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Food Chemistry 139 (2013) 578588

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Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Ellagic acid derivatives, ellagitannins, proanthocyanidins and other phenolics,


vitamin C and antioxidant capacity of two powder products from camu-camu
fruit (Myrciaria dubia)
Daniela Fracassetti a,1, Carlos Costa a, Leila Moulay b, Francisco A. Toms-Barbern a,
a
b

Quality, Safety and Bioactivity of Plant Foods, CEBAS-CSIC, P.O. Box 164, Espinardo, 30100 Murcia, Spain
Agrcola San Juan de la Amazonia Europa (ASJAEU), Valencia, Spain

a r t i c l e

i n f o

Article history:
Received 17 September 2012
Received in revised form 19 January 2013
Accepted 21 January 2013
Available online 13 February 2013
Keywords:
Camu-camu
Myrciaria dubia
Phenolics
Ellagitannins
Ellagic acid conjugates
Proanthocyanidins
Antioxidant capacity
Vitamin C
Dehydrated powders

a b s t r a c t
The aims of this study were the evaluation of polyphenols and vitamin C content, and antioxidant capacity of dehydrated pulp powder and the dried our obtained from the skin and seeds residue remaining
after pulp preparation from camu-camu (Myrciaria dudia). Fifty-three different phenolics were characterised by HPLCDADESI-MSMS and UPLC-HR-QTOF-MS-MS. The phenolic content of camu-camu our
was higher than that of the pulp powder (4007.95 mg/100 g vs. 48.54 mg/100 g). In both products the
avonol myricetin and conjugates, ellagic acid and conjugates and ellagitannins were detected. Cyanidin
3-glucoside, and quercetin and its glycosides were only found in the pulp powder, while proanthocyanidins were only present in the our (3.5 g/100 g, mean degree of polymerisation 3). The vitamin C content
was lower in pulp powder (3.5%) than in the our (9.1%). The radical-scavenging capacity of both
powders was determined by the DPPH, ABTS and ORAC assays, and was higher for camu-camu our as
could be expected for its higher phenolics and vitamin C content. Comparative analyses with fresh
camu-camu berries indicate that some transformations occur during processing. Analysis of fresh
berries showed that ellagic acid derivatives and ellagitannins were mainly present in the seeds, while
proanthocyanidins were present both in the seeds and skin.
2013 Elsevier Ltd. All rights reserved.

1. Introduction
Consumption of fruits and vegetables is known to lower the risk
of several diseases, including cardiovascular diseases, cancer and
stroke (Willett, 2002) and such health benets are mainly attributed to the content in antioxidant compounds and particularly
vitamin C and phytochemicals such as polyphenols and carotenoids (Steinmetz & Potter, 1996).
Among fruits, berries are reported to exhibit many benecial effects in human health (Seeram, 2010; Seeram, 2012). This is well
documented by several studies and has been the focus of much
current research on chemoprevention of cardiovascular diseases
(Basu, Rhone, & Lyons, 2010). As well as being a good source of
vitamin C, dietary bre, and minerals, berries contain high levels
of natural polyphenol components that act as potent antioxidants.
Among the berries, those produced by the genus Myrciaria have
received attention recently due to their high content in antioxidants including vitamin C and polyphenols. Thus Myrciaria dubia
Corresponding author. Tel.: +34 968396334; fax: +34 968396213.
E-mail address: fatomas@cebas.csic.es (F.A. Toms-Barbern).
Address: Department of Food, Environmental and Nutritional Sciences (DeFENS),
Universit degli Studi diMilano, Via Celoria, 2 20133 Milano, Italy.
1

0308-8146/$ - see front matter 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.foodchem.2013.01.121

(camu-camu) (DaSilva et al., 2012), Myrciaria jaboticaba (Leite


et al., 2011; Wu, Dastmalchi, Long, & Kennelly, 2012) and Myrciaria
vexator (Dastmalchi et al., 2012) fruits have been recently studied.
An increase of plasma antioxidant potential of rats after the intake
of freeze-dried jaboticaba peel has recently been demonstrated
(Leite et al., 2011).
Camu-camu is a native Amazonian bush from the Myrtaceae
family. Its fruits are round berries having an average diameter of
2.5 cm; its pulp is pink, while its skin is green when immature
and changes during the ripening process from green to red-purple
due to the presence of anthocyanins (Zanatta & Mercadante, 2005).
Camu-camu is appreciated for its high content of ascorbic acid,
which varies from 1.9 to 2.3 g/100 g fresh matter depending
on the maturity stage (Chirinos, Galarza, Betalleluz-Pallardel,
Pedresch, & Campos, 2010). Compared to other fruits, camu-camu
is considered one of the richest sources of vitamin C, with a higher
content than acerola (Runo et al., 2010).
Camu camu fruits have a signicant use history as edible and as
traditional medicines with different ethnobotanical uses throughout the tropical and subtropical world (Flores, 1998).
In addition to vitamin C, several studies have reported that
camu-camu is a good source of bioactive phytochemicals. Different
concentrations of avan-3-ols, avonols, avanones, gallic acid and

D. Fracassetti et al. / Food Chemistry 139 (2013) 578588

ellagic acid were also detected (Chirinos et al., 2010; De Souza


Schmidt Gonalves, Lajolo, & Genovese, 2010; Genovese, Da Silva
Pinto, De Souza Schmidt Gonalves, & Lajolo, 2008; Reynertson,
Yang, Jiang, Basile, & Kennelly, 2008; Runo et al., 2010), although
no complete study with a detailed characterisation of the main
phenolic compounds (proanthocyanidins, ellagitannins and ellagic
acid conjugates) has been reported to date.
Research on phenolic compounds and health has been a focus of
interest in the last decade due to their antioxidant activity and free
radical-scavenging ability (Tomas-Barberan & Andres-Lacueva,
2012). In particular, the polyphenols seem to be involved in
several benecial effects in human health (Ross & Kasum, 2002;
Tomas-Barberan & Andres-Lacueva, 2012; Traka & Mitchen,
2011). The results obtained so far suggest the potential application
and positive biological effects of camu-camu berries and derived
food products in human health (Inoue, Komoda, Uchida, & Node,
2008; Yazawa, Suga, Homna, Shirosaki, & Koyama, 2010), although
human intervention studies are necessary.
Camu-camu consumption as fresh fruit is limited due to its high
acidity and bitterness (Flores, 1998). It is mainly consumed as
juice, or as an ingredient for jellies, ice-creams, liquors, wines or
other foods (Cavalieri, 1993; Franco & Janzantti, 2005; Villachica,
1997). Its commercial interest has particularly increased due to
its high vitamin C content. However, due to the loss of this vitamin
during postharvest storage and processing, alternative technological processes need to be developed to preserve the nutritional value of camu-camu berries. Water is the main component of the
fruits and has a direct implication on quality loss through its effect
on many physicochemical and biological attributes. Therefore, the
dehydration process has been suggested as an alternative to obtain
camu-camu ingredients that in powdered form can be used to enhance vitamin C and bioactive compounds of different food products (Rodrigues et al., 2004).
Few studies have reported a thorough chemical composition,
particularly phenolic compounds characterisation, of camu-camu
fruit. The phenolics content has been often evaluated as total polyphenols in the fruits, and the main components previously identied were ellagic acid, quercetin, rutin and gallic acid (Chirinos
et al., 2010; De Souza Schmidt Gonalves, Lajolo, & Genovese,
2010; Reynertson et al., 2008; Runo et al., 2010). No information
has previously been reported either on the proanthocyanidin or
ellagitannin composition of camu-camu fruit or the powder products obtained from the berry pulp and the remaining skins and
seeds by dehydration processes.
Therefore, this study aims to the chemical characterisation of
two different powders obtained from camu-camu fruit, one from
the pulp and the second one from the remaining peel, seeds and
adhered pulp (our). Our main objective was to characterise the
phenolic compounds, quantify the vitamin C and determine the
antioxidant capacity due to the nutritional potential which they
might exhibit.

