Food and Bioproducts Processing 1 1 7 ( 2 0 1 9 ) 224–230

Contents lists available at ScienceDirect

Food and Bioproducts Processing

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

Highly efficient pumpkin-seed extraction with the

simultaneous recovery of lipophilic and
hydrophilic compounds

Daniele F. Ferreira a,b , Juliano S. Barin a , Arianna Binello b ,

Valery V. Veselov c , Giancarlo Cravotto b,c,∗
a Departamento de Tecnologia e Ciência dos Alimentos, Universidade Federal de Santa Maria, CEP 97105-900 Santa
Maria, Brazil
b Dipartimento di Scienza e Tecnologia del Farmaco, University of Turin, Via P. Giuria 9, 10125, Turin, Italy
c Sechenov First Moscow State Medical University, Center of Bioanalytical Research and Molecular Design, 8

Trubetskaya ul, Moscow, Russia

a r t i c l e i n f o a b s t r a c t

Article history: Enabling technologies have led to the design of new extraction protocols for naturally
Received 18 May 2019 occurring compounds that are fast, sustainable and have low energy demands. Ultrasound-
Received in revised form 29 June assisted extraction (UAE) and microwave-assisted extraction (MAE) have been used, in this
2019 study, for the recovery of valuable nutraceuticals and primary metabolites from pump-
Accepted 24 July 2019 kin seeds (Curcubita sp.) and have been compared with conventional methods. In order
Available online 31 July 2019 to optimise extraction parameters and conditions, we have compared yields, total phe-
nolic content, antioxidant activity, as well as fatty-acid and lipid profiles. The best yield,
Keywords: in both lipophilic and hydrophilic compounds, was achieved under UAE with a ternary
Non-conventional extraction solvent mixture (hexane/ethanol/water - 30:49:21). However, the highest phenolic content
Microwaves and antioxidant capacity were obtained using MAE under subcritical conditions with good
Ultrasound extraction yields. The oil composition, as obtained using GC-FID analyses, showed a preva-
Subcritical solvents lence for unsaturated fatty acids; mainly linoleic and oleic acids. The use of UAE with ternary
Pumpkin seeds mixtures presented the highest lipid content and a slightly higher percentage of unsaturated
Phenolic compounds compounds. Preliminary tests to verify the scalability of UAE with a ternary mixture showed
Combined ultrasound/microwave that the extraction performance was maintained when using larger volumes (factor 10). This
extraction study will pave the road for the potential exploitation of UAE as a fast and efficient proce-
dure for the simultaneous recovery of lipophilic and hydrophilic compounds from pumpkin
seeds.
© 2019 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

1. Introduction represent a rich source of valuable oil and several health-promoting

The Cucurbitaceae family encompasses many edible plants, includ-
ing pumpkins (Cucurbita sp.), which can be considered one of the
most important species in food industries thanks to the production
of soups, jams, juices and alcoholic beverages (Dhiman et al., 2009).
Industrial processing leaves a large quantity of by-products (shells,
peels etc.), which are commonly discarded as agro-industrial waste
(Peiretti et al., 2017; Saavedra et al., 2013). However, pumpkin seeds
compounds (antioxidant, antidiabetic, cholesterol-lowering, cardiopro-
tective, antiinflammatory and diuretic properties), which can also
alleviate negative symptoms of benign prostatic hyperplasia. Consider-
able attention has therefore been paid to pumpkin-seed valorisation in
the agro-food industry as they are a valuable functional-food resource
(Patel, 2013; Fruhwirth and Hermetter, 2007). Traditional extraction
methods, which are often time- and energy-consuming, do not comply
with current demands for sustainability and may affect thermally sen-

