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Contents lists available at ScienceDirect

The Journal of Supercritical Fluids

journal homepage: www.elsevier.com/locate/supflu

Extraction of bioactive compounds from peach palm pulp

(Bactris gasipaes) using supercritical CO2
Faber A. Espinosa-Pardo a , Julian Martinez b , Hugo A. Martinez-Correa c,
a
b
c

Universidad Nacional de Colombia, Departamento de Ingeniera, Cr 32 #12-00, Palmira, Colombia

Food Engineering Department, Faculty of Food Engineering, University of Campinas, P.O. Box 6121, 13083-862 Campinas, So Paulo, Brazil
Universidad Nacional de Colombia, Cr 32 # 12-00, Palmira, Colombia

a r t i c l e

i n f o

Article history:
Received 26 September 2013
Received in revised form 2 March 2014
Accepted 7 May 2014
Available online xxx
Keywords:
Bactris gasipaes
Supercritical uid
Biocompounds
Carotenoids
Antioxidant activity

a b s t r a c t
Natural compounds with biological activity have recently attracted special interest in the agro-industry
as sources of additives in nutraceutical food production and pharmaceutical industries. Herein, we evaluated extracts obtained from peach palm fruit (Bactris gasipaes) using supercritical carbon dioxide, in terms
of yield, total phenolic content, total avonoids, total carotenoids, and antioxidant activity by -carotene
bleaching method. Extractions were performed at 40, 50, and 60 C and 100, 200, and 300 bar; additionally, Soxhlet (with petroleum ether) and methanol extraction were conducted. The results showed that
supercritical CO2 allows obtaining extracts rich in carotenoids and, although it presents lower yield than
conventional extraction (SOX), supercritical CO2 represents a technique with greater advantages. The
best operation condition for supercritical extraction was 300 bar40 C, given that the highest concentration of carotenoids was obtained, without the yield being signicantly different from that obtained
with 300 bar60 C, this extract had antioxidant activity comparable to that of commercial caffeic acid.
2014 Elsevier B.V. All rights reserved.

1. Introduction
Bioactive compounds are naturally widespread in the plant
kingdom because they are synthesized as secondary metabolites
with defense functions, besides being responsible for properties
of color, astringency, and avor of fruits and vegetables. They
are increasingly important because given their chemical structure,
these compounds are suitable for scavenging free radicals found
in the human body; said free radicals behave as reactive oxygen
species (ROS) enabling development of chronic multifactorial diseases [1].
These compounds are credited for the mass consumption of
fruits due to their high content of antioxidants; with benecial effects in preventing cardiovascular and circulatory diseases,
cancer, and neurological diseases, given their anti-inammatory,
anti-allergic, antimicrobial, antithrombotic, and antineoplastic
activity [2,3]. Tropical countries like Colombia and Brazil, because
of their abundance of exotic fruits, have a huge potential for
the exploitation of the resource to obtain bioactive compounds
of underutilized fruits like the peach palm fruit (Bactris gasipaes), which could be used as active ingredients in pharmaceutical

Corresponding author. Tel.: +57 2 286 8888; ext. 34350, fax: +57 2 286 8808.
E-mail addresses: hamartinezco@unal.edu.co, hamartinez@yahoo.com
(H.A. Martinez-Correa).

products, in controlling oxidation processes in food processing, in

nutraceutical food production with high added value, as well as in
the cosmetic industry [4].
Peach palm fruit (B. gasipaes) is a palm of the Arecaceae family,
cultivated in tropical America from Costa Rica to Brazil and Bolivia
in wet and low zones. It is commonly known as cachipay, chontaduro, pejib, and pu
nu
na (Amazon). The fruit has a brous
and eshy mesocarp of deep yellow or orange color; it may be considered a fruit with high nutritional value due to its high content
of ber, oils, -carotene, for eight essential amino acids [46], and
for its energy value. However, the main feature of the recent interest in working with this fruit is the -carotene content that could
be obtained from it as a vitamin A precursor of high antioxidant
activity because it can capture free radicals due to its conjugated
double-bond system [2].
Extraction with supercritical uids is a technique that uses the
properties of uids over their critical points to selectively extract
soluble components from raw plant materials; additionally, supercritical carbon dioxide is recognized as an ideal solvent to extract
bioactive compounds because it is nontoxic, non-explosive, readily available, easy to remove from the nal extract, does not cause
major disruptions in biocompounds, and its biological properties
can be preserved [79].
This work sought to evaluate bioactive compounds of extracts
obtained from the peach palm fruit using supercritical carbon
dioxide (Sc-CO2 ) at different pressures and temperatures. For this

http://dx.doi.org/10.1016/j.supu.2014.05.010
0896-8446/ 2014 Elsevier B.V. All rights reserved.

