Food Chemistry 286 (2019) 354–361

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

Hass avocado (Persea americana Mill.) oil enriched in phenolic compounds T

and tocopherols by expeller-pressing the unpeeled microwave dried fruit
⁎
Isabelle Santanaa, , Vanessa Naciuk Castelo-Brancob, Bárbara Mello Guimarãesc,
Laís de Oliveira Silvad, Vanessa Oliveira Di Sarli Peixotod, Lourdes Maria Corrêa Cabrale,
⁎
Suely Pereira Freitasc, Alexandre Guedes Torresd,
a
Institute of Nutrition, Rio de Janeiro State University, Brazil
b
School of Pharmacy, Federal Fluminense University, Brazil
c
Chemical Engineering Laboratory, Federal University of Rio de Janeiro, Brazil
d
Laboratory of Food Science and Nutritional Biochemistry, Institute of Chemistry, Federal University of Rio de Janeiro, Brazil
e
Embrapa Food Technology, Rio de Janeiro, Brazil

A R T I C LE I N FO A B S T R A C T

Keywords: This study investigated how the quality of avocado oil is affected by the fruit ripening stage and peeling, and the
Hass avocado oil drying process used. Expeller pressed avocado oils were obtained from unripe or ripe pitted avocados after
Drying technology drying peeled or unpeeled pulps by convection oven, microwave or freeze-drying. Oils from the unpeeled mi-
Oxidative stability crowave dried pulp (from unripe or ripe avocados) showed the highest induction period (54.2–83.6 h) and
Ripening
antioxidant capacity (4.07–5.26 mmol TE/kg), and high amounts (mg/100 g) of α-tocopherol (11.6–21.0), β-
Phenolic compounds
carotene (0.49–0.65) and chlorophyll (44.3–54.0), and unsaponifiable matter (2.48–2.99 g/100 g). Pulp drying
process and avocado (un)peeling were the major contributors to the induction period (R2 = 0.61; p = 0.0139)
and antioxidant capacity (R2 = 0.62; p = 0.011), and the oils from microwave dried unpeeled pulp were those
that presented the best performance. The phenolic composition of these oils improved with ripening and keeping
the peel during the pressing process.

1. Introduction
Avocado oil stands out as one of the fruit’s derived products with the
highest market values, with appreciable flavor and color, and with
varied positive health effects (Duarte, Chaves, Borges, & Mendonça,
2016). The pressed oil can be consumed pure and added cold to salads,
or used in food preparations, or in formulations for pharmaceutical and
cosmetic applications. In these last cases, the unsaponifiable matter,
containing carotenoids, chlorophylls and tocopherols, and the phenolic
compounds are of major interest, for their antioxidant and anti-in-
flammatory properties (Zhang, Huber, & Rao, 2013; Kosínska et al.,
2012).
Avocado oil oxidative stability determines its quality during storage
and heat processing, eventually affecting flavor, color, and nutrient and
bioactives composition. Oil stability on its side is affected by the ori-
ginal avocado oil composition, and may be affected by processing fac-
tors, such as pulp drying and oil extraction technologies (Krumreich,
Borges, Mendonça, Jansen-Alves, & Zambiazi, 2018; Santana, dos Reis,
Torres, Cabral, & Freitas, 2015; Costagli & Betti, 2015). However, there
is scarce information concerning the influence of the avocado fruit
characteristics, for instance ripening stage, on the quality of the ex-
tracted oil (Villa-Rodríguez, Molina-Corral, Ayala-Zavala, Olivas, &
González-Aguilar, 2011). Avocado matures when attached to the tree,
increasing in size and weight in parallel to exponential lipid accumu-
lation in the mesocarp and losing approximately 30% of the fruit’s
water (Ozdemir & Topuz, 2004). Oleic acid content increases at higher
rates compared to the other fatty acids, although large differences in
fatty acid composition can be observed even for the same cultivar de-
pending on the growing areas and harvesting season (Carvalho, Bernal,
Velásquez, & Cartagena, 2015; Duque, Londoño-Londoño, Álvarez, Paz,
& Salazar, 2012). Moreover, plant secondary metabolites also accu-
mulate, increasing the contents of unsaponifiable matter and phenolic
compounds (Zhang et al., 2013; Kosínska et al., 2012).
Commercially, avocado fruit ripening starts after harvesting, and
generally net lipid synthesis does not occur after that (Meyer & Terry,
2008). As a climacteric fruit, postharvest ripening induces physiological

⁎
Corresponding authors at: Instituto de Química, Universidade Federal do Rio de Janeiro, Av. Athos da Silveira Ramos, 149, CT, Bloco A, 528A, 21941-909 Rio de
Janeiro, RJ, Brazil (A.G. Torres).
E-mail address: torres@iq.ufrj.br (A.G. Torres).