2. Materials and methods


2.1. Chemicals
Standards of gallic acid, ellagic acid, quercetin, myricetin, rutin,
catechin, and cyanidin 3-O-glucoside (Cy-glc), sodium acetate,
potassium phosphate, o-phenylene diamine (OPDA) manganese
dioxide, sodium uoride, ethylenediaminetetraacetic acid (EDTA),
2,20 -azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS),
2,2 0 -azobis(2-amidino-propane)dihydrochloride (AAPH), and
2,2-diphenyl-1-picrylhydrazyl (DPPH) were purchased from Sigma
Aldrich (Darmstadt, Germany). Phloroglucinol, ascorbic acid,
dehydro-ascorbic acid, potassium chloride, hydrochloric acid,

579

methanol, acetonitrile, acetic acid, and formic acid were from


Merck (Darmstadt, Germany). Vescalagin was used as external
standard for ellagitannin quantication and was kindly provided
by Prof. Stephan Quideau (University of Bordeaux, France). Water
was obtained from Milli-Q apparatus (Millipore, Milford, MA, US).
2.2. Samples
Two powders from camu-camu fruit were provided by Agricola
San Juan de la Amazonia Europa (ASJAEU) (Valencia, Spain). Both
powders were obtained from mature turning colour maturity state
Myrciaria dubia fruits harvested during the summer 2010 in Pucallpa Ucayali (Peru). One powder was produced from the pulp
(pulp powder), dried in a spray drier at an inlet temperature of
185 C and an outlet temperature of 95 C with addition of 10%
maltodextrin to decrease the stickiness of the resulting powder.
The second powder (our) was produced from the remaining peel
and seeds with adhered pulp after pulp extraction, and was dried
in a uid bed drier at a temperature of 4555 C. Therefore two different drying processes were applied to obtain these powders and
this was due to the different water content of the raw materials
which is generally around 90% for the pulp fraction and about
60% for the remaining skin/seeds fraction. Both powders were
packaged under vacuum and stored at 4 C until used. Unprocessed
fresh camu-camu berries were also collected and stored frozen
(20 C) for comparative phenolic compounds analyses. Fruit skins
were 24.2% of the total fruit weight (fresh weight) and had a water
content of 81.9%; the pulp weighed 43.6% of the total fruit with a
water content of 92%; the seeds constituted 32.2% of the fruit
weight (fresh weight) and had a water content of 62%.
2.3. Determination of phenolic compounds
The extraction of phenolics was performed as follows: 1 g pulp
powder was added to 25 mL of 50% methanol in water acidied
with1% formic acid; 1 g camu-camu our was added to 25 mL of
50% methanol in water. Different extractions were carried out in
order to achieve the better phenols recovery using variable ratio
of water and methanol, with and without formic acid. The use of
50% methanol in water acidied with 1% formic acid and methanol
50% for pulp powder and camu-camu our, respectively, allowed
the better recovery of phenols. The powders were vortexed for
2 min, sonicated for 15 min (Sonicator Branson 5510, Emerson,
Danbury, CT, US) and centrifuged at 3000 rpm for 15 min at 4 C
(Centrifuge 5804 R, Eppendorf, Hamburg, Germany). The supernatants were recovered, freeze-dried under vacuum, suspended in
2 mL of the corresponding extraction solvent, then ltered with a
PVDF lter 0.22 lm (Millipore, Billerica, MA, US) and injected in
LC/MS.
The identication and quantication of phenols were performed using an Agilent 1100 Series equipment (Agilent, Santa
Clara, CA, US) equipped with G1312A binary pump, G1313A autosampler, G1315B photodiode array detector, and G1322A degasser
controlled by the Agilent software v. A08.03. HPLC was coupled
with a detector MSD Trap 1100 Series (Agilent) with an electrospray ionisation system (ESI), with the following conditions: the
heated capillary was 350 C and 33.5 kV voltage, mass scan
(MS) and MS/MS were measured from 100 to 1500 m/z. Collisioninduced fragmentation experiments were performed in the ion
trap using helium as the collision gas, and the collision energy
was set at 75%. Mass spectrometry data were acquired in the negative ionisation mode. A column Pursuit XRs C18 250  40 mm
from Varian (Agilent, Santa Clara, CA, US) was used and a ow rate
of 0.8 mL min1. The used solvents were 1% formic acid in water
(A) and acetonitrile (B) which was in the following separation gradient: 1% B in A at 0 min, 9% B at 10 min, 35% B at 48 min, and 95%

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D. Fracassetti et al. / Food Chemistry 139 (2013) 578588