Please cite this article in press as: F.A. Espinosa-Pardo, et al., Extraction of bioactive compounds from peach palm pulp (Bactris gasipaes)
using supercritical CO2 , J. Supercrit. Fluids (2014), http://dx.doi.org/10.1016/j.supu.2014.05.010

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Table 1
Extraction conditions.

Fig. 1. Extraction owchart [11]. B1: CO2 pump, R1: CO2 reservoir, CA: adsorption
column, T: thermocouples, FR: collection bottle, TF: ow totalizer, FI: lter, V1:
locking valve, LF: extraction bed, V2: retention valve, M: manometer, V3: valve.

Experiment

Temperature ( C)

Pressure (bar)

E1
E2
E3
E4
E5
E6
E7
E8
E9

40
40
40
50
50
50
60
60
60

100
200
300
100
200
300
100
200
300

methanol (CHEMCO, absolute grade); the solution was stirred for

24 h at 25 C; then, the solution was vacuum ltered (0.45 m lter, Vaccuo Tecnalpump TE-0581) and rotor-evaporated (Heidolph
220V). Soxhlet extraction was performed for 6 h using 2 g of raw
material and 60 mL of petroleum ether (ECIBRA) as solvent; the
ether-extract solution was rotor-evaporated (Heidolph 220V) at
40 C.
2.3. Extraction yield

purpose, the following variables were evaluated: extraction yield,

total phenolic content (TPC), total avonoids (TF), total carotenoids
and antioxidant activity (AA%) by -carotene bleaching (BCB)
method of each of the extracts obtained.

The amount of extract obtained (E) in relation to the amount

of raw material (RM) used in each type of extraction is a significant factor in assessing bioactive extracts and on the extraction
techniques.

2. Materials and methods

2.4. Chemical characterization: TPC, TF, and total carotenoids

2.1. Raw material and chemical characterization

For all Sc-CO2 , MET, and SOX extracts total phenols content
(TPC) were quantied, expressed as (mg GAE)/g extract through
the FolinCiocalteu method [12]. Total avonoids (TF) were quantied via spectrophotometric method, according to the methodology
described by Zhishen et al. [13]. Gallic acid (Sigma Aldrich) was
used to construct the calibration curve for TPC at different concentrations and, nally, the following linear equation came about: Abs
B = 0.090 + 0.002, R2 = 0.998, where Abs is absorbance (nm) and B is
phenolic content (mg mL1 ).
Catechin was used as pattern for TF (Sigma Aldrich) and the
equation obtained was: C = 0.217 Abs, R2 = 0.999, where C is the
avonoid content (GAE mg mL1 ).
Total carotenoid content was determined according to the
methodology described by Szydowska-Czerniak et al. [14] modied. Extract samples (5.08.0 mg) were diluted in 10 mL of
n-hexane (96% purity, EMSURE Merck); subsequently, the solution was loaded onto the spectrophotometer (FEMTO 800 XI) at
450 nm absorbance, using a 1 cm quartz cell. The calibration curve
was prepared by using standard pattern -carotene (97.0% purity,
Fluka Analytical) at different concentrations ranging from 0.02 to
6.1 mg mL1 . The resulting calibration curve was D = 0.006 Abs,
R2 = 0.995, where D is the total carotenoid content expressed as
-carotene equivalent (mg mL1 ).
-Carotene bleaching is based on a spectrophotometric method
monitoring oxidation products due to degradation of linoleic acid.
The methodology used was described by Martinez-Correa et al.
[15]. Briey, 5 mL of a dry emulsion of -carotene and linoleic
acid transferred to a test tube and 0.2 mL of extract diluted in
ethanol, at a 200-g/mL concentration was added. Similar standard solutions, quercetin and caffeic acid solutions (200 g/mL)
were used as positive controls (standard solutions). The control
solution was prepared the same way, except that the solution was
replaced by 0.2 mL of pure ethanolic extract. Both tubes with the
extract and the control were subjected to thermal auto-oxidation
at 50 C for 120 min and absorbance was measured at 464 nm
(spectrophotometer FEMTO 800 XI) at 30-min intervals, against a

