss reached an average of ∼70%
at 35 days for the three purple cultivars (Fig. 2d).
However, slight differences in the rate of anthocyanin
degradation between these cultivars were observed
(1.8% day−1 , 2.2% day−1 , 2% day−1 for DP0224,
ARB5241 and AGM5109 respectively). The storage
conditions used in the study (10 h average bright
sunlight and 20 ◦ C, maximum temperature) affected
the rapid decrease in anthocyanin and most likely
the degradation would be related to a photo-
bleaching effect, oxidation,40 temperature41 or a
combination of these. Anthocyanins are considered
to be unstable in storage.41,42 However, in studies
with colored potato during cold storage, an increase in
anthocyanin synthesis was reported.30 Therefore, for
color retention in mashua tubers it would be important
to ensure that storage conditions are optimized for
maximum tuber color.
The total carotenoid contents between 7 and 28 days
were generally higher than at 0 and 35 days. Different
genotypic trends were also observed (Fig. 2e). The
increase in carotenoid content for ARB 5576 could be
related with higher rate in carotenoid synthesis during
the sunning storage compared to other cultivars.
443
R Chirinos et al.

Similarly, an increase in carotenoid content and Changes in glucosinolates concentration during post-
differences in the levels of individual carotenoids was harvest storage has been reported for different mashua
shown in potatoes,43 during tuber storage for 9 months cultivars.11 Furthermore isothiocyanates yielded upon
at 4 ◦ C. enzymatic hydrolysis of the glucosinolates such as
The correlations between antioxidant capacity benzyl-isothiocyanate found in mashua,45 have been
(ABTS assay) and flavan 3-ol, anthocyanin or total shown to have antioxidant capacity as well.46 In
phenolic content during post-harvest storage are general, the results of post-harvest storage of this crop
shown in the Table 2. The correlation between flavan are interesting because the metabolism of bioactive
3-ol and antioxidant capacity during the sunning compounds continues although the tuber is detached
storage was high for the three purple cultivars (DP from the plant (i.e. the nutrient source). One goal in
0224, ARB 5241 and AGM 5109) and for two yellow using post-harvest technology on this crop would be
cultivars (DP 0203 and DP 0215) (0.604 < r < 0.948, to manipulate its metabolism to enhance the health
P < 0.01), while it was variable for the other cultivars. functionality of mashua tubers.13 The habitual time
ARB 5241 again presented a high correlation between of sunlight exposure for consumption of mashua is
antioxidant capacity and anthocyanin (r = 0.867, 5–7 days, thus according to our observations during
P < 0.01), while low correlations were found for the this period there is enough time for induced synthesis
others purple cultivars. The antioxidant capacity and of new phenolic and carotenoid compounds.
total phenolics during the sunning post-harvest storage In general, the high content of total phenolics, flavan
were highly correlated for most cultivars including 3-ols, anthocyanins and carotenoids of some mashua
DP 0224, ARB 5241, AVM 5562, M6COL2c, cultivars indicates that this crop can be considered
DP 0203 and DP 0207 (values 0.639 < r < 0.928, an important source of health-protective antioxidants.
P < 0.01). The low and/or negatively correlation of A harvest time 6 months after being planted may be
the other cultivars could indicate differences in the recommended when mashua cultivars with the highest
ratios and profiles of phenolic compounds and/or antioxidant capacity are required. However, harvest
the presence of other bioactive compounds that may after 7 months may be recommended for increased
contribute to the hydrophilic antioxidant capacity, carotenoid and anthocyanin content in yellow and
including ascorbic acid7 and glucosinolates.11,44 purple genotypes, respectively. A sunning post-harvest
storage higher than 7 days has been shown to be detri-
Table 2. Pearson’s correlation coefficientsa of total anthocyanins, mental for the stability of many of the compounds
flavan 3-ol, total phenolics and antioxidant capacity (ABTS assay) tested except for carotenoids, which presented almost
during the sunning post-harvest storage constant values. Further research is needed in terms of
Color, identification of specific bioactive compounds, as well
Cultivar peel/flesh FV:ABTS ACY:ABTS TPH:ABTS as understanding how pre-harvest and post-harvest
conditions affect the pathway for secondary metabo-
DP 0224 P/P 0.941∗∗ 0.024 0.829∗∗ lites in mashua. Among the 10 cultivars of mashua
ARB 5241 P/Y 0.948∗∗ 0.867∗∗ 0.928∗∗
under study, purple cultivars DP 0224, ARB 5241
AGM 5109 P/Y 0.886∗∗ 0.420 0.459∗
ARB 5576 Y/Y −0.206 0.393
and AGM 5109 (Fig. 3), appear not only to have
AVM 5562 Y/Y −0.314 0.735∗∗ high amounts of phenolic compounds, but also a high
M6COL2C Y/Y 0.127 0.885∗∗ antioxidant capacity or free-radical scavenging proper-
DP 0203 Y/Y 0.761∗∗ 0.690∗∗ ties. This is in agreement with the folk medicine belief
DP 0223 Y/Y 0.005 −0.036 about greater health properties of the dark varieties.