B at 52 min, following by washing and conditioning steps. Data


were registered from 250 nm to 700 nm and the phenolic compounds were quantied at 280 mn, 360 mn, and 520 nm, depending on the type of phenolic compound. Integrations were
performed by Agilent ChemStation for LC 3D, Rev. B.01.03 SR1.
MS trap control was carried out Bruker Daltonic version 5.2.
Quantication of gallic acid, ellagic acid, myricetin and
their derivatives, and ellagitannins was carried out with the
calibration curves obtained for gallic acid (1300 mg L1), ellagic
acid (1300 mg L 1 ), rutin (1300 mg L 1 ), and vescalagin
(0.1100 mg L1), respectively, at the appropriate wavelengths.
All the samples and standards were injected in triplicate.
Moreover, samples of pulp powder and our were analysed by
UPLC-Q-TOF (Agilent) in order to further conrm the phenolic
compounds identied by MS Trap. The Q-TOF equipment had
the following conditions: ESI gas temperature 280 C, drying gas
9 l/min, nebulizer 35 psig, sheath gas temp 400 C, sheath gas ow
12 l/min. MS TOF fragmentor 100 V, mas range 1001500, negative
mode. The column was Poroshell 120, EC-C18, 2.7 lm,
30  100 mm (Agilent); the eluents were 0.1% formic acid in water
(A) and acetonitrile acidied with 0.1% formic acid (B). The separation gradient started with 1% B in A at 0 min, 9% at 3 min, 48% at
20 min, and 95% at 23 min, following by washing and conditioning
steps. The volume injected was 2 lL and the ow rate was 0.4 mL/
min. The determinations were carried out in triplicate.
2.4. Analysis of proanthocyanidins
Proanthocyanidins were quantied as previously reported by
Kennedy and Jones (2001) using an acid catalysis in the presence
of phloroglucinol. Briey, 50 mg sample were dissolved in 800 lL
of phloroglucinol (50 mg mL1) added with ascorbic acid
(10 mg mL1) dissolved in methanol acidied with 0.1 N HCl. The
reaction mix was vortexed and incubated at 50 C for 20 min.
The reaction tube was placed in ice and 1 mL 40 mM sodium acetate was added in order to stop the reaction. The sample was centrifuged, ltered with a 0.22 lm PVDF lter, and injected in LC/MS.
The identication and quantication of catechin, epicatechin and
their derivatives was carried out by Agilent 1100 Series apparatus
equipped with detector MSD Trap 1100 Series (Agilent), as previously described (Buenda et al., 2010; Vallejo, Marn, & TomsBarbern, 2012). Briey, the column used was an Atlantis C18
(250 mm  4.6 mm, 5 lm particle size; Water, Milford, MA, US)
operating at a ow rate of 1 mL min1; the injection volume was
10 lL. The solvents were 2.5% acetic acid in water (A) and acetonitrile (B) with a separation gradient starting with 3% B in A at 0 min,
9% at 5 min, 16% at 15 min, 50% at 45 min followed by washing and
conditioning steps. The phenolic compounds were quantied at
280 nm with a calibration curve of catechin (1300 mg L1). The
MS detector operated in negative ion-mode. The Trap interface
and ion optics settings were the following: spray potential
65 psi; nebulization gas (nitrogen) relative ow value 11; capillary
temperature 325 C. Full-scan mass spectra were acquired scanning the range 100800 m/z.
2.5. Determination of ascorbic and dehydroascorbic acids
Quantication of ascorbic acid and dehydroascorbic acid was
carried out as previously described by Zapata and Dufour (1992).
Briey, 50 mg sample were dissolved in 10 ml of 5% methanol
added with citric acid (21 g L1) and EDTA (0.5 g L1). The homogenate was ltered through a 0.45 lm PVDF lter and puried on a
C18 Sep-Pak cartridge (Waters, Mil-ford, MA, US). The HPLC analysis was achieved after derivatization of DHAA into the uorophore
3-(1,2-dihydroxyethyl) furol [3,4-b]quinoxaline-1-one (DFQ), with
1,2-phenylenediamine dihydrochloride (OPDA). Standard solu-

tions, column conditioning, mobile phase, ow rate, wavelengths


and derivatization procedures used were previously reported by
Gil, Ferreres, and Toms-Barbern (1999), and Martnez-Snchez,
Tudela, Luna, Allende, and Gil (2011). The results were expressed
as g ascorbic acid and dehydroascorbic acid per 100 g powder.
2.6. Antioxidant capacity assays
The antioxidant capacity of both powders was carried out
through three different methods measuring the free radical scavenging capacity (DPPH, ABTS and ORAC assays).
The free radical scavenging activity determined with DPPH followed the method of Brand-Williams, Cuvelier, and Berset (1995)
with some modications (Espn, Soler-Rivas, Wichers, & GarcaViguera, 2000; Llorach, Toms-Barbern, & Ferreres, 2004). The
DPPH solution was diluted with methanol to an absorbance of
1.00 (0.03) at 515 nm. In a 96-wells micro plate (Nunc, Roskilde,
Denmark), 250 lL of DPPH solution were placed in each well and
2 lL sample were added. The sample was dissolved in 70% methanol (20 g L1) and, after centrifugation, it was serially diluted.
The ABTS method was performed as reported by Mena et al.
(2011). The ABTS solution was diluted with water to an absorbance
of 1.00 (0.03) at 414 nm. In a 96-wells micro plate (Nunc,
Roskilde), 250 lL of ABTS solution were put in each well and
2 lL sample were added. The sample was dissolved in water
(20 g L1) and, after centrifugation, it was serially diluted. For both
assays, the reaction kinetic was monitored for 50 min at 25 C by
micro plate reader (Innite M200, Tecan, Grdig, Austria). A
calibration curve was made by adding increasing concentration
of Trolox ranged from 50 to 1000 lM; each concentration was
assayed in quadruplicate, as well each sample.
The free radical scavenging activity determined by the ORAC assay followed the method reported by Prior et al. (2003) with some
modications. In a 96-wells micro plate, 100 lL 14 lM uorescein
prepared in 75 mM phosphate buffer pH 7.4, 20 lL sample (or
standard) and 30 lL 75 mM phosphate buffer pH 7.4 were placed
into each well. After 15 min incubation at 37 C, 30 lL AAPH
(21.6 mg/mL) were added. Readings were carried out with a uorescent microplate reader (Multi-Detection Microplate Reader,
Synergy HT, Biotek Instruments USA) which was programmed
to read the uorescence with an excitation wavelength of
485 nm and an emission wavelength of 528 nm at 1 min interval
for 90 min using software Gen 5. The calibration curve was obtained with increasing concentrations of Trolox prepared in
75 mM phosphate buffer pH 7.4, ranging from 5 to 50 lM.
The results were expressed as lM of Trolox per gram of sample
(dry matter). Each concentration was assayed in quadruplicate, as
well as each sample.
3. Results and discussion
3.1. Phenolics characterisation
The phenolic compounds characterised in camu-camu berries
and powders are shown in Table 1 and Fig. 1AC. The UV spectra
of the different compounds recorded with a Diode Array Detector
(DAD) showed that avonols, ellagic acid conjugates, ellagitannins
and proanthocyanidins were the main polyphenols present. One
anthocyanin was detected in pulp powder, and at least two
hydroxycinnamic acid derivatives, which showed the characteristic UV/Vis spectrum of a caffeic acid derivative (maximum at
332 nm, and shoulder at 298 nm and m/z 353). Their molecular
weights, however, could not be clearly established from the
HPLCMS analyses. The different compounds were characterised
by their UV spectra, their molecular ions and fragments obtained

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D. Fracassetti et al. / Food Chemistry 139 (2013) 578588


Table 1
HPLCDADESI-MSMS analysis of phenolic compounds in camu-camu powders.
[MH]

Number

Compound

Retention time (min)

Flavonols
4
6
6
26
27
28

Myricetin 3-O-hexoside
Myricetin 3-O-pentoside
Myricetin 3-O-pentoside
Quercetin 3-O-hexoside
Quercetin 3-O-pentoside
Myricetin

27.5
29.2
30.4
31.7
35.5
38.3

479
449
449
463
433
317

Anthocyanins
29

Cyanidin 3-O-glucoside

kmax (nm)
264,
258,
272,
256,
256,
256,

MS fragments

358
356
356
362
352
374

316, 221, 179


317
316
417, 301
301, 179, 151
317, 179, 151

520

285

374
362
360
364
368
362
362
360
362
362

425
301
301
300
229
301
301
415, 301
301
301

20.6

447

Ellagic acid derivatives


1
Valoneic acid dilactone
2
Ellagic acid hexoside
3
Ellagic acid pentoside
5
Ellagic acid desoxyhexoside
7
Ellagic acid
8
Ellagic acetyl rhamnoside
9
Ellagic acetyl rhamnoside
10
Ellagic acid derivative
12
Ellagic acid derivative
13
Ellagic acid derivative