Peach palm fruit grown in el Tambo, Cauca (Colombia) were

acquired in the municipal market in Cali (Colombia), then the exocarp was removed and the mesocarp was cut, lyophilized, ground,
vacuum packed, and refrigerated until its later use.
Chemical characterization of peach palm fruit (proximate analysis) for moisture, ash, crude fat, crude ber, and protein content was
performed by using methods 966.02, 923.03, 920.39, 920.87, and
962.09, respectively, from the Ofcial Analytical Chemists Association (AOAC) [10].
2.2. Supercritical extraction
Supercritical extractions were carried out in a supercritical unit
(Fig. 1), which operates up to a maximum pressure of 35 MPa. The
unit has a high-pressure pump for the solvent (Thermo Separation
Products, model 2000, Florida, USA), two programmable thermostatic baths (Marconi, model MA-159 and Marconi, model MA-184,
Piracicaba, SP, Brazil), a ow totalizer (LAO, model G 0.6 0.001 m3 ,
So Paulo, SP, Brazil), thermocouples, and three control manometers (Record (50.0 0.5) MPa, So Paulo, SP, Brazil). The extraction
bed has an internal 3.41 cm diameter and 46 cm height [11].
All extractions were performed using 10.0 0.010 g of raw
material, constant CO2 ow of 3.00 l/min for 91 min and a
solvent/raw material S/F ratio (w/w) of 46. The supercritical extractions were performed at different temperature and pressure values;
40, 50, and 60 C and 100, 200 and 300 bar, respectively (Table 1).
After extraction, the collection bottles, kept at 5 1 C during
extraction, were freed of residual CO2 , hermetically re-sealed, isolated from direct light, and stored under refrigeration (Metalfrio
Freezer) until its subsequent analysis.
Additionally, Soxhlet extraction (SOX) was conducted with
petroleum ether and methanolic extraction (MET) to compare
Sc-CO2 extraction to traditional methods. For MET, 2 g of peach
palm fruit were taken and added to a beaker containing 30 mL of

Please cite this article in press as: F.A. Espinosa-Pardo, et al., Extraction of bioactive compounds from peach palm pulp (Bactris gasipaes)
using supercritical CO2 , J. Supercrit. Fluids (2014), http://dx.doi.org/10.1016/j.supu.2014.05.010

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Table 2
Composition of peach palm pulp (Bactris gasipaes).
Component

Content (%)

Dry matter
Ash
Protein
Oil
Neutral ber
Carbohydrates

30.84
2.64
8.17
18.73
8.18
62.28

target solution prepared with 5 mL of the emulsion without the

-carotene plus 0.2 mL of ethanol. Antioxidant activity (AA%) was
calculated as percentage of inhibition related to the control that
represents 100% oxidation, using Eq. (1).

AA (%) =

(AC0 ACt ) (Am

Am
t )
0
AC0 ACt

100

(1)

where C represents the control solution and m the sample; A0

and At are the absorbance values at 0 and t minutes.

Fig. 2. Extraction yield and concentration of carotenoid for experiments. Lowercase letters correspond to signicant differences between the different extraction
conditions.