∗
Corresponding author.
https://doi.org/10.1016/j.fbp.2019.07.014
0960-3085/© 2019 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.
 Food and Bioproducts Processing 1 1 7 ( 2 0 1 9 ) 224–230
sitive compounds (Hernández-Santos et al., 2016). This problem can be
resolved by means of enabling technologies that protect active com-
pounds from degradation. Academia and industry are concentrating
efforts on alternatives techniques that are considered to be more effi-
cient, as well as economically and environmentally-friendly (Chemat
et al., 2012; Chemat et al., 2015; Koubaa et al., 2016). This goal has been
achieved by means of ultrasound-assisted extraction (UAE) (Cravotto
and Binello, 2016) and microwave-assisted extraction (MAE) (Chemat
and Cravotto, 2013), which are widely recognized as fast and sus-
tainable alternatives for extraction processes (Chemat et al., 2017; Li
et al., 2013; Koubaa et al., 2016). The main advantages that can be
derived from efficient cell-wall rupture include its ability to provide
high extraction yields with significant time and energy savings. Fur-
thermore, it allows higher quality extracts to be obtained by ensuring
that target compounds and biological properties can be successfully
processed without losses. These features make UAE and MAE green and
cost-effective approaches for industrial application in the extraction of
natural products (Grillo et al., 2019; Ciriminna et al., 2016; Casazza et al.,
2010).
There is growing worldwide interest in economically viable strate-
gies for the efficient management of industrial waste and reductions
in environmental impact. This study exploits agro-industrial pump-
kin seeds to obtain value-added compounds via the use of green
extraction methods to achieve precisely those goals. MAE, UAE, a
microwave/ultrasound (MW/US) combination and MAE under sub-
critical conditions have been compared with conventional extraction
methods in this study. Extract yields, total phenolic contents, antiox-
idant activities and fatty acids profiles have all been investigated in
order to evaluate extraction efficiency.
2. Materials and methods
2.1. Plant material and chemicals
The pumpkin seeds were manually separated from agro-
industrial waste for the extractions and stored at −18 ◦ C. The
seeds presented a moisture content of 54.4%. Ethanol and hex-
ane (ACS grade, ≥99%, Merck-Sigma Aldrich, Milan, Italy) were
used for the extractions. Methanol and chloroform (ACS grade,
≥99%, Merck-Sigma Aldrich, Milan, Italy) were used for extract
dilution. The Folin-Ciocalteau reagent, gallic acid and sodium
carbonate were used for total phenolic-content analyses and
2,2-diphenyl-1-picrylhydrazyl (DPPH) for antioxidant-activity
analysis. They were all purchased from Merck-Sigma Aldrich.
The standards FAME C8-C24 and methyl heptadecanoate (Me
C17) were used for fatty-acid analyses.
2.2. Extraction procedure
All of the extractions were performed using ground pumpkin
seeds (0.8 – 2.0 mm) that were suspended at a 1:15 solid/liquid
ratio. The obtained extracts were filtered on a paper filter
under vacuum and the solvent evaporated to determine the
extraction yield (w/w).
2.2.1. Conventional extraction (CE)
Conventional extraction was performed under solvent reflux
according to Grillo et al. (2019). In this procedure 6 g of material
in either ethanol 60% v/v (CE-EtOH 60%) or hexane (CE-
hex) was used. The two extractions provided the contents of
hydrophilic and lipophilic compounds, respectively. Both were
performed in an oil bath under reflux and stirring at 80 ◦ C for
4 h.
225
2.2.2. MAE
Extraction was performed using a multi-mode microwave
oven working at 2.45 GHz (MicroSynth by Milestone, Bergamo,
Italy). The procedure was carried out in a closed PTFE (Teflon)
vessel using 6 g of sample in ethanol 60% v/v for 20 min. MW
extraction was operated in a temperature-control mode and
kept at 100 ◦ C.
2.2.3. MAE under subcritical conditions
Subcritical extraction was performed in a SynthWave, a mul-
timode pressure-resistant MW reactor working at 2.45 GHz
(Milestone, Bergamo, Italy). Extraction was performed in a
PTFE (Teflon) vessel with 10 g of sample in one of two differ-
ent solvents (ethanol 60% v/v (MAE-sub EtOH 60%) or distilled
water (MAE-sub H2 O), in a nitrogen atmosphere at 20 bar for
20 min. MW extraction was operated in temperature-control
mode and kept at 150 ◦ C.
2.2.4. Combined MW/US extraction
In this hybrid technology, a Pyrex® US horn (Danacamerini,
Turin, Italy) was inserted into a MicroSynth oven (Milestone,
Bergamo, Italy) from the top. The extraction was performed
using 6 g of sample in ethanol 60% v/v at 20 kHz for 20 min. MW
extraction was operated under temperature control at 50 ◦ C.
2.2.5. US-assisted extraction (UAE)
A probe system that was equipped with a titanium horn (ø
=15 mm) (Danacamerini, Turin, Italy) was used for extraction.
The sample (6 g) was extracted using two different solvents:
namely ethanol 60% v/v (UAE - EtOH 60%) or a ternary mix-
ture of hexane/ethanol/water (30:49:21) (UAE - hex/EtOH/ H2 O).
Samples were sonicated at 20 kHz, 100 W at a temperature of
around 40 ◦ C for 20 min.
2.3. Extract Characterisation
The peculiar nature of the extracts, which contain hydrophilic
and lipophilic compounds, required the use of a double liquid-
liquid extraction technique with hexane and methanol/water
(60:40, v/v). The mixture was vortexed for 2 min and cen-
trifuged at 3500 rpm for 5 min. The resulting hydrophilic
phase was used to analyse the total phenolic compounds
and antioxidant capacity, while the lipophilic phase was used
to determine the lipid-content and fatty-acid profiles. The
results were expressed relative to dry weight.
2.3.1. Total phenolic content (TPC)