https://doi.org/10.1016/j.foodchem.2019.02.014
Received 10 September 2018; Received in revised form 6 February 2019; Accepted 8 February 2019
Available online 12 February 2019
0308-8146/ © 2019 Elsevier Ltd. All rights reserved.
I. Santana, et al.
changes culminating in full avocado ripening after 3–8 days (Wang,
Zheng, Khuong, & Lovatt, 2012; Ozdemir & Topuz, 2004), when at
21–25 °C. Regarding Hass avocado, ripening modifies the skin color
from green to purple/black, due to the accumulation of anthocyanins
(Cox, Mcghie, White, & Woolf, 2004) and chlorophyll degradation
(Villa-Rodríguez et al., 2011; Ashton et al., 2006). Therefore, oils ex-
tracted from avocados with different ripening stages could present
distinct composition. Nevertheless, the extraction of oil from unripe
avocados has been poorly addressed so far (Ashton et al., 2006; Mostert,
Botha, Plessis, & Duodu, 2007). Besides eliminating the demanded time
for ripening, processing the unripe avocados may limit postharvest is-
sues linked to microbiological and biochemical agents leading to un-
wanted changes in chemical and physical characteristics. Furthermore,
the contents of avocado fruit secondary metabolites, several of which
present antioxidant activity and potential bioactivity, might be en-
hanced in the oil from unripe avocados, but this was not investigated
previously.
Another factor that could enhance avocado oil contents of natural
antioxidants and bioactives is keeping the fruits’ peel or other parts,
besides the pulp, during oil extraction. Antioxidant capacity and con-
tents of phenolic compounds are much higher in avocado peel than in
the fruit’s pulp (Kosínska et al., 2012; Rodríguez-Carpena, Morcuende,
Andrade, Kylli, & Estévez, 2011; Rodríguez-Carpena, Morcuende, &
Estévez, 2011; Wang, Bostic, & Gu, 2010). Even though presenting low
lipid amounts, using the peel for extraction might reduce extra waste
disposal, while allowing the potential enhancement of antioxidants in
the extracted oil.
Our aim was to investigate the effects of Hass avocado ripening
stage and peeling on the composition and quality of the oil obtained by
expeller pressing, in combination with the effects of the fruit drying
techniques adopted, either in a convection oven with forced air circu-
lation (60 °C), in a microwave-oven or by freeze-drying.
2. Materials and methods
2.1. Reagents and standards
Tocopherols (α-, β-, γ- and δ-) and phenolic compounds standards
(quercetin, 3,4-dihydroxyphenylacetic, 3,4-dihydroxybenzoic, gallic,
vanillic, syringic, p-coumaric, m-coumaric, ferulic, ellagic, trans-cin-
namic acids), fatty acid methyl esters (37 FAME mix), 6-hydroxy-
2,5,7,8 tetramethylchromane-2-carboxylic acid (trolox) and 2,2′-azino-
bis (3-ethylbenzothiazoline)-6-sulfonic acid (ABTS) were purchased
from Sigma-Aldrich (São Paulo, Brazil). β-Carotene was isolated from
carrot extract by open column chromatography and chlorophylls a and
b were obtained from spinach, as described by Silva et al., 2017. All
pigment standards showed purity grades > 95%, determined by HPLC-
PDA. All solvents used were of chromatography grade (Tedia, Rio de
Janeiro, Brazil) and ultrapure water (Milli-Q system, Millipore, Bed-
ford, MA, USA) was used throughout the experiments.
2.2. Raw material
Avocado (Persea americana Mill. cv. Hass) fruits grown in Carandaí
(Minas Gerais, Brazil) were purchased from a central wholesale dis-
tributor of fruits and vegetables (CADEG, Rio de Janeiro, Brazil). The
fruits were acquired in the unripe stage after 1 day of harvest during
winter (July), having reached the horticultural maturity defined by the
fruit dry matter content (25.7 ± 0.4 g/100 g). Avocado fruits were
stored in a cold chamber at 4 °C immediately after purchase. Ripening
was carried out in a darkroom at 20 ± 2 °C, and the ideal point of
ripeness was based on the skin color change from green to purplish/
black (> 75% of the skin) and pulp softening by hand (Cox et al., 2004;
Osuna-García, Doyon, Salazar-García, Goenaga, & González-Durán,
2010), which occurred in 6–7 days. Fruits were daily turned approxi-
mately 90° around their longer axis to keep air exposure uniform and to
355
Food Chemistry 286 (2019) 354–361
avoid deformations resulting from fruit softening during ripening.
Avocados were sanitized with chlorine solution (100 mg/kg) prior to
processing.
2.3. Hass avocado pulp processing and oil extraction
Before oil extraction, the pulps from unripe (two days after harvest)
and ripe avocados were classified into four groups: 1) unripe and
peeled; 2) unripe and unpeeled; 3) ripe and peeled and 4) ripe and
unpeeled. All avocados were pitted before processing. Resulting pulps
were crushed in a pilot scale cutter (Geiger; São Paulo, Brazil) to obtain
small irregular pieces (up to 7 mm), in the case of unripe avocado pulps,
or a homogeneous creamy mass when ripe avocado pulp was processed,
and in this case peel fragments with up to 4 mm were dispersed in this
mass. Subsequently, avocado pulp was dehydrated by convection oven
with forced air circulation at 60 °C, freeze-drying or microwave-oven
drying, as detailed below.
For convection oven drying (60 °C), fruit pulp was spread on
stainless steel trays (80 × 100 cm) in layers up to 10 mm thick, and
placed in a forced air circulation oven, until reaching equilibrium