15.4
24.7
28.9
29.7
30.9
35.9
36.5
37.2
40.8
41.5

469
463
433
447
301
489
489
585
719
719

Ellagitannins
15
16
48
18
47
19
20
21
23
24

11.5
13.9
15.3
16.2
19.5
19.8
20.5
21.8
25.9
27.9

933
933
633
784
935
633
935
785
935
937

240,
240,
240,
240,
240,
240,

246
246
270
240
270
270
270
272
270
276

915, 889, 631


915, 889, 631
463, 301
481, 301
917, 633, 301
463, 301
917, 633, 301
484, 301
917, 633, 301
767, 741, 465, 301

Gallic acid derivatives


14
Gallic acid
25
Gallic acid derivative

9.0
37.8

169
569

274
238, 274

125
551, 523, 169

Proanthocyanidins
17
Gallocatechin-gallocatechin gallate-gallocatechingallate
46
Gallocatechin-gallate
22
Gallocatechin-gallate-dimer

15.8
24.0
24.1

1221
457
915

240,274sh
240, 274
240, 274

915
331,305,169
457, 169

Vescalagin
Castalagin
HHDP-galloyl-glucose
Di-HHDP-glucose (pedunculagin)
Di-HHDP-galloyl-glucose (casuarictin/potentillin)
HHDP-galloyl-glucose
Di-HHDP-galloyl-glucose (casuarictin/potentillin)
Digalloyl-HHDP-glucose
Di-HHDP-galloyl-glucose (casuarictin)
Tri-galloyl-HHDP-glucose (tellimagrandin II)

255,
255,
254,
254,
256,
254,
254,
254,
254,
254,

240,

Bold values are the main fragments.

with an ESI-MSMS detector (Table 1) and comparison, wherever


possible, with authentic markers. In addition, the different compounds were conrmed using a high resolution analysis on a
UPLC-Q-TOF equipment in which the molecular formulae were calculated (Table 2). All identied compounds showed a low error
(3.30) and a high score (better than 92.15) that indicated the
accuracy of the exact mass and the molecular formulae obtained.
The avonols myricetin (3,5,7,30 ,40 ,50 -hexahydroxyavone) (28),
its 3-O-hexoside (4) (probably glucoside) and two 3-O-pentoside
isomers (6) (probably arabinoside and xyloside), were detected.
They showed characteristic UV spectra of avonols with a free hydroxyl at 3 position for myricetin (UV band I maximum at 374 nm),
and a blocked hydroxyl at 3 for the O-glycosides (UV band I maximum at 356 nm) (Table 1). The pseudomolecular ions recorded
with the HPLC-ESI MS and the fragments obtained conrmed these
structures with the characteristic losses of a hexosyl and a pentosyl
residue, respectively, leading to the myricetin aglycone fragment at
m/z 317. The High-resolution Mass Spectrometry Analysis conrmed these structures (Table 2). In addition two quercetin
(3,5,7,30 ,40 -pentahydroxyavone) 3-O-glycosides were detected,
quercetin 3-O-hexoside (26) and 3-O-pentoside (27) and these
two were only detected in the pulp powder. Rutin (quercetin
3-O-rutinoside) that was previously reported in camu-camu
(Reynertson et al., 2008), was not detected in the analysed
powders or in the fresh berry fruits. In addition, the avanones
naringenin and eriodictyol, that had previously been reported
(Akter, Oh, Eun, & Ahmed, 2011) were not detected here either.

The anthocyanins were easily detected with UVVis detector set


at 520 nm. Only one pigment was detected in the pulp powder and
our, and its UVVis spectrum with a maximum at 520 nm suggested that this was a cyanidin derivative, and this was conrmed
by the MSMS analysis that showed that this was a cyanidin
hexoside. This is most probably Cyanidin 3-O-glucoside (29) that
was previously reported in fresh camu-camu fruits (Zanatta &
Mercadante, 2005). The structure was conrmed by high-resolution
Q-TOF MSMS (Table 2). No delphinidin 3-O-glucoside was
detected in the powders, although this was present in the analysed
fresh berries, both in the skin and the pulp (Table 3), and had been
previously reported in camu-camu fruits (Zanatta & Mercadante,
2005). This is not unexpected as delphinidin is more susceptible
than cyanidin to oxidative and thermal degradation.
A number of compounds with the characteristic UV spectrum
ellagic acid were detected (Table 1). The main one was free ellagic
acid (7) that showed a pseudomolecular ion at m/z 301 and coincided chromatographically with an authentic standard of this polyphenol. Nine other different compounds showed a UV spectrum
similar to that of ellagic acid and all of them but compound 1 produced a m/z 301 fragment in the MS analysis conrming that they
were ellagic acid conjugates. Compound 1 showed a pseudomeolecular ion at m/z 469 and a fragment at m/z 425 that are characteristic of valoneic acid bilactone, a compound that often occurs
in plants containing ellagitannins. The conjugates included an
O-hexoside (2) (most probably glucoside), an O-pentoside (3)
(most probably arabinoside or xyloside), and an O-deoxyhexoside

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D. Fracassetti et al. / Food Chemistry 139 (2013) 578588


DAD1 C, Sig=360,16 Ref=off (Y:\CAMU\05-05\11051203.D)
mAU

60

50

40

30

20

10

10

15

20

25

30

35

40

min

25

30

35

40

min

DAD1 C, Sig=360,16 Ref=off (Y:\CAMU\05-05\04051214.D)


mAU

350

300

250

200

150

100

4.9 60

50

0
5

10

15

20

DAD1 B, Sig=280,16 Ref=off (Y:\CAMU\05-05\04051214.D)


mAU

1200

1000

800

600

400

200

0
5

10

15

20

25

30

35

40

min

Fig. 1. HPLC analyses of phenolic compounds from camu-camu pulp powder at 360 nm (A), camu-camu our at 360 nm (B) and at 280 nm (C). : unidentied compounds. For
compound identication see Table 1. (D) HPLCDAD chromatogram (280 nm) of camu-camu our proanthocyanidins analysed after acid catalysed degradation in the
presence of phloroglucinol. For compound identication see Table 4.