The total phenolic content of the extracts was determined
using the Folin-Ciocalteu spectrophotometric method (Cicco
et al., 2009). A 1 mL aliquot of the extract solution (2 mg/mL)
was mixed with 250 ␮L of the Folin-Ciocalteu reagent (diluted
in distilled water 1:1), 500 ␮L of 10% sodium carbonate and
4 mL of distilled water. The solution was gently shaken and
then allowed to stand for 25 min at room temperature. Absorp-
tion measurements were performed at 740 nm using a Cary 60
UV-VIS spectrophotometer (Agilent Technologies, Santa Clara,
USA). Gallic acid (5 to 160 mg/L) was used to prepare the cali-
bration curve. The TPC was expressed as milligrams of gallic
acid equivalent per gram of extract and per gram of matrix.
2.3.2. Antioxidant-activity evaluation
Antioxidant activity was determined using the DPPH radical-
scavenging method, according to the protocol described in
Brand-Williams et al. (1995). Firstly, a DPPH solution, with
226 Food and Bioproducts Processing 1 1 7 ( 2 0 1 9 ) 224–230
Fig. 1 – Extraction yields and total phenolic content (TPC) expressed as g/100 g of pumpkin seeds over dry weight (DW),
obtained using the various extraction methods.
absorption of between 0.45 and 0.55, was prepared. The extract
was sequentially diluted in methanol (from 0.04 to 1.2 mg/mL)
and mixed with the DPPH solution. After 20 min in the dark,
the absorbance was measured at 517 nm using a Cary 60
UV-VIS spectrophotometer (Agilent Technologies, Santa Clara,
USA). The EC50 value (concentration that is able to promote
the bleaching of DPPH in 50%) was calculated using Microsoft
Visual Basic 6.0 software (Microsoft Corporation, Redmond,
WA, USA) and the results were expressed mg dry extract per
mL solution.
2.3.3. Determination of fatty acids
The organic phase containing the lipids was dried under nitro-
gen flow and derivatised as fatty-acid methyl esters (FAME)
according to the method described by Hartman and Lago
(1973). In a tube containing the lipid mass and 1 mg of inter-
nal standard methyl heptadecanoate (C17:0 Me), 1 mL of KOH
in methanol (0.4 mol/L) was added and heated at 100 ◦ C for
10 min. Afterwards, 3 mL of sulfuric acid in methanol (1 mol/L)
was added and heated at 100 ◦ C for 10 min. The mixture was
cooled and 2 mL of hexane was added and homogenised in
a vortex for 1 min. The upper part was collected for analysis
in gas-chromatography equipment (7820A, Agilent Technolo-
gies, Santa Clara, USA), which was equipped with a flame
ionisation detector (GC-FID). The injection was performed
using 1 ␮L of sample, which was separated in a capillary
column (Mega WAX, 30 m ×0.25 mm ×0.25 ␮m, Mega S.r.l., Leg-
nano, Italy). FAME identification was performed by comparing
the retention time of the analytes with the standard C8 – C24.
2.3.4. Principal component analysis (PCA)
An exploratory analysis of data using PCA was performed
using the Pirouette 3.11 software (Infometrix Co., USA). Before
analysis the data matrix was auto-scaled for each variable
assuming the same weight for all variables.
3. Results and discussion
3.1. Extraction yield
Firstly, the effect of the different technologies on the extrac-
tion yield from pumpkin seeds was evaluated and then
compared with that of the conventional method, as can
be seen in Fig. 1. The extraction yields ranged from 9.86
to 35.57 g/100 g of dry weight pumpkin seeds. Conventional
extraction using hexane gave the highest value of 35.57 g/100 g
of seeds, and mainly favoured the recovery of lipophilic
compounds. While the hydrophilic fraction, recovered using
ethanol 60% v/v, only presented a 11.1 g/100 g yield. These
results are in agreement with the literature, confirming that
pumpkin seeds are a rich source of oil. In general, the oil con-
tent present in pumpkin seeds is around 25 to 50 % (Younis
et al., 2000; Silva and Jorge, 2014; Cuco et al., 2019).
Now moving to the non-conventional techniques only, UAE
using the ternary mixture (Hex/EtOH/H2 O) gave the highest
extraction yield, of 35.06 g/100 g of seeds, with 26.10 being in
the apolar phase and 8.91 in the polar phase. This procedure
was able to recover 73% of the lipophilic compounds and 80%
of the hydrophilic ones in a single extraction step of 20 min.
This required 4 h using conventional extraction. This result
may be related to the high ultrasonic energy used and the
two organic solvents, hexane and ethanol 60%, which have
a considerable ability to recover lipophilic and hydrophilic
compounds, respectively. These results are in agreement with
those reported by Grillo et al., 2019, who also found higher
extraction yields when using UAE with the same ternary mix-
ture.
The application of MAE under subcritical conditions also
gave interesting extract yields using green solvents. In this
case, subcritical EtOH 60% furnished 23.92 g/100 g and sub-
critical water, 22.21 g/100 g. These results may be due to the
high temperature and pressure conditions, which were able
to improve solvent solubility and mass transfer. These con-
ditions enable subcritical extraction to recover a large range
of compounds with both polar and nonpolar characteristics,
explaining the high extraction yield presented in this study
(Nastić et al., 2018; Herrero et al., 2015). Castejón et al. (2018)
also found higher yields from echium seeds using subcritical
ethanol than using UAE and MAE with ethanol.
Extraction using UAE and combined MW/US with EtOH 60%
gave the lowest yield values, 15.59 and 9.86 g/100 g of seeds,
respectively. Since these procedures are performed in open
vessels, low extraction temperatures of around 40-50 ◦ C were
required. In this way, it is possible that ethanol is not suit-
 Food and Bioproducts Processing 1 1 7 ( 2 0 1 9 ) 224–230 227