moisture, which took 18.0 ± 0.3 h. For freeze-drying, pulp was dried
in a freeze-dryer (Edwards Pirani 501, West Sussex, United Kingdom) at
−40 °C for 8–10 h. For microwave-oven drying, pulp was spread in the
rotating plate and dried for 19.0 ± 0.4 min, pausing after 10 min to stir
the sample, in a domestic microwave oven (Brastemp, 2450 MHz,
1140 W; Brazil) set at 80% power. At the end of the drying processes,
equilibrium moisture was between 5.4 and 11.7 g/100 g
(Supplementary Table 1). All drying processes were performed in tri-
plicate and dried pulps were vacuum packaged in dark containers and
stored at −18 °C until oil extraction.
Crude oil extraction was conducted in triplicate in a continuous
expeller (Oekotec, CA59G, Germany) at room temperature (25 ± 2 °C).
Oil temperature exiting the expeller, measured by a thermo-par coupled
to a digital thermometer varied between 40 and 45 °C, thus character-
izing the samples as cold pressed. Crude oils were decanted and the
liquid portion was separated from the sludge, resulting in the following
oil groups: 1) oil from unripe and peeled avocado, oven dried at 60 °C;
2) oil from unripe and unpeeled avocado, oven dried at 60 °C; 3) oil
from ripe and peeled avocado, oven dried at 60 °C; 4) oil from ripe and
unpeeled avocado, oven dried at 60 °C; 5) oil from unripe and peeled
freeze-dried avocado; 6) oil from unripe and unpeeled freeze-dried
avocado; 7) oil from ripe and peeled freeze-dried avocado; 8) oil from
ripe and unpeeled freeze-dried avocado; 9) oil from unripe and peeled
microwave-dried avocado; 10) oil from unripe and unpeeled micro-
wave-dried avocado; 11) oil from ripe and peeled microwave-dried
avocado; and 12) oil from ripe and unpeeled microwave-dried avocado.
All crude oils were stored at −18 °C, in the dark, in nitrogen atmo-
sphere until analysis.
2.4. Assessment of avocado oil quality
2.4.1. Fresh oil quality
The quality of fresh crude avocado oils was determined by con-
ventional quality indices such as acid (method Cd3d-63), peroxide
(method Cd8-53) and iodine (method Cd1c-85) values and refractive
index (method CC7-25) as described by AOCS (2012). Refractive index
was determined in a digital refractometer (Atago® PAL-BX/RI) at 25 °C.
Temperature correction to 40 °C was provided by Eq. (1)
R1 = R2 + K (T 2 − T 1) (1)
where, R1 = refractive index adjusted to 40 °C; R2 = refractive index
read in the apparatus; K = 3.885 × 10−4; T2 = reading temperature;
T1 = adjustment temperature (40 °C).
2.4.2. Oxidative stability and total antioxidant capacity (TAC)
Oxidative stability was determined by Rancimat 743 (Metrohm AG,
I. Santana, et al.
Herisau, Switzerland) where 3 g of sample were oxidized at 110 °C with
an air flow of 20 L/h. Electrical conductivity (μS) was measured by
electrodes immersed in 50 mL of distilled water and the induction
period (IP) was determined by the instrument’s software package as the
time (h) elapsed until a sharp increase in water conductivity due to the
building-up of oxidation products. TAC was determined spectro-
photometrically (UV-1800, Shimadzu, Japan) by TEAC assay exactly as
previously described (Castelo-Branco & Torres, 2012), and results were
expressed as mmol of trolox equivalents/kg oil (mmol TE/kg).
2.5. Analysis of avocado oil chemical components
2.5.1. Fatty acid composition by GC-FID
Fatty acid methyl esters (FAME) obtained after direct transester-
ification of samples (Lepage & Roy, 1986) were analyzed using a gas
chromatograph (GC-2010, Shimadzu, Japan), an Omegawax-320
(30 m × 0.32 mm, 0.25 μm; Supelco, Co., EUA) column, flame ioniza-
tion detector (FID), and with split injection at 1:30 split ratio. Column
temperature was programmed as follows: 160 °C constant for 2 min,
followed by a gradient of 2.5 °C/min up to 190 °C, kept constant for
5 min, followed by a temperature gradient of 3.5 °C/min up to 220 °C,
remaining constant for 15 min. The injector and detector were operated
at 260 °C and 280 °C, respectively, and He was used as carrier gas. Peak
identities in samples’ chromatograms were determined by comparison
of relative retention times with those of a commercial mixture of fatty
acid methyl esters standards (37 FAME-mix; Sigma-Aldrich; São Paulo,
Brazil). Quantification was performed by internal normalization after
correction of peak areas by their respective theoretical correction fac-
tors (Wolf, Bayard, & Fabien, 1995). Fatty acid content in the samples
was expressed as g/100 g of total fatty acids.
2.5.2. Unsaponifiable matter
Samples (2.5 g) were cold-saponified with saturated KOH in ethanol
95% (v/v) for 30 min. The supernatant was collected and first washed
with cyclohexane and aqueous ethanol 50% (v/v), followed by washing
with ethanol 50% (v/v) and distilled water. The solvent was evaporated
until dryness, and the remaining mass weighed to the nearest 0.1 mg.
Results were expressed as g/100 g oil, adapted from Hartman, Viana,
and Freitas (1994).
2.5.3. Analysis of tocopherols, β-carotene and chlorophyll by HPLC-PDA
The contents of tocopherols (α, β, γ and δ), β-carotene and chlor-
ophylls in avocado oils were determined simultaneously by normal-
phase HPLC with a photon-diode array detector (PDA), as previously
described by Silva et al. (2017). Briefly, the oil was dissolved in n-