583

D. Fracassetti et al. / Food Chemistry 139 (2013) 578588


Table 2
High-resolution UPLC-Q-TOF characterisation of phenolic compounds in camu-camu powders.
Number

Compound

Exact mass

Exact mass theorical

Error

Score

Retention time (min)

Formula

Flavonols
4
6
6
6
26
27
28

Myricetin 3-O-hexoside
Myricetin 3-O-pentoside
Myricetin 3-O-pentoside
Myricetin 3-O pentoside
Quercetin 3-O-hexoside
Quercitin 3-O-pentoside
Myricetin

479.0842
449.0734
449.0734
449.0735
463.0890
433.0787
317.0314

479.0831
449.0725
449.0725
449.0725
463.0882
433.0776
317.0303

2.85
2.32
2.37
0.20
2.18
2.35
3.59

91.42
96.00
95.17
94.43
96.46
97.02
95.54

8.2
8.6
9.0
9.1
9.2
10.3
11.4

C21H20O13
C20H18O12
C20H18O12
C20H18O12
C21H20O12
C20H18O11
C15H10O8

447.0944

447.0933

2.63

95.31

6.2

C21H21O11

469.0053
463.0533
433.0423
447.0593
301.0000
489.0679
489.0678
585.0526
719.2199
719.2206

469.0049
463.0518
433.0412
447.0569
300.9990
489.0675
489.0675
585.0522
921.2193
719.2193

0.95
0.42
2.53
4.62
3.42
0.76
0.68
0.97
0.94
1.20

99.06
98.59
96.75
85.05
96.08
99.15
98.76
98.17
96.34
96.14

5.0
7.3
8.4
8.7
9.1
10.8
10.9
11.2
12.7
12.8

C21H9O13
C20H16O13
C19H14O12
C20H16O12
C14H6O8
C22H18O13
C22H18O13
C26H18O16
C34H39O17
C34H40O17

933.0645
933.0640
633.0746
783.0695
935.0794
633.0747
935.0801
785.0857
935.0797
937.0945

933.0640
933.0640
633.0733
783.0686
935.0796
633.0733
935.0796
785.0843
935.0796
937.0953

0.33
0.31
3.35
0.22
0.36
3.10
0.96
2.06
0.00
0.45

98.45
99.68
93.54
98.40
95.48
85.20
97.97
95.92
99.93
99.37

3.9
4.6
4.8
5.0
5.1
5.2
6.4
6.7
8.0
8.5

C41H26O26
C41H26O26
C27H22O18
C34H24O22
C41H28O26
C27H22O18
C41H28O26
C34H26O22
C41H28O26
C41H30O26

305.0674
289.0723
577.1349
913.1461
457.0779
441.0828

305.0667
289.0718
577.1351
913.1469
457.0776
441.0827

1.25
2.27
0.66
0.62
2.21
1.49

99.38
97.85
99.02
99.36
97.4
98.27

4.3
5.0
5.6
6.9
7.7
9.7

C15H14O7
C15H14O6
C30H26O12
C44H34O22
C22H18O11
C22H18O10

169.0147
569.2246

169.0142
569.2240

2.58
0.75

98.89
97.97

3.1
10.9

C7H6O5
C27H38O13

Anthocyanins
29
Cyanidin 3-O-glucoside
Ellagic acid derivatives
1
Valoneic acid dilactone
2
Ellagic acid hexoside
3
Ellagic acid pentoside
5
Ellagic acid desoxyhexoside
7
Ellagic acid
8
Ellagic acid acetyl-rhamnoside
9
Ellagic acid acetyl-rhamnoside
10
Ellagic acid derivative
12
Ellagic acid derivative
13
Ellagic acid derivative
Ellagitannins
15
Vescalagin
16
Castalagin
48
HHDP-galloyl-glucose
18
Di-HHDP-glucose
47
Di-HHDP-galloyl-glucose
19
HHDP-galloyl-glucose
20
Di-HHDP-galloyl-glucose
21
HHDP-digalloyl-glucose
23
Di-HHDP-galloyl-glucose
24
Tri-galloyl-HHDP-glucose
Proanthocyanidins
49
Gallocatechin
50
Catechin
51
Procyanidin dimer B-type
22
Gallocatechin gallate dimer
46
Gallocatechin gallate
53
Catechin gallate
Gallic acid and derivatives
14
Gallic acid
25
Gallic acid derivative

(5) (most probably rhamnoside). In addition, two isomeric compounds with pseudomolecular ion at m/z 489 (8, 9) were detected.
The fragmentation and the high resolutions MSMS coincided with
ellagic acid acetyl rhamnoside, previously reported in several
Myrtaceae species, but that were not reported in camu-camu. In
addition other ellagic acid derivatives were detected with pseudomolecular ions at m/z 585 (10) and m/z 719 (12, 13). All these
compounds produced fragments at m/z 301 for ellagic acid, and
the molecular formulae were established with the high-resolution
Q-TOF equipment, but these compounds were not completely
characterised as the fragmentation did not provide enough information to suggest a chemical structure.
Ten ellagitannins were detected in camu-camu powders. Their
UV spectra provided information on the number of hexahydroxydiphenoyl (HHDP) and galloyl residues that every compound had attached to the glucose nucleus (Salminen, Ossipov, Loponen,
Haukioja, & Pihlaja, 1999). The isomeric C-glucosides vescalagin
(15) and castalagin (16) were characterised by the pseudomolecular ion at m/z 933, and the characteristic fragments that did not include the ellagic acid fragment at m/z 301, and were conrmed by
chromatographic comparisons with authentic standards. Two isomers of HHDP-galloyl-glucose (19, 48), with a pseudomolecular
ion at m/z 633 and fragments at m/z 463 (M-H-galloyl) and 301 (ellagic acid), were characterised. Two isomers of di-HHDP-galloylglucose (causarictin/potentillin) (23, 47), with a pseudomolecular

ion at m/z 935 and fragments at m/z 917 (M-H-H20), 633 (M-HHHDP) and 301 (ellagic acid), were also detected. Di-HHDP-glucose
(pedunculagin) (18) with a pseudomolecular ion at m/z 783 and
fragments at m/z 481 (M-H-HHDP) and 301 was also detected, as
well as digalloyl-HHDP-glucose (21) with m/z 785 and fragments
at m/z 483 (M-H-HHDP) and 301, and trigalloyl-HHDP-glucose (tellimagrandin II) (24) with m/z 937 and fragments at m/z 767 (M-Hgalloyl), 741, 465 (M-H-galloyl-HHDP) and 301 (ellagic acid).
The characterisation of the different ellagitannins was conrmed by High Resolution MS analyses using a Q-TOF detector,
with the calculation of the corresponding molecular formulae
(Table 2).
Gallic acid (14) was also detected as well as an unidentied gallic acid derivative (25) with a pseudomolecular ion at m/z 569.
Proanthocyanidins were also present in the camu-camu our,
although the UV spectra and UV response was not relevant compared with those of the rest of phenolic compounds that had a
higher UV absorption coefcient. Three main proanthocyanidins
were detected in the HPLCDAD-MSMS chromatograms (Table 1),
in which gallocatechin-gallate (46) with a pseudomolecular ion at
m/z 457, a dimer of gallocatechin-gallate (22) at m/z 915 and fragments at m/z 457 and m/z 169 (gallate), and a trimer of gallocatechingallocatechingallategallocatechingaltate (17) at m/z 1221
with a main fragment at m/z 915 (M-H-gallocatechin), were detected. These were conrmed by High-resolution Q-TOF, with the