2.5. Statistical analysis

6

3. Results and discussion

3.1. Chemical characterization
As seen in Table 2, which shows the percentage composition
of peach palm pulp, the dry matter content is signicantly high
compared to other tropical fruits and it is similar to that reported
by Leterme et al. [16] which permits obtaining higher yields of
extracts; however, a signicant variation in the total mineral content was presented from the same study. Compared to Amazonian
fruit, it was observed that dry matter content is lower [6], but the
Colombian sample has higher protein content (8.17%) and ether
extract content (18.73%).
Table 2 also shows that the fruit has high lipid content (18.73%),
which together with a reddish color in the mesocarp might indicate
its high carotenoid content [17,18]. Fiber, carbohydrate, and protein
contents are similar to those reported by Ros et al. [19] for different
tropical Colombian peach palm varieties, showing their potential
for agro-industrialization.
3.2. Extraction yield
As seen in Fig. 2, the extraction yield for all Sc-CO2 extractions
showed lower values compared to SOX (up to 394%) and no significant differences with MET (except those at 100 bar). Supercritical
extraction yields reported by other authors for bioactive compounds and/or oils are relatively low compared to those obtained
in this work [8,9,20,21], this may be due to the selectivity and
afnity of peach palm soluble compounds in Sc-CO2 compared to
conventional extraction methods that obtain higher yields with little selectivity. The disadvantage with MET and SOX extractions is
the limitation in the subsequent recovery, separation, and purication of the extracts. Additionally, the extract has limitations for
food applications due to important reduction of solvents approved
for this industry [22].
The supercritical extraction yield is related to a complex balance
between the reduction of CO2 density and increased vapor pressure of the compounds as temperature increases, which basically

40C

50C

60C

Global yield (%)

All experiments were performed in duplicate and the results

were analyzed in the SAS statistical software, managing an array
of completely randomized blocks. A Tukey test was also performed
with a condence range 95%.

4
3
2
1
0
80

100

120

140

160

180

200

220

240

260

280

300

320

Pressure (bar)

Fig. 3. Global yield isotherms obtained by Sc-CO2 extraction. Experimental points

connected for better visualization.

represents the solubility of the bio-compounds in solvent [23].

The yield increases with increasing pressure; behavior observed
in Fig. 2. However, it is observed that in treatment E9 the pattern
changes, introducing the highest yield at 4.9%.
The behavior of the global yield isotherms obtained via SFE at
40, 50, and 60 C under different pressures is presented in Fig. 3.
As noted, the global yield of extraction decreases with increasing
temperature at 100 bar due to the effect of the decreased density of
the solvent versus the effect increased vapor pressure of the solute
[24,25]. The crossover phenomenon was not observed between the
isotherms at 40 and 50 C; however, at 50 and 60 C the crossover
phenomenon occurs at 200 bar, while considering the extractions
performed at 40 and 60 C the retrograde point is higher, close to
250 bar. These values are close to those reported by Filho et al. [21]
who also worked on carotenoid extraction.
The yield increases with pressure at three temperatures, the
lowest value was 0.19% and was obtained at the lower pressure
and higher temperature; an increase of 24.7% was obtained for
300 bar60 C. For 40 and 50 C isotherms yield increases in pressure from 200 to 300 bar do not have great inuence on extraction,
while for 60 C increases in pressure had a signicant inuence on
global extraction yield. The statistical analysis performed for only
the Sc-CO2 extractions showed signicant differences only with
the variation of pressure but not with temperature variation, as
reported in other studies [9,24,26].

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3.3. Chemical characterization of extracts