hexane, centrifuged (10,000×g, 5 min) and the supernatant was fil-
tered through a PTFE syringe filter (0.45 μm) and injected (20 μL) in the
HPLC system (Shimadzu, Kyoto, Japan) equipped with a quaternary
pump (LC-20AT), system controller (CBM-20A), degasser DGU-20A5,
and SPD-M20A PDA detector. Chromatographic separation was
achieved using a normal-phase silica column (ZORBAX Rx-Sil, 5 µm,
4.6 mm × 250 mm, Agilent Technologies, USA) with isocratic elution
(n-hexane:2-propanol; 99:1, v/v) at 1.0 mL/min. Tocopherol isoforms,
β-carotene and chlorophyll a were detected at 295, 450 and 665 nm,
respectively. The samples chromatographic peaks identities were de-
termined based on standards’ retention times, and UV/Vis spectra, and
confirmed by perfect coelution and spectra matching with standards.
Contents were calculated by using calibration curves ranging from 0.5
to 3.0 μg/mL for tocols (R2 > 0.9979; p < 0.0001; LODmax = 0.091
μg/mL; LOQmax = 0.30 μg/mL), 0.5 to 5.0 μg/mL for β-carotene
(R2 = 0.9965; p < 0.0001; LODmax = 0.32 μg/mL; LOQmax = 1.08 μg/
mL) and 6.25 to 100.0 µg/mL for chlorophylls (R2 = 0.9983;
p < 0.0001; LODmax = 5.4 μg/mL; LOQmax = 17.81 μg/mL). Results
were expressed in mg/100 g oil for each tocopherol, β-carotene or
chlorophyll.
356
Food Chemistry 286 (2019) 354–361
2.5.4. Analysis of phenolic compounds by HPLC-PDA
Phenolic compounds were extracted from avocado oil by liquid–li-
quid extraction with aqueous methanol 80% (v/v) after dissolving the
oil in n-hexane (2:1, w/v). Extraction was repeated twice. The upper-
phases from the three extractions were combined, and the solvent was
evaporated. The residue was reconstituted with 3 mL of methanol and
analyzed by HPLC-PDA. The LC system used was the same described in
2.6.3 and chromatographic separations were achieved in a reversed-
phase column (C18, 4.6 mm i.d × 150 mm, 5 μm particle size;
Kromasil®) and the mobile phase consisted of a gradient of 0.3% aqu-
eous formic acid (eluent A), methanol (eluent B) and acetonitrile
(eluent C) at flow rate of 1.0 mL/min as follow: 24% (B) at 8 min, 28%
(B) at 18 min, 33% (B) at 30 min, 65% (B) at 60 min, followed by re-
equilibration for 15 min. Eluent C was kept constant at 1% during
analysis. Phenolic compounds were monitored by the PDA detector
from λ = 190 to 370 nm. Chromatographic peaks in samples were
identified as phenolic compounds by analytical comparisons with
commercial standards behavior, as follows: retention time, UV-spectra
analysis, perfect co-elution with phenolic standards, and perfect over-
lapping of the co-eluted peaks’ UV-spectra. Quantitative analysis was
based on external calibration curves with concentrations ranging from
1.0 to 20.0 μg/mL (R2 > 0.9965, p < 0.0001; LODmax = 0.42 μg/mL,
LOQmax = 1.41 μg/mL). Only the oil samples with the highest oxidative
stability and TAC values were analyzed.
2.6. Statistical analyses
All results are presented as mean ± standard deviation of triplicate
processes. Multivariate analysis of variance (MANOVA) followed by
Fischer LSD post-hoc test was used to compare the composition of the
oils obtained, according to the three investigated factors: avocado
peeling (peeled or unpeeled) and ripening stage (mature-unripe or
mature-ripe), and pulp drying processes applied (convection oven,
microwave-oven or freeze-drying). To investigate the effects of pro-
cessing factors (avocado ripeness, fruit peeling, and pulp drying pro-
cess) on parameters of fresh avocado oil quality, data was analyzed in a
Multi-factor categorical design matrix, with a desirability function, in
order to determine the best combination of factors that enhances avo-
cado oil quality. Pearson’s correlation analysis was used to investigate
associations between continuous variables of oil composition and to
select variables to insert into a general linear model (GLM) matrix. The
GLM, which allowed including continuous and categorical factors in the
models, was used to investigate the major determinants of oil quality
variability. The dependent variable in the model was induction period
(IP) of oils and independent variables were α-tocopherol, β-carotene and
chlorophyll contents, as continuous variables; and ripening stage, peeled/
unpeeled and pulp drying process, as categorical factors. All analyses were
performed using Statgraphics v. 18 (Manugistics, EUA) and P va-
lues < 0.05 were considered as statistically significant.
3. Results and discussion
3.1. Oil yield, avocado oil quality, oxidative stability and antioxidant
capacity were affected by the fruit ripening stage, peeling, and pulp drying
technology
Avocado oil yield was, in general, lower for the fruits dried by
freeze-drying, especially the unripe avocado (Supplementary Table 1).
Additionally, processing the unripe fruit markedly reduced oil yield
independently of the fruit-drying process used, except for the micro-
wave-dried pulp, that was not affected by avocado ripening. The
highest values of oil yield were observed for oven-dried unpeeled ripe
avocados, which showed yield values roughly 20% higher (p < 0.05; t-
test) than those of unripe microwave-dried fruits, either peeled or not
(Supplementary Table 1). The fruit ripening stage and peeling, and the
drying process used had influence on the quality of pressed oils
I. Santana, et al. Food Chemistry 286 (2019) 354–361