584

D. Fracassetti et al. / Food Chemistry 139 (2013) 578588

Table 3
Content of phenolic compounds in camu-camu powders.
Number

Compound

Pulp powder (mg/


100 g powder)

Flour (mg/100 g
powder)

Peel (mg/100 g
fresh weight)

Pulp (mg/100 g
fresh weight)

Seeds (mg/100 g
fresh weight)

Myricetin 3-O-hexoside
Myricetin 3-O-pentoside
Myricetin 3-O-pentoside
Quercetin 3-O-hexoside
Quercetin 3-O-pentoside
Myricetin
Total

0.55 0.11
Trace
1.47 0.07
Trace
0.05 0.01
0.98 0.04
3.05 0.07

1.40 0.05
Nd
Trace
Nd
Nd
5.28 0.10
6.68 0.16

0.81
0.26
2.29
Trace
Nd
Nd
3.36

0.38
0.08
1.33
Nd
0.14
Nd
1.93

0.55
0.91
Trace
Nd
Nd
Nd
1.46

Anthocyanins
29
Cyanidin 3-O-glucoside

19.63 0.60

Nd

0.67

0.32

Nd

Ellagic acid derivatives


1
Valoneic acid dilactone
2
Ellagic acid hexoside
3
Ellagic acid pentoside
5
Ellagic acid desoxyhexoside
7
Ellagic acid
8
Ellagic acetyl rhamnoside
9
Ellagic acetyl rhamnoside
10
Ellagic acid derivative
12
Ellagic acid derivative
13
Ellagic acid derivative
Total

Nd
Nd
1.22 0.07
2.84 0.11
5.60 0.11
Nd
Nd
Nd
Nd
Nd
9.75 0.10

5.46 0.10
7.52 0.08
20.42 0.16
12.94 0.28
76.49 0.49
3.90 0.06
3.13 0.06
2.27 0.14
1.51 0.03
1.61 0.01
129.80 0.15

Nd
Nd
0.28
Nd
Nd
Nd
Nd
Nd
Nd
Nd
0.28

Nd
Nd
0.06
Nd
Nd
Nd
Nd
Nd
Nd
Nd
0.06

2.00
4.62
10.43
6.44
5.04
3.15
2.14
0.99
0.15
0.40
35.36

Nd
12.71 0.51
3.39 0.16
Nd

64.51 1.11
228.88 1.89
17.93 0.17
29.89 0.21

2.70
1.43
2.73
Nd

3.59
1.45
1.63
Nd

81.30
114.67
15.29
8.81

Nd
Nd

37.80 0.17
38.37 2.78

Nd
Nd

Nd
Nd

6.56
19.00

Nd
Nd
Nd

7.89 0.41
5.71 0.35
4.89 0.17

Nd
Nd
Nd

Nd
Nd
Nd

Nd
Nd
0.99

16.10 0.33

435.86 0.98

6.86

6.67

246.62

Flavonols
4
6
6
26
27
28

Ellagitannins
15
Vescalagin
16
Castalagin
18
Di-HHDP-glucose (pedunculagin)
47
Galloyl-bis-HHDP-glucose
(casuarinin/potentillin)
19
HHDP-galloyl-glucose
20
Di-HHDP-galloyl-glucose (casuarictin/
potentillin)
21
HHDP-galloyl-glucose
23
Di-HHDP-galloyl-glucose (casuarictin)
24
Tri-galloyl-HHDP-glucose
(tellimagrandin II)
Total
Gallic acid derivatives
14
Gallic acid
25
Gallic acid derivative
Total
Proanthocyanidins
17
Gallocatechin-gallocatechin gallategallocatechingallate
46
Gallocatechin-gallate
22
Gallocatechin-gallate-dimer
Total
Total

Nd
Nd

29.56 0.71
6.40 0.50
35.96 0.54

Nd
Nd

Nd
Nd

8.20
8.62
16.82

Nd

19.38 0.78

Nd

Nd

27.69

Nd
Nd

27.35 0.89
17.46 0.39
64.19 0.54

Nd
Nd

Nd
Nd

3.02
5.06
35.77

10.50

8.66

336.03

48.54 0.28

672.49 0.55

Values are mean values (n = 3) with standard deviation.

determination of the structural formulae (Table 2). In addition, gallocatechin (49), catechin (50), a B-type procyanidin dimer (51) at
m/z 577, a gallocatechin-gallate dimer (22), gallocatechin gallate
(46), and catechin gallate (53) were detected. The sensitivity of
the Q-TOF equipment allowed the detection of these proanthocyanidins, although some of them were not detected in the HPLC
ESI-MSMS analyses (Table 1).
3.2. Phenolics quantication in powders and changes during the drying
process
The total phenolics content of both camu-camu powders and
the fresh berries were calculated as an addition of the individual
characterised compounds by their UV absorbance at the convenient wavelength, and using the appropriate external standards
for each type of compound (Table 3). In the case of proanthocyanidins, they were quantied by HPLCUV after the acid catalysed

degradation in the presence of phloroglucinol (Kennedy & Jones,


2001) (Table 4).
The phenolic content in pulp powder was 48.54 mg/100 g (Table 3). Among the phenols the total concentration of myricetin
and its 3-O-glycosides (3 mg/100 g) was higher than that previously reported in fresh fruit (Reynertson et al., 2008). When comparing with fresh berries, it is evident that some degradation of
avonols occurs during the drying process, and that the aglycone
myricetin, which is not detected in the fresh berry, is released (Table 3). No anthocyanins were detected in the our, while they were
present in the skin and pulp, showing that anthocyanins are also
sensitive to the thermal treatment. The content of ellagic acid
and its conjugated derivatives was 9.75 mg/100 g powder, of which
the 60% was for free ellagic acid. The amount of quercetin and its
glycosides (<0.05 mg/100 g) was lower than that reported in literature related to either fresh fruit (Reynertson et al., 2008) or dry
matter (De Souza Schmidt Gonalves et al., 2010). Neither gallic,

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D. Fracassetti et al. / Food Chemistry 139 (2013) 578588

Table 4
Proanthocyanidin analysis of camu-camu our and fresh berry skins and seeds by phloroglucinolysis. HPLC Retention times, UV/Vis spectra, HPLCESI-MSMS fragmentations.
Number

Compound

Retention time (min)

[MH]

kmax (nm)

MS fragments

30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45

Catechin
Epicatechin
(epi)Catechin aduct
(epi)Catechin aduct
(epi)Catechin gallate
(epi)Catechin gallate
(epi)Catechin gallate aduct
(epi)Catechin gallate aduct
(epi)Gallocatechin
(epi)Gallocatechin
(epi)Gallocatechin aduct
(epi)Gallocatechin aduct
(epi)Gallocatechin gallate
(epi)Gallocatechin gallate
(epi)Gallocatechin gallate aduct
(epi)Gallocatechin gallate aduct