DP 0207 Y/Y −0.025 0.639∗∗
DP 0215 Y/Y 0.604∗∗ 0.438
a, ∗, ∗∗
Significant at P ≤ 0.05 and 0.01, respectively. ACY = antho- ACKNOWLEDGEMENTS
cyanin; TPH = total phenolics; FV = flavan 3-ols. P = purple, Y = The authors gratefully acknowledge Manuel Perez for
yellow. his technical assistance and the personnel from CIP’s

Figure 3. Mashua tubers with high antioxidant capacity and secondary metabolites harvested at 7.5 months, (a) ARB 5241, (b) DP 0224 and
(c) AGM 5109.

444 J Sci Food Agric 87:437–446 (2007)

DOI: 10.1002/jsfa

experimental station for their help in plant cultivation,
harvesting and handling. This research was funded
by USAID through the Association Liaison Office for
University Cooperation in Development (ALO); con-
tract/grant number: HNE-A-00-97-00059-00, Con-
sejo Nacional de Ciencia y Tecnologı́a del Perú
(Concytec) and by the CUI project of the Belgian
Coopération Universitaire au Développement (CUD).
REFERENCES
1 Gibbs PE, Marshall D and Brunton D, Studies on the cytology
of Oxalis tuberosum and Tropaeolum tuberosum. Notes R Bot Gar
Edinb 37:215–220 (1978).
2 Soria SL, Vega R, Damsteegt VD, McDaniel LL, Kitto SL and
Evans TA, Occurrence and partial characterization of a
new mechanically transmissible virus in Mashua from the
Ecuatorian highlands. Plant Dis 82:69–73 (1998).
3 Grau A, Ortega R, Nieto C and Hermann M, Mashua (Tropae-
olum tuberosum Ruı́z & Pav.), Promoting the Conser-
vation and Use of Underutilized and Neglected Crops, 25,
ed. by Engels JMM. International Potato Center, Lima,
Peru/International Plant Genetic Resources Institute, Rome,
Italy. pp. 19–20 (2003).
4 Sperling CR and King SR, Andean tuber crops: Worldwide
potential, in Advances in New Crops, ed. by Janick J and
Simon JE. Timber Press, Portland, OR, pp. 428–435 (1990).
5 National Research Council, Roots and tubers, in Lost Crops of
the Incas: Little Known Plants of the Andes with Promise for
Worldwide Cultivation. National Academy Press, Washington,
DC, pp. 67–73 (1989).
6 Vaughn JG and Geissler CA, The New Oxford Book of Food
Plants. Oxford University Press, Oxford (1997).
7 Collazos C, Alvistur E, Vásquez J, Quiroz A, Herrera N,
Robles N, et al, Tablas peruanas de composición de alimentos,
7th edition. Ministerio de Salud, Instituto Nacional de Salud,
Lima, Peru (in Spanish) (1996).
8 Johns T, Kitts WD, Newsome F and Towers GHN, Anti-
reproductive and other medicinal effects of Tropaeolum
tuberosum. J Ethnopharmacol 5:149–161 (1982).
9 Campos D, Noratto G, Chirinos R, Arbizu C, Roca W and
Cisneros-Zevallos L, Antioxidant capacity and secondary
metabolites in four species of Andean tuber crops: Native
potato (Solanum sp.), Mashua (Tropaeolum tuberosum Ruiz
& Pavon), Oca (Oxalis tuberosa Molina) and Ulluco (Ullucus
tuberosus Caldas). J Sci Food Agric 86:1481–1488 (2006).