Table 1
Acid value (mg KOH/g), iodine value, refractive index (at 40 °C) and peroxide value (mEq O2/kg oil) of pitted unripe and ripe avocado (Persea americana Mill. cv.
Hass) oils obtained from three pulp drying technologies.
Hass avocado characteristics and pulp drying technology used Expeller-pressed Hass avocado oil quality indices

Acid value (mg KOH/g) Iodine value Refractive index Peroxide value (mEq O2/kg oil)

Pitted unripe avocado

Peeled Oven drying (60 °C) 1.09 ± 0.03b 72.7 ± 1.77b 1.4598 ± 0.0002d,* 8.04 ± 0.06a,*
Microwave-oven 0.47 ± 0.02f 72.7 ± 0.44b 1.4601 ± 0.0006c,* 2.83 ± 0.05f,*
Freeze-drying 1.27 ± 0.14a 73.0 ± 1.81b 1.4581 ± 0.0002f,* 5.05 ± 0.21d,*
Unpeeled Oven drying (60 °C) 0.80 ± 0.08c 74.8 ± 1.37a 1.4602 ± 0.0002b,* 5.10 ± 0.43c*
Microwave-oven 0.53 ± 0.06e 74.8 ± 0.14a 1.4609 ± 0.0006a,* 2.90 ± 0.10e,*
Freeze-drying 0.67 ± 0.10d 75.6 ± 0.73a 1.4596 ± 0.0012e,* 5.14 ± 0.06b,*

Pitted ripe avocado

Peeled Oven drying (60 °C) 0.97 ± 0.11B 71.1 ± 1.53B 1.4596 ± 0.0007D 10.7 ± 0.21A
Microwave-oven 0.47 ± 0.02F 73.9 ± 0.70B 1.4596 ± 0.0003C 4.96 ± 0.30E
Freeze-drying 1.28 ± 0.03A 72.8 ± 0.88B 1.4586 ± 0.0002F 7.78 ± 0.12B
Unpeeled Oven drying (60 °C) 0.69 ± 0.04D 75.5 ± 1.47A 1.4601 ± 0.0001B 6.33 ± 0.51C
Microwave-oven 0.57 ± 0.03E 74.4 ± 0.47A 1.4602 ± 0.0001A 4.49 ± 0.16F
Freeze-drying 0.84 ± 0.12C 75.6 ± 0.45A 1.4590 ± 0.0010E 5.68 ± 0.60D

Results are expressed as mean ± standard deviation of triplicate processes. Different superscript lowercase letters in the same column indicate significant differences
between oils from unripe avocado. Different superscript capital letters in the same column indicate significant differences between oils from pitted ripe avocado.
* Significantly different from ripe avocado (peeled vs. peeled and unpeeled vs. unpeeled; for each drying process). MANOVA followed by Fischer’s LSD test
(p ≤ 0.05).

concerning acid, peroxide and iodine values and refractive index
(Table 1). Even so, all oils presented acid and peroxide values below the
maximum limits recommended for pressed oils (Codex standard for
named vegetable oils, amended 2013) (≤4.0 mg KOH/g oil and 15 mEq
O2/kg oil, respectively), indicating that no extensive hydrolytic or
oxidative degradation happened during avocado pulp processing and/
or oil extraction. In this sense, pressed oils from microwave dried pulps
presented the lowest acid and peroxide values in both ripening stages,
both from peeled and unpeeled fruits.
In parallel, iodine values varied from 71.1 to 75.6, only being in-
fluenced by peeling. Refractive indexes ranged from 1.458 to 1.465, at
40 °C, for all samples, which is in agreement with Mexican standards for
avocado oil (NMX-F-052-SCFI-2008, 2008).
Oxidative stability (expressed as IP, in hours), and TAC (by TEAC
assay), were influenced by ripening, fruit peeling and pulp drying
process used (Fig. 1). Oils from microwave dried pulp exhibited the
highest oxidative stability (Fig. 1A) and TAC values (Fig. 1B). Ad-
ditionally, keeping the avocado peel during expeller pressing clearly
improved these parameters for all extracted oils, especially those from
the microwave dried pulps, for which IP and TAC were approximately
1.6-fold higher in the oils from unpeeled avocados in relation to their
paired peeled samples.
In the present study, IP varied from 7.03 h in the oil from ripe
peeled oven-dried avocados to 83.6 h in the unripe and unpeeled mi-
crowave dried ones. Sensibly lower IP values were previously reported,
for instance 2.8 h (at 110 °C and 20 L/h) for solvent-extracted and re-
fined avocado oils (Knothe, 2013), and 3.93 and 5.24 for centrifuged
oils from Hass and Fuerte avocados (Ferrari, 2015). However, there is
scarce information regarding avocado oils’ IP by Rancimat, and there-
fore, more data is needed before attempting to rank the samples ac-
cording to this parameter. But, except for the ripe peeled avocados
dried in the oven or freeze-dried, all oils presented IP above 10 h, which
is generally considered a reasonable threshold for highly stable oils
such as those from macadamia (up to 37.4 h; 110 °C) (Souza,
Antoniassi, Freitas, & Bizzo, 2007), olive (6 to 11 h, 120 °C) and palm (7
to 12 h, 120 °C) (Metrohm Application Bulletin 240/02). Taken to-
gether, the results of oxidative stability and PV agree well with the
upper temperature limit that was achieved during oil extraction
(≤45 °C), indicating that the avocado oils processed and analyzed
herein are cold-pressed crude oils.
Similarly to IP, there is limited data of avocado oils’ TAC, and
specifically using the TEAC assay to the best of our knowledge there
have been no reports, and Krumreich et al. (2018) used DPPH assay for
pressed Breda avocado oil dried at 60 °C, but results are not directly
comparable. But by comparing the TEAC values obtained herein, it is
fair to say that pressed avocado oil, especially those samples obtained
by using the processes with best results, are among the oils with the
highest antioxidant activity values (Castelo-Branco, Santana, Di-Sarli, &
Torres, 2016; Pellegrini et al., 2003).
3.2. Avocado oil fatty acid composition was only marginally affected by
fruit ripening stage and peeling, and was not affected by pulp drying
technology
As expected, all avocado oils showed 18:1n-9 as the major fatty acid
(> 44%), followed by palmitic (16:0), palmitoleic (16:1n-7) and lino-
leic (18:2n-6) acids (Table 2). Only 18:2n-6 content varied slightly,
though significantly between oil samples, and showed the following
descending order, regardless the pulp drying technology used: ripe,
unpeeled > unripe, unpeeled > unripe, peeled > ripe, peeled.
Therefore, it might be stated that avocado pulp drying technologies
tested herein did not affect avocado oil fatty acid composition, and that
the fruit ripening stage and peeling affected it only marginally.
Oleic acid content was lower than the values established by Mexican
standards for avocado oil (56–74%) (NMX-F-052-SCFI-2008, 2008),
and some previous reports (Knothe, 2013; Haiyan, Bedgood, Bishop,
Prenzler, & Robards, 2007), although similar values to those herein
reported have been found elsewhere (Carvalho et al., 2015; Meyer &
Terry, 2008). Lower amounts of 18:1n-9 are related to premature har-
vest but also depend on environmental factors (Donetti & Terry, 2014;
Landahl, Meyer, & Terry, 2009). Because the avocados used in the
present study were harvested when horticulturally mature, as con-
firmed by dry matter content (> 21.5%), other factors than fruit ma-
turity stage at harvest, such as environmental ones (Carvalho et al.,
2015), might have led to reduced contents of oleic acid in the oil. Thus,
technical parameters adopted in specific countries’ legislations for
avocado oil quality and identity might not be universally adopted,
raising the need for local regulations and acceptable levels of technical
standards whenever feasible.
3.3. Avocado oil minor components: Unsaponifiable matter, tocopherol, β-
carotene and chlorophyll
Unsaponifiable matter varied from 0.89 g% in oils from ripe,