The antioxidant-activity results confirm the efficiency of

Table 1 – Total phenolic content and antioxidant activity
in the extracts from pumpkin seeds. MAE in extracting phenolic compounds under subcritical
conditions. Furthermore, the extracts obtained using this
Extraction Total phenolic content DPPH EC50
technique also showed higher antioxidant activity. The DPPH
method
mg/g matrix mg/g extract mg/ml extract EC50 values were 0.51 mg/ml of extract using MAE subcriti-
cal ethanol and 0.79 mg/ml of extract using MAE subcritical
CE - hex 1.18 3.49 -
CE - EtOH 60% 3.93 42.42 1.88 water. For other techniques, UAE, MAE and MW/US, the antiox-
UAE - hex/EtOH/H2 O 3.02 39.41 1.52 idant activity of the extracts followed the behaviour MAE > UAE
UAE - EtOH 60% 2.04 34.19 1.96 with ternary mixture > UAE with ethanol > MW/US. With the
MW/US - EtOH 60% 2.43 31.10 2.07 exception of the extracts obtained using UAE with ethanol
MAE - EtOH 60% 2.61 35.02 1.23 and MW/US, all of the others presented higher antioxidant
MAE - sub EtOH 60% 8.12 83.95 0.51
activity than those obtained using conventional extraction
MAE - sub H2 O 8.71 55.52 0.79
(1.77 mg/ml of extract).