10 Noratto G, Cisneros-Zevallos L and Mo H, Tropaeolum tubero-
sum (mashua) extracts suppress tumor cell proliferation.
FASEB JOURNAL 18(5):A886–A886 Suppl. S MAR 24
(2004).
11 Ramallo R, Wathelet JP, le Boulengé E, Torres E, Marlier M,
Ledent JF, et al, Glucosinolates in isaño (Tropaeolum tubero-
sum) tubers: qualitative and quantitative content and changes
after maturity. J Sci Food Agric 84:701–706 (2004).
12 Kalt W, Forney CF, Martin A and Prior RL, Antioxidant
capacity, vitamin C, phenolics, and anthocyanins after fresh
stored of small fruits. J Agric Food Chem 47:4638–4644
(1999).
13 Cisneros-Zevallos L, The use of controlled postharvest abiotic
stresses as a tool for enhancing the nutraceutical content
and adding-value of fresh fruits and vegetables. J Food Sci
68:1560–1565 (2003).
14 Arnao M, Cano A and Acosta M, The hydrophilic and
lipophilic contribution to total antioxidant activity. Food Chem
73:239–244 (2001).
15 Awika JM, Rooney LW, Wu X, Prior RL and Cisneros-
Zevallos L, Screening methods to measure antioxidant activity
of sorghum (Sorghum bicolor) and sorghum products. J Agric
Food Chem 51:6657–6662 (2003).
J Sci Food Agric 87:437–446 (2007)
DOI: 10.1002/jsfa
Antioxidants during maturity and storage of Mashua tubers
16 Ou B, Hampsch-Woodill M and Prior RL, Development and
validation of an improved oxygen radical absorbance capacity
assay using fluorescein as the fluorescent probe. J Agric Food
Chem 49:4619–4626 (2001).
17 Huang D, Ou B, Hampsch-Woodill M, Flanagan JA and Prior
RL, High-throughput assay of oxygen radical absorbance
capacity (ORAC) using a multichannel liquid handling system
coupled with a microplate fluorescence reader in 96-well
format. J Agric Food Chem 50:4437–4444 (2002).
18 Cao GH and Prior RL, Measurement of oxygen radical
absorbance capacity in biological samples. Oxidants and
antioxidants. Methods Enzymol 299:50–62 (1999).
19 Swain T and Hillis W, The phenolic constituents of Prunus
domestica. I. The Quantitative analysis of phenolic con-
stituents. J Sci Food Agric 10:63–68 (1959).
20 Giusti MM and Wolrstad RE, Anthocyanins. Characterization
and measurement with UV–visible spectroscopy, in Current
Protocols in Food Analytical Chemistry, ed. by Wrolstad RE.
John Wiley, New York (2001).
21 Abdel-Aal ES, A rapid method for quantifying total antho-
cyanins in blue aleurone and purple pericarp wheats. Cereal
Chem 76:350–354 (1999).
22 Delcour JA and de Varebeke DJ, A new colorimetric assay for
flavonoids in pilsner beers. J Inst Brew 91:37–40 (1985).
23 McMurrough I and McDowell J, Chromatrographic separation
and automated analysis of flavonols. Anal Biochem 91:92–100
(1978).
24 Talcott S and Howard R, Phenolic autooxidation is responsible
for color degradation in processed carrot puree. J Agric Food
Chem 47:2109–2115 (1999).
25 Wang S and Lin HS, Antioxidant activity in fruits and leaves of
blackberry, raspberry, and strawberry varies with cultivar and
developmental stage. J Agric Food Chem 48:140–146 (2000).
26 Cevallos-Casals BA and Cisneros-Zevallos L, Stoichiometric
and kinetic studies of phenolic antioxidants from andean
purple corn and red-fleshed sweet potato. J Agric Food Chem
51:3313–3319 (2003).
27 Breithaupt DE and Bamedi A, Carotenoids and carotenoid
esters in potatoes (Solanum tuberosum L.): New insights into an
ancient vegetable. J Agric Food Chem 50:7175–7181 (2002).
28 Iwanzik W, Tevini M, Stute R and Hilbert R, Carotinoigehalt
und -zusammensetzung verschiedener deutscher Kartoffel-
sorten un deren Bedeutung für die Fleischfarbe der Knolle.