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Fig. 1. Oxidative stability (A) and total antioxidant capacity (B) of pitted unripe and ripe avocado (Persea americana Mill. cv. Hass) oils obtained from three pulp
drying technologies.
unpeeled freeze-dried avocado to 3.89 g% in unripe unpeeled oven-
dried avocado (Table 3). For each raw-material, oils obtained from
freeze-dried pulps showed the lowest contents of unsaponifiable matter,
independently of fruit ripening stage or peeling. Additionally, for the
oils from unripe and peeled avocados, drying under microwaves pro-
moted higher contents of unsaponifiables than in oven-drying, whereas
the opposite was observed for ripe unpeeled oils. It is not easy to ex-
plain these results by considering the current understanding of the ef-
fects of avocado ripening on the contents of unsaponifiables in the fruit
pulp and peel, and merits future investigations. Among the components
of the unsaponifiable fraction analyzed, chlorophyll was the major one,
followed by total tocopherols and β-carotene (Table 3).
α-Tocopherol was the major tocol in all avocado oil samples. β-
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Food Chemistry 286 (2019) 354–361
Tocopherol was present only in the oil from unripe oven-dried avocado,
while δ-tocopherol was present in most of the unpeeled samples, with
the exception of those dried under microwaves, suggesting that the peel
altered tocol isoforms profile. This was especially the case for δ-toco-
pherol, which is more effective in protecting vegetable oils against lipid
oxidation than α-tocopherol. Additionally, pressing unpeeled avocado
markedly increased α-tocopherol contents in the oils, except for the oil
from microwave dried ripe avocado. Concerning the drying technique
used, oils from microwave dried pulps showed highest contents of α-
tocopherol, whereas those from oven dried and freeze-dried pulps re-
sulted in oils with only slight differences between them.
β-Carotene was also higher in the unpeeled samples, but reduced
from unripe to ripe, and with all microwave dried samples showing the
I. Santana, et al. Food Chemistry 286 (2019) 354–361