30

Total carotenoids: Carotenoids are the most interesting bioactive

organic pigments for fruits like peach palm, which have characteristic reddish color and are rich in oil; these pigments are responsible
for the color of the fruit are vitamin A precursors, while collaborating in immune system regulation, oxidation inhibitors, and
peroxidation of fats [27].
Fig. 2 shows that for supercritical extraction carotenoid concentration in the extracts increases as the yield rises; it means that it
is increased with pressure and decreases with increasing temperature. The E9 treatment (300 bar60 C) allows obtaining a higher
extraction yield and signicant carotenoid content. The carotenoid
content in the extracts obtained from treatments E1E9 reached
maximum values at highest pressure, and decreased with increasing temperature, which may be due to carotenoid sensitivity to
thermal degradation [18]. This same behavior was also reported by
Gracia et al. [26].
Regarding conventional extractions, SOX showed the highest
concentration of all extracts (4.157 mg/g), with signicant differences compared to MET and Sc-CO2 . Because of its non-polar
nature, it presents afnity for carotenoids and allows for higher
yields, as reported by Cadoni et al. [27] and Vgi et al. [28]. The
opposite occurs with MET, which is a polar solvent and gets the
lowest concentration of carotenoids. For both cases, the extracts
are not entirely free of residual solvent, and the use of petroleum
ether as solvent is not recommended for some applications because
of its carcinogenicity.
The ANOVA for carotenoid content showed that both temperature variation and pressure produce signicant differences in the
nal carotenoid concentration in the Sc-CO2 extracts; however, the
experiments conducted at 300 bar and 40 and 50 C showed no signicant difference from those obtained at 200 bar40 and 50 C;
hence, for extractions aiming to recover carotenoids, it is enough
to work at 200 bar and 40 C to obtain the high carotenoid content
(2.01 mg/gE).
Total phenolic and total avonoid tests were not detected via
spectrophotometry (TPC, TF <0.0102 mg mL1 ) in the extracts. Contreras et al. [29] reports total phenol content for peach palm fruit
signicantly lower compared to other Colombian exotic fruits, with
a value of 65.7 mg GAE/100 g of fresh raw material; additionally,
the non-polar nature of Sc-CO2 solubilizes and extracts basically
non-polar compounds, which correspond to the lipid fraction of
the array. It is not an effective method to obtain extracts rich in
phenolic compounds with polar nature [30].
The use of polar solvents like methanol allows the extraction
of phenolic compounds of polar nature. Martinez-Correa et al.
[15] reported that primary extraction of the lipid phase of plant
matrix (for example Sc-CO2 ) facilitates subsequent removal of the
polar compounds (e.g., phenolic) using conventional solvents like
ethanol and water, thereby, obtaining more concentrated polyphenol extracts.

25

A.A (%)

20
b

AA (%)
15

a,b
a

10

E1

E2

a
a

E5

E6

E7

5
0
E3

E4

E8

Extraction

E9 SOX MET Ac. Quer

Caf.

Fig. 4. Antioxidant activity of the different extracts. Lowercase letters correspond

to signicant differences between the different extraction conditions.

non-polar nature of the compounds in SOX extract allows that

during the test, these can be in the water/lipid interface, thus, protecting the emulsion and inhibiting oxidation of -carotene [15].
The same situation is explained for the AA value obtained for standard quercetin, which scored 26%.
Extracts resulting from supercritical technology presented
lower ability to inhibit oxidation reactions than the MET extract,
even with a greater carotenoid concentration than MET. This behavior suggests that the antioxidant activity determined by the DBC
method would not be related to the carotenoid content in the
extracts, as reported by Rodriguez-Amaya [31].
4. Conclusions
This study allowed showing that the use of supercritical CO2 produces carotenoid-rich extracts from peach palm pulp and, although
presenting lower yield than conventional extraction (SOX), the
Sc-CO2 technique offers advantages compared to conventional
extraction. The recommended operating condition for supercritical
extraction is 300 bar40 C because it allows obtaining the highest carotenoid concentration. The Sc-CO2 extracts obtained an AA
(%) comparable to standard antioxidants. The importance of carotene is mainly as vitamin A precursor, which provides several
benets to the human organism operation; as a natural colorant
in the food industry, making this bio-compound an alternative for
food (nutraceuticals) and pharmaceutical products.
Acknowledgements
The authors thank the staff at the LASEFI Laboratory in Brazil for
their cooperation in this research.

3.4. Antioxidant activity (AA)

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Fig. 4 shows the antioxidant activity (AA%) obtained by MET, SOX
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antioxidants (caffeic acid and quercetin). For supercritical extracts,
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MET also presented a statistically equal value, although it presented
the lowest concentration of carotenoids of all types of extracts.
The highest antioxidant activity was obtained by the SOX
extract, reaching 25% inhibition; it also presented the highest
overall extraction yield and the highest carotenoids concentration, which could be related to the high antioxidant activity. The

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Please cite this article in press as: F.A. Espinosa-Pardo, et al., Extraction of bioactive compounds from peach palm pulp (Bactris gasipaes)
using supercritical CO2 , J. Supercrit. Fluids (2014), http://dx.doi.org/10.1016/j.supu.2014.05.010