able for efficient extraction under these conditions. This is
confirmed by studies that have reported the effect of tempera-
ture on the improvement of extraction yields from seeds using
ethanol, both when obtaining oils and phenolic compounds
(Castejón et al., 2018; Zardo et al., 2019).
3.2. Total-phenolic content and antioxidant capacity
A comparison of the different extraction procedures, in terms
of total phenolic content and antioxidant activity, are shown
in Table 1. The phenolic content is reported as the amount of
phenolics per gram of extract and also considers the amount
of phenolics extracted per gram of matrix (dry weight).
It is possible to note that MAE under subcritical condi-
tions was the most efficient extraction method for phenolic
compounds. In terms of phenolic content recovered from the
matrix, the values were similar for both subcritical solvents,
ethanol and water, 8.12 and 8.71 mg gallic acid equivalents/g
(GAE/g) matrix, respectively. However, the extract obtained
using subcritical ethanol was shown to be more concentrated
in phenolic content as it gave 83.95 mg GAE/g extract, while
subcritical water gave 55.52 mg GAE/g extract. These results
were higher than those obtained for conventional extraction;
3.93 mg GAE/g matrix and 42.42 mg GAE/g extract. This can be
attributed to the reduction of solvent polarity under subcriti-
cal conditions, which promotes the solubilisation of a number
of phenolic compounds (Luo et al., 2018; Corazza et al., 2018).
If we compare MAE, UAE and MW/US, we can see that these
techniques showed similar phenolic-compound extraction
efficiencies. However, UAE with the ternary mixture presented
slightly higher values, with 39.41 mg GAE/g matrix and 3.02 mg
GAE/g extract, while UAE using only ethanol 60% had the
lowest phenolic values, presenting 2.04 mg GAE/g matrix and
34.19 mg GAE/g extract. From these results, we can see that the
ternary mixture had a significant effect on UAE. This result is
probably related to the easier recovery of phenolic compounds
after extraction that this procedure allows.
3.3. Fatty-acid composition
The fatty-acid profiles from pumpkin seeds, as analysed by
GC-FID, and total lipid content are presented in Table 2. A com-
parison of the non-conventional methods shows that there is
a significant difference in the amount of lipids extracted, rang-
ing from 26.10 to 3.38%. The highest lipid content was achieved
using UAE with the ternary mixture, followed by MAE with sub-
critical and non-subcritical ethanol. MW/US and MAE using
subcritical water displayed poor-lipid extraction efficiencies.
Similar fatty-acid compositions were found for all the
samples. The results show that the predominant fatty acids
are linoleic acid C18:2n-6 (36.31 – 45.79%), oleic acid C18:1n-
9 (23.69 – 35.88%), palmitic acid C16:0 (15.17 – 19.12%) and
stearic acid C18:0 (9.26 – 11.47%). Pumpkin seeds are there-
fore characterised by high percentages of unsaturated fatty
acids, representing around 71% of the total fatty acids. Similar
results for the fatty-acid compositions from pumpkin seeds
were found by Salgin and Korkmaz (2011), Silva and Jorge
(2014) and Cuco et al. (2019).
Although the fatty-acid profiles were similar, some changes
were observed in the different extraction methods. The con-
ventional technique presented 28.82% of saturated fatty acids
and 71.14% of unsaturated fatty acids, of which 39.78% was
C18:2 n-6 and 31.35% C18:1 n-9. A similar profile was obtained
using MAE with ethanol 60% as well as UAE with ethanol 60%
and the ternary mixture. However, UAE was able to extract
slightly higher amounts of unsaturated fatty acids; 72.51%
with the ternary mixture and 72.20% with ethanol 60%. This
procedure enabled the recovery of a good amount of both
linoleic acid (C18:2n-6) and oleic acid (C18:1n-9), with around
35% of each being extracted. On the other hand, MW/US and
MAE in subcritical conditions presented lower sums of unsat-
urated fatty acids, although a higher percentage of C18:2n-6
(around 45%) was observed.
Independently of the extraction method used, it can be
observed that pumpkin seeds are a rich source of linoleic acid

Table 2 – Fatty-acid composition, as determined by GC-FID.

Fatty acids (%) CE. hexane UAE Hex/EtOH/H2 O UAE EtOH 60% US/MW EtOH 60% MAE EtOH 60% MAE Sub. EtOH 60% MAE Sub. H2 O

C14:0 0.14 0.11 0.17 0.19 0.15 0.20 0.30

C16:0 17.04 15.17 17.11 18.47 17.20 19.00 19.12
C18:1 31.39 35.15 35.88 26.28 32.35 24.56 23.69
C18:2 39.83 37.45 36.31 44.97 39.15 45.68 45.79
C18:0 10.70 11.47 9.63 9.26 9.81 9.56 10.40
C20:0 0.73 0.00 0.69 0.00 1.12 0.71 0.51
C22:0 0.17 0.66 0.21 0.82 0.22 0.30 0.18
Lipid content* 35.57 26.10 9.71 3.38 10.81 10.61 4.08