Table 2 moderate temperatures, in the present work this was not the case for
Major fatty acids (> 1 g/100 g fatty acids) of pitted unripe and ripe avocado Hass avocado’s minor components during drying for oil extraction.
(Persea americana Mill. cv. Hass) oils obtained by pressing the fruit pulps dried Freeze-drying the Hass avocado pulp resulted in lower contents of to-
by varied methods. copherols, chlorophyll and total unsaponifiables in the oil (Table 2).
Fatty acid Pitted unripe avocado Pitted ripe avocado From our data it is not possible to determine what factors have pro-
composition moted the loss of these compounds, but we may speculate that enzymes
Peeled Unpeeled Peeled Unpeeled that remained active during freeze-drying might have caused these
Oven drying (60 °C)/Microwave/Freeze-drying
losses. Freshly freeze-dried avocado pulps had light-green color that
was preserved during freezing under vacuum packaging. However, after
Fatty acids and groups of fatty acids opening the package the pulp rapidly turned brown, as typically occurs
16:0 22.0 ± 0.77 21.2 ± 0.59 21.9 ± 0.75 21.8 ± 0.29 during enzymatic browning. Although polyphenoloxidase is the major
Total SFA 22.1 ± 1.12 21.7 ± 0.82 22.2 ± 0.99 22.2 ± 0.42
16:1n-7 12.5 ± 1.05 11.4 ± 0.71 11.7 ± 0.75 12.3 ± 0.69
enzyme causing these browning reactions, peroxidase and lipoxygenase
18:1n-9 45.2 ± 0.71 46.7 ± 0.62 46.6 ± 0.94 45.6 ± 1.18 activity might parallel and contribute to these transformations, pro-
Total MUFA 64.7 ± 2.60 65.2 ± 2.29 66.1 ± 1.17 65.4 ± 2.92 moting the oxidation of other compounds, such as lipids and toco-
18:2n-6 9.74 ± 0.29c 11.0 ± 0.33b 10.0 ± 0.26d 11.4 ± 0.35a pherols, which in turn may oxidize other fruit components. These en-
Total PUFA 11.4 ± 0.74 12.9 ± 0.63 12.0 ± 0.58 12.6 ± 0.96
zymatic activities have been shown in avocado (Jacobo-Velázquez,
Meaningful fatty acids ratios Hernández-Brenes, Cisneros-Zevallos, & Benavides, 2010; Vanini,
UFA/SFA 3.43 ± 0.06 3.60 ± 0.02 3.52 ± 0.15 3.52 ± 0.11 Kwiatkowski, & Clemente, 2010) and would possibly resist the mild
PUFA/SFA 0.51 ± 0.01 0.59 ± 0.01 0.54 ± 0.00 0.57 ± 0.03
18:1n-9/ 4.59 ± 0.15 4.21 ± 0.18 4.51 ± 0.14 4.00 ± 0.18
processing conditions during freeze-drying. In a previous work, freeze-
18:2n-6 dried guava powder contained less soluble phenolic compounds than
18:2n-6/ 19.36 ± 0.33 18.87 ± 0.50 19.05 ± 0.33 18.85 ± 0.51 the oven-dried powder (Nunes et al., 2016). These results indicate that
18:3n-3 preservation of fruits components during freeze-drying might be com-
pound- and fruit-specific, and this deserves proper direct investigations
*Values were presented as mean ± standard deviation of means from the three
in future studies.
drying processes used, and this samples’ grouping was adopted because fruit
pulp drying technology did not affect fatty acid composition. Values in the same
row with different superscript letters are significantly different (MANOVA, with
Fischer’s LSD post-hoc test; p ≤ 0.05). MUFA: monounsaturated fatty acids; 3.4. Identifying the main determinants of avocado oil quality
PUFA: polyunsaturated fatty acids; SFA: saturated fatty acids; UFA: unsaturated
fatty acids. To identify the main factors that influenced the quality of avocado
oils, we used a multi-factor categorical experimental design for data
highest contents. Therefore, oils from ripe avocados and microwave analysis and GLM to model avocado oil oxidative stability. The multi-
dried pulps will have technological benefits from the α-tocopherol an- factor design showed that unpeeling the avocados significantly
tioxidant activity, whereas oils from unripe avocados will benefit from (p < 0.05) improved all the response variables investigated herein (β-
β-carotene properties as natural pigment and singlet oxygen scavenger. carotene, α-tocopherol, chlorophyll-a, oxidative stability, and anti-
Total chlorophyll contents were higher in oils extracted from un- oxidant capacity) (Supplementary Fig. 2). Additionally, both oxidative
peeled avocados, especially when obtained from the unripe avocado stability and antioxidant capacity of avocado oil were improved by
fruits, because chlorophyll is hydrolyzed by chlorophyllases during ri- microwave-drying the fruit pulp, compared to oven- or freeze-drying
pening (Ashton et al., 2006). Besides that, the drying methods showing (Supplementary Fig. 2). A desirability function was only marginally
the best performances in preserving chlorophyll were microwave and significant (p < 0.1) for (adj. R2) chlorophyll-a (0.8845), α-tocopherol
oven drying, consistently with the higher stability of other minor (0.8924), oxidative stability (0.9035), and antioxidant capacity
components in avocado oil shown by these technologies. (0.9247).
Although freeze-drying is generally considered as a gold-standard GLM is a multivariate statistical method that combines multivariate
fruit drying method for vitamin preservation, because it occurs at regression and analysis of variance, therefore it allows considering si-
multaneously the contribution of categorical and continuous

Table 3
Minor components of pitted ripe and unripe avocado (Persea americana Mill. cv. Hass) oils: unsaponifiable matter, tocopherols, β-carotene, and chlorophyll a.
Hass avocado characteristics and pulp drying Unsaponifiable matter (g/ Tocopherols (mg/100 g) β-carotene (mg/ Chlorophyll a (mg/
technology used 100 g) 100 g) 100 g)
α β δ