Potato Res 26:149–162 (1983).
29 Prior RL, Cao G, Martı́n A, Sofic E, McEwen J, O’Brien C,
et al, Antioxidant capacity as influenced by total phenolic
and anthocyanin content, maturity, and variety of Vaccinium
species. J Agric Food Chem 46:2686–2693 (1998).
30 Lewis CE, Walker JRL and Lancaster JE, Changes in antho-
cyanins, flavonoid and phenolic acid concentrations during
development and storage of coloured potato (Solanum tubero-
sum L) tubers. J Sci Food Agric 79:311–316 (1999).
31 Reyes LF, Miller Jr JC and Cisneros-Zevallos L, Environmental
conditions influence the content and yield of anthocyanins
and total phenolics in purple- and red-flesh potatoes during
tuber development. Am J Potato Res 81:187–193 (2004).
32 Plumb GW, de Pascual-Teresa S, Santos-Buelga C, Cheynier V
and Willianson G, Antioxidant properties of catechins and
proanthocyanidins: Effect of polymerization, galloylation and
glycosylation. Free Radical Res 29:351–358 (1998).
33 Matsuda H, Chisaka T, Kubomura Y, Yamahara J, Sawaka T
and Fujimura H, Effects of crude drugs on experimen-
tal hipercolesterolemia. I. Tea and its active principes.
J Ethnopharmacol 17:213–224 (1986).
34 Rice-Evans CA, Miller NJ and Paganga G, Antioxidant prop-
erties of phenolic compounds. Trends Plant Sci 2:152–159
(1997).
35 Burda S, Oleszek W and Lee CY, Phenolic compounds and
their changes in apples during maturation. J Agric Food Chem
38:945–948 (1999).
36 Vvedenskaya IO and Vorsa N, Flavonoid composition over
fruit development and maturation in American cranberry,
Vaccinium macrocarpon Ait. Plant Sci 167:1043–1054 (2004).
445
R Chirinos et al.
37 Jaakola L, Määta K, Pirttilä AM, Törrönen R, Kärenlampi S
and Hohtola A, Expression of genes involved in anthocyanin
biosynthesis in relation to anthocyanin, proanthocyanidin,
and flavanol levels during bilberry fruit development. Plant
Physiol 130:729–739 (2002).
38 Buschmann H, Reilly K, Rodriguez MX, Tohme J and Beech-
ing JR, Hydrogen peroxide and flavan-3-ols in storage roots of
cassava (Manithot esculenta Crantz) during postharvest deteri-
oration. J Agric Food Chem 48:5522–5529 (2000).
39 Berryman AA and Christiansen E, Induced responses in
phenolic metabolism in 2 Norway spruce clones after
wounding and inoculations with Ophiostoma polonicum, a bark
beetle-associated fungus. Plant Physiol 109:821–827 (1995).
40 Francis FJ, Food colorants: anthocyanins Crit Rev Food Sci Nutr
28:273–314 (1989).
41 Hall IV and Stark R, Anthocyanin production in cranberry
leaves and fruit, related to cool temperatures at a low light
intensity. Hortic Res 12:183–186 (1972).
446
42 Van Buren JP, Bertino JJ and Robinson WB, The stability of
wine anthocyanins on exposure to heat and light. Am J Enol
Vitic 19:147 (1968).
43 Morris WL, Ducreux L, Griffiths DW, Stewart D, Davies
HV and Taylor MA, Carotenogenesis during tuber
development and storage in potato. J Exp Bot 55:975–982
(2004).
44 Guimares RL, Momotani K and Flores HE, Study of glucosi-
nolates from mashua (Tropaeolum tuberosum). Annual meeting
Plant Biology Abstracts (2000). URL http://rycomusa.com/
aspp2000/public/P43/0904.html [accessed 15 December
2003].
45 Johns T and Towers GHN, Isothiocyanates and thioureas in
enzyme hydrolysates of Tropaeolum tuberosum. Phytochemistry
20:2687–2689 (1981).
46 Rao BN, Bioactive phytochemicals in Indian foods and their
potential in health promotion and disease prevention. Asia
Pacif J Clin Nutr 12:9–22 (2003).
J Sci Food Agric 87:437–446 (2007)
DOI: 10.1002/jsfa