∗
values expressed in g/100 g of pumpkin seeds.
228 Food and Bioproducts Processing 1 1 7 ( 2 0 1 9 ) 224–230
Fig. 2 – Plots of (A) scores (samples) and (B) loading
(variables) obtained in principal component analysis,
showing the two major principal components of the
analysis extracts obtained from pumpkin seeds using the
various extraction methods.
C18:2n-6 and oleic acid C18:1n-9. These compounds are con-
sidered to be important fatty acids as they provide relevant
effects on human health, especially in the prevention of can-
cers and cardiovascular diseases. However, these seeds can
also provide a promising matrix for use as nutraceutical prod-
ucts.
3.4. Exploratory multivariate analysis
Principal Components Analysis (PCA) was used to better
visualise the effects of the procedures on the extraction of
hydrophilic and lipophilic compounds from pumpkin seeds
(Fig. 2). The two major components (PC1 and PC2) together
incorporated 81.88% of the overall variability. PC1 separated
the samples mainly according to the solvent used for extrac-
tion and consequently their ability to extract hydrophilic and
lipophilic compounds. UAE with the ternary mixture and CE-
hex were located on the opposite side to CE – EtOH 60%.
PC2 clearly separated the samples according to extrac-
tion technology. The bottom quadrant contains the samples
obtained by MAE under subcritical conditions, which are char-
acterised by higher phenolic content and antioxidant capacity.
These samples also had higher percentages of PUFA, as did
those that were extracted using MAE with EtOH 60%. The sam-
ples extracted by UAE can be also be grouped, and this is
Fig. 3 – US horn employed for pumpkin seeds in typical lab
test (left) and 10-fold larger-scale extraction (60 g seeds,
right).
probably related to the higher percentages of MUFA. It was
possible to separate the samples obtained by conventional
extractions in the upper part of the quadrant.
3.5. Extraction scale-up
In view of the promising results obtained by UAE with the
ternary phase, a scale-up of this procedure was performed.
For this purpose, the same extraction protocol was applied
on a 10-fold larger scale of sample and solvent (Fig. 3). It is
worth to mention the challenging scaling-up of UAE and MAE,
because the variability from one equipment to another can be
significant, and inhomogeneous distribution of pressure field
by change the scale could impact the energy transfer efficien-
cies (Gomes et al. (2018)). Therefore, the increase of scale could
be used as a strategy to test the potentiality of the proposed
process towards its ultimate application in pilot settings. This
scale-up confirmed the previous results in terms of yield and
extract composition, phenolic content, antioxidant capacity
and fatty-acid profile, as shown in Table 3. It may be noted that
the synergism of UAE with the ternary solvent mixture was an
efficient in one-step extraction of compounds with different
polarity. In terms of industrial application, it could provide
numerous advantages over conventional procedures for being
a fast and efficient process that allows the enhancement of
productivity with energy and cost saving. Big scale extractions
require flow mode UAE as reported by Alexandru et al. (2013).
The solvents employed in this process are already hugely
used in industrial extraction in particular in food processing.
Common facilities enable hexane recycling for a cost-effective
process with a low environmental impact. The scale-up fea-
sibility indicates the potential gains to be made by designing
non-conventional methods for larger scale applications.
 Food and Bioproducts Processing 1 1 7 ( 2 0 1 9 ) 224–230
Table 3 – Results obtained by the scaled-up extraction
using UAE with ternary phase.
Yield (mg/100 g DW) 34.78
Lipid content (mg/100 g DW) 25.48
Fatty acids (%)
C14:0 0.12
C16:0 15.46
C18:1 35.5
C18:2 37.24
C18:0 11.17
C20:0 0
C22:0 0.51
TPC (mg/g DW) 3.69
DPPH EC50 (mg/mL extract) 1.48
4. Conclusion
The present investigation on the extraction of pumpkin seeds
compares non-conventional techniques with classic methods
and has resulted in significant differences in extract yields and
compositions being observed. In general, UAE with the ternary
mixture gave the most promising results, mainly in terms of
extraction yield, total phenolic content and fatty-acid com-
position. With a single process step of 20 min, this procedure
can be considered a viable and fast alternative for the recov-
ery of valuable compounds from pumpkin seeds. A scaling of
factor 10 has confirmed these results. It is also worth mention-
ing that MAE under subcritical conditions provided excellent
results using only green solvents (ethanol and water); this
technique was able to grant a good extract yield with dou-
ble the amount of phenolic compounds, higher antioxidant
capacity and a higher linoleic-acid amount.
Acknowledgment
The authors are grateful to the Coordenação de
Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
for supporting to DF the doctoral fellowship CAPES/PDSE/
88881.188528/2018-01 and to the University of Turin for the
financial support (Ricerca locale 2018).
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