15.3
19.1
11.5
12.0
24.9
27.4
17.4
23.6
10.7
14.4
6.1
8.6
19.3
22.2
10.2
12.4

289
289
413
413
441
441
565
565
305
305
429
429
457
457
581
581

275

245, 205, 179

277
277

287, 261, 175


331, 289, 169

277

439, 413, 395

277
277

287, 219, 178


287, 219, 178

275
275

303, 261, 177


331, 305, 287

275

455, 429, 319

Number

Compound

mg/100 g powder (average sd)

Peel mg/100 g dry weight

Seeds mg/100 g dry weight

30
31
33
34
36
38
39
41
42
45

Catechin
Epicatechin
(epi)Catechin aduct
(epi)Catechin gallate
(epi)Catechin gallate adduct
(epi)Gallocatechin
(epi)Gallocatechin
(epi)Gallocatechin aduct
(epi)Gallocatechin gallate
(epi)Gallocatechin gallate aduct

213.53 3.27
Trace
845.48 1.89
52.02 0.40
677.73 11.99
45.28 4.72
60.10 0.67
409.20 6.47
352.75 0.13
767.46 0.13

4.46
Trace
24.57
Trace
41.88
316.51
Trace
438.43
Trace
Trace

70.17
Trace
889.03
96.66
276.59
126.04
37.38
276.59
629.89
689.69

mpd

3.00

2.39

2.87

Total

3525.54 3.30

825.85

3092.04

Bold values are the main fragments.

proanthocyanidins or kaempferol derivatives were detected in


pulp powder. Thermal processing released free ellagic acid from
ellagitannins both in pulp powder or in the our (Table 3). This
is consistent with previous publications in which thermal treatments released ellagic acid from raspberry ellagitannins during
jam processing (Zafrilla, Ferreres, & Toms-Barbern, 2001).
Camu-camu our contained a high ellagitannin content (435 mg/
100 g d.w.), and this compound came mainly from the seeds. Thermal treatment induced some transformations as the conversion of
vescalagin into castalagin (Table 3). In addition HHDP-galloyl-glucose (19) was partially converted to an isomer (21) in the our, and
Di-HHDP-galloyl-glucose (20) was also transformed to an isomer
(22), and both isomers 21 and 22 were not detected in the fresh
berry seeds suggesting that they were formed during the drying
process.
The highest amounts of phenols were detected in the camucamu our and in the berry seeds (Table 3, Table 4, Fig. 1). As for
the pulp powder, only the identied peaks in the chromatograms
were quantied. No quercetin, rutin, kaempferol, and anthocyanins
were detected in the camu-camu our analysed although these
compounds were found in the camu-camu fresh berries analysed
and in previous studies (Reynertson et al., 2008; De Souza Schmidt
Gonalves et al., 2010). The total concentration of phenols in camucamu our, quantied by addition of the individual compounds
characterised, was 4007.95 mg/100 g (Table 3, Table 4). The
amount of proanthocyanidins, namely catechin, and gallocatechin,
and their gallate derivatives (and trace amounts of the epicatechin
isomers) was up to 3423.5 mg/100 g (Table 4, Fig. 2), which is more
than twice the content of other fruits recognised as a good source
of proanthocyanidins, such as strawberry (1170 mg/100 g dry matter), apple (1460 mg/100 g dry matter), and grape (1420 mg/100 g
dry matter) (Tarascou et al., 2010). Moreover, the content of these
compounds was higher than that found in cocoa powder (278 mg/
100 g dry matter) (Lee, Kim, Lee, & Lee, 2003) and Brazil nut

(419 mg/100 g dry matter) (John & Shahidi, 2010). Among the proanthocyanidins, the most abundant compound was (epi)gallocatechin gallate (1120.21 mg/100 g), followed by catechin gallate
(729.8 mg/100 g) and (epi)gallocatechin (514.58 mg/100 g). Analysis of the fresh berries showed that (epi)gallocatechin gallate was
mainly present in the seeds while (epi)gallocatechin was more
abundant in the skins (Table 4). The proanthocyanidin composition
resulted to be similar to the phenolic prole of green tea which has
been reported to have a relevant nutritional potential (McKay &
Blumberg, 2002). Catechin-gallates have recently been described
as potent inhibitors of a-amylase and a-glucosidase with implications in obesity prevention (Yilmazer-Musa, Grifth, Schneider, &
Frei, 2012). In addition, the polymerisation degree of the proanthocyanidins present in this powder was around 3, suggesting a relatively high absorption in humans (Scalbert & Williamson, 2000)
which can consequently have potential biological activities. It has
been reported that proanthocyanidins play an important role in
several biological processes resulting in health benets. Several
benecial effects of proanthocyanidins have been reported such
as antioxidant, anti-inammatory, antimicrobial, antiproliferative,
cardioprotective, hypolipidemic and antidiabetic properties
(Nandakumar, Singh, & Katiyar, 2008; Serrano, Puupponen-Pimia,
Dauer, Aura, & Saura-Calixto, 2009; Blad, Arola, & Salvado, 2010).
Ellagitannins were also relevant constituents of camu-camu
our although the concentration found was lower than that of
the proanthocyanidins (435.9 mg/100 g) (Table 3). However, the
level of ellagitannins was higher than that of strawberry
(162 mg/100 g dry matter) (Buenda et al., 2010) and similar to
that of raspberry (415 mg/100 g dry matter) (Bobimait, Viskles,
& Venskutonis, 2012). The most abundant ellagitannins were castalagin (228.9 mg/100 g), followed by vescalagin (64.5 mg/100 g)
and di-HHDP-galloyl glucose (38.4 mg/100 g). In addition, the content of free ellagic acid and its glycosidic conjugates (129.80 mg/
100 g) was higher than those previously reported in strawberry

586

D. Fracassetti et al. / Food Chemistry 139 (2013) 578588

ascorbic acid concentration in berries depends on the ripening


stage (Marques, Ferreira, & Freire, 2007). The ascorbic acid content
in these camu-camu powders was much higher than those reported for freeze-dried fruits, such as red pummel (0.02 g/100 g
dry matter), strawberry (0.6 g/100 g dry matter), and tomato
(0.6 g/100 g dry matter), which are known to be a source of vitamin
C (Tsai, Chang, & Chang, 2007; Asami, Hong, Barrett, & Mitchell,
2003; Chang, Lin, Chang, & Liu, 2006).
The dehydroascorbic acid values were of 0.69 0.31 g/100 g and
0.578 0.027 g/100 g for camu-camu pulp powder and camucamu our, respectively (Table 5). Dehydroascorbic acid usually
represents about 1020% of the ascorbic acid present in fruit and
vegetable products (Riemer & Karel, 1978; Borowski, Szajdek,
Borowska, Ciska, & Zielinski, 2008), and is considered to be the rst
compound produced in the oxidative degradation of ascorbic acid
(Chang et al., 2006). In the present study, dehydroascorbic acid
represented the 19.5% of the vitamin C (ascorbic + dehydroascorbic
acid) for pulp powder and 6.4% for camu-camu our, conrming
the data previously reported in literature, and suggesting that the
higher temperature used for the production of the pulp powder led
to a higher transformation of ascorbic acid into dehydroascorbic
acid. In addition, the higher content of antioxidant polyphenols
in the skin than in the pulp could also help preventing the transformation of ascorbic acid into dehydroascorbic acid.