Pitted unripe avocado

Peeled Oven drying (60 °C) 2.35 ± 0.15b,* 4.49 ± 0.49 d,* 2.41 ± 0.12 ND 0.30 ± 0.02 d,* 24.1 ± 0.83 e,*
Microwave-oven 3.83 ± 0.06 a,* 10.4 ± 1.00c,* ND ND 0.42 ± 0.01c,* 26.0 ± 1.00 d,*
Freeze-drying 2.24 ± 0.19c,* 6.17 ± 0.03 d,* ND ND 0.37 ± 0.02c,* 23.8 ± 1.13f,*
Unpeeled Oven drying (60 °C) 3.89 ± 0.15 a,* 17.1 ± 0.18b,* 5.32 ± 0.08 3.23 ± 0.23 0.67 ± 0.02b,* 57.1 ± 0.27 a,*
Microwave-oven 2.99 ± 0.17b,* 21.0 ± 0.12 a,* ND ND 0.65 ± 0.04b,* 54.0 ± 1.12b,*
Freeze-drying 1.51 ± 0.13 d,* 15.2 ± 1.02b,* ND 9.15 ± 0.12 0.83 ± 0.02 a,* 28.9 ± 2.22c,*

Pitted ripe avocado

Peeled Oven drying (60 °C) 3.44 ± 0.42 A 7.42 ± 0.96 D ND ND 0.24 ± 0.01 D 17.3 ± 0.57F
Microwave-oven 2.37 ± 0.41B 12.8 ± 2.25 A ND ND 0.33 ± 0.01C 28.1 ± 1.59 E
Freeze-drying 2.09 ± 0.06C 8.65 ± 0.16 D ND ND 0.28 ± 0.01C 40.1 ± 0.55C
Unpeeled Oven drying (60 °C) 1.41 ± 0.18B 11.7 ± 1.02B ND 6.96 ± 1.03 0.44 ± 0.02 A 32.1 ± 2.08 D
Microwave-oven 2.48 ± 0.15B 11.6 ± 3.13C ND ND 0.49 ± 0.14 A 44.3 ± 1.42 A
Freeze-drying 0.89 ± 0.01 D 12.0 ± 0.21B ND 4.40 ± 0.33 0.38 ± 0.03B 44.1 ± 2.22B

Results are expressed as mean ± standard deviation of triplicate processes. Different lowercase superscript letters in the same column indicate significant differences
between oils from unripe avocado. Different capital superscript letters in the same column indicate significant differences between oils from pitted ripe avocado.
*Significantly different from ripe avocado (peeled vs. peeled and unpeeled vs. unpeeled; for each drying process). MANOVA followed by Fischer’s LSD test (p ≤ 0.05).

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I. Santana, et al. Food Chemistry 286 (2019) 354–361

Table 4
Phenolic acids and quercetin composition (mg/100 g) in pressed avocado oil obtained from microwave-oven dried pulps, varying in ripening stage and peeling.
Phenolic compounds Contents of phenolic compounds (mg/100 g) in oil from avocado pulp dried in microwave-oven

Unripe Ripe

Peeled Unpeeled Peeled Unpeeled

Gallic acid ND 0.45 ± 0.02c 0.52 ± 0.01b 0.94 ± 0.03a

3,4 di-OH-p-phenylacetic acid ND ND 0.51 ± 0.03b 1.39 ± 0.05a
p-OH-benzoic acid ND 0.30 ± 0.06a ND 0.17 ± 0.06b
Vanillic acid ND 0.17 ± 0.01a ND 0.19 ± 0.01a
p-Coumaric acid 0.10 ± 0.00d 0.50 ± 0.01c 1.29 ± 0.03a 0.93 ± 0.14b
Ferullic acid ND 0.12 ± 0.00c 0.72 ± 0.06a 0.64 ± 0.16b
Quercetin 0.14 ± 0.01a 0.11 ± 0.00a 0.11 ± 0.01a 0.13 ± 0.03a
Total phenolics contents 0.24d 1.65c 3.15b 4.39a

Results are expressed as mean ± standard deviation of triplicate processes. Different lowercase superscript letters in the same row indicate significant differences
between oils. MANOVA followed by Fischer LSD test (p ≤ 0.05).

independent factors, such as ripening stages and contents of chemical
compounds, as determinants for a dependent variable. In the present
investigation we were interested in investigating the determinants of
avocado oil oxidative stability (IP, h). Continuous variables that cor-
related (Pearson’s) with IP were included in the GLM matrix as in-
dependent factors. In addition to these continuous factors, avocado ri-
pening stage, fruit peeling and drying process were also included as
independent categorical factors. Only the categorical variables fruit
peeling and drying process were retained as significant predictors of
avocado oils’ oxidative stability (p = 0.0312 and p = 0.0193, respec-
tively).
Taken together, these analyzes confirmed the general impression
captured from Fig. 1 and Table 3, which was that extracting avocado
oils from unpeeled and microwave dried pulps, regardless the avocado
ripening stage, resulted in the oils with the best quality, considering
oxidative stability and contents of minor components.
Heating avocado oils over 40 °C reduced its nutritional value and
oxidative stability (Santos, Alicieo, Pereira, Ramis-Ramos, & Mendonça,
2014). However, comparisons between microwave-drying and freeze-
drying showed that process duration was very important to the re-
sulting avocado oil quality. In the case of convection oven, the oxida-
tive atmosphere (high temperature and forced air circulation) was po-
tentially a decisive factor to decrease avocado oil quality. Concerning
the drying process and independently of other factors, avocado oil
stability tended to be improved by the fast process of microwave-
drying. Krumreich et al. (2018) showed that longer drying processing
decreased oil quality, even when occurring at lower temperatures, more
than high temperature drying for shorter periods. Although this seems
counterintuitive, there are two factors that might explain these results.
Because drying occurs before oil extraction, the food matrix compo-
nents serve as physical