(8.820.6 mg/100 g dry matter) (Bobimait et al., 2012), and Brazil


nut (11.4 mg/100 g dry matter) (John & Shahidi, 2010). More than
half of the ellagic acid was present as free ellagic acid (Table 3,
Fig. 1). A small quantity of myricetin derivatives was also present
(6.68 mg/100 g) although in lower concentration than that
reported in camu-camu berries (24 mg/100 g dry matter)
(Reynertson et al., 2008).
The phenolic compounds in camu-camu contain a high number
of hydroxyls, in which three phenolic hydroxyl groupings are present as it occurs in gallic acid, ellagitannins, myricetin, and gallocatechin-gallate, and this chemical feature could be linked to the
high antioxidant activity found.
3.3. Ascorbic acid and dehydroascorbic acid quantication
The ascorbic acid contents were 3.51 0.97 g/100 g and
9.04 0.95 g/100 g in camu-camu pulp powder and camu-camu
our, respectively (Table 5). These values are higher than those reported in fresh camu camu fruit (Chirinos et al., 2010; Runo et al.,
2010) due to the lower moisture content of the powder (13%) (De
Souza Schmidt Gonalves et al., 2010). This content was very relevant although the amount of ascorbic acid present in fresh berries
could be partly degraded during the production of these powders
from the fresh fruit that needs thermal treatments (Shoan et al.,
2011), and some variations in the content could had happened as

3.4. Antioxidant capacity

Table 5
Content of ascorbic acid, dehydroascorbic acid and antioxidant capacity (ABTS, DPPH
and ORAC assays) in camu-camu pulp powder and our.
Camu-camu
pulp powder
Ascorbic acid (g/100 g powder)
Dehydroscorbic acid (g/100 g powder)
Antioxidant capacity
ABTS assay (lmol Trolox/100 g powder)
DPPH assay (lmol Trolox/100 g powder)
ORAC assay (lmol Trolox/100 g powder)

The antioxidant capacity was determined in both pulp powder


and our using ABTS, DPPH and ORAC assays (Table 5). The ABTS
assay showed the free-radical scavenging activity for pulp powder
(167.5 54.4 lmol Trolox/g dry powder) which is about 4-fold lower than that of camu-camu our (752.3 41.0 lmol Trolox/g dry
our). The dilution factor of the addition of 10% maltodextrin
during the pulp drying process would have a minor contribution
to the difference observed in antioxidant capacity. These values
are higher than those reported in foods well known for their high
antioxidant capacity, such as guava (about 98 lmol Trolox/g dry
product) (Corral-Aguayo, Yahia, Carrillo-Lopez, & Gonzlez-Aguilar,

Camu-camu
our

3.51 0.97
0.69 0.31

9.04 0.95
0.578 0.027

167.5 54.4
510.5 156.1
337.1 77.9

752.3 41.0
1036.4 211.2
755.2 77.9

DAD1 A, Sig=280,100 Ref=off (Y:\CAMU\FLOROGLUCINOLISIS\05-05\07051212.D)


mAU

275

250

225

200

175

150

125

100

10

12

14

16

18

20

22

24

min

Fig. 2. HPLCDAD chromatogram (280 nm) of camu-camu our proanthocyanidins analyzed after acid catalyzed degradation in the presence of phloroglucinol. For
compound identication see Table 4.

D. Fracassetti et al. / Food Chemistry 139 (2013) 578588

2008), strawberry (about 250 l mol Trolox/g dry product)


(Proteggente et al., 2002), and cocoa powder (617 lmol Trolox/g
dry matter) (Lee et al., 2003). Similarly, the antioxidant capacity
determined as DPPH was higher in camu-camu our
(1036.4 211.2 lmol Trolox/g dry our) than in pulp powder
(510.5 156.1 lmol Trolox/g dry powder). These values were again
higher than those reported for other polyphenol-rich powders
including guava (300 lmol Trolox/g dery matter) (Corral-Aguayo
et al., 2008), apple and kiwi (37 lmol Trolox/g dry matter)
(Wijngaard, Rle, & Brunton, 2009), cocoa (458 lmol Trolox/g
dry matter) (Lee et al., 2003), and broccoli (3 lmol Trolox/g dry
matter) (Borowski et al., 2008).
The camu-camu our revealed an antioxidant capacity value of
ORAC 2-fold higher (755.2 77.9 lmol Trolox/g dry our) than
that of the pulp powder (337.1 77.9 lmol Trolox/g dry powder).
The high value obtained in the camu-camu our was expected as
this product revealed high contents of both vitamin C and phenolic
compounds. This antioxidant capacity was higher than those reported for strawberry (153.6 lmol Trolox/g dry matter) (Wang,
Cao, & Prior, 1996), and blackberry (204 lmol Trolox/g dry matter)
(Wang & Lin, 2000) and were consistent with those previously reported for camu-camu (De Souza Schmidt Gonalves et al., 2010).
The antioxidant capacity determined by the ABTS assay was
lower than that determined by the DPPH assay for the camu-camu
our and pulp powder, and this differs from previous studies with
other plant extracts (Arnao, 2000). This could be explained by the
higher content of phenolics, particularly proanthocyanidins, in the
our. Nevertheless, the high antioxidant capacities of the our
determined by the ABTS, the DPPH and the ORAC assays can also
be related to both the high content of vitamin C and phenolic compounds present. The ORAC values were also lower than those
determined as DPPH for both powders. This could again be due
to the extraction of the camu-camu powders in water which can
probably limit the extraction of some phenolics, such as the
proanthocyanidins.
4. Conclusion
Camu-camu fruit is considered as a good source of bioactive
compounds potentially benecial for human health. The consumption of this fruit is limited by its acidity and bitter taste, and therefore the dehydration process is a good alternative for its use as a
powdered food ingredient. These results revealed that the two
camu-camu powder products studied are excellent sources of bioactive substances including vitamin C and phenolic compounds
which resulted in high antioxidant capacity values. Camu-camu
our showed a higher concentration of vitamin C than the camucamu pulp powder and high proanthocyanidins content. Therefore,
camu-camu our has a good potential to be used as an ingredient
for the formulation of functional foods. The results provide further
evidence that this our is a rich source of bioactive compounds
with potential health-promoting properties such as antioxidant,
anti-inammatory, and hypocholesterolemic activities which have
been related to vitamin C and phenolic compounds such as avonoids and ellagitannins. However, in vivo and intervention studies
are needed to assess the nutritional and functional potential of this
camu-camu product.

Acknowledgements
FATB is grateful to the Spanish MICINN (Consolider Ingenio
2010- Fun-C-Food CSD2007-0063 and AGL2004-06076-C02-01)
and Fundacin Seneca de la Region de Murcia (grupo de excelencia
GERM 06, 04486).

587

D.F. was supported by the post-doctoral fellow Dote Ricerca:


FSE, Regione Lombardia and Cariplo Foundation (Rif. Pratica 20102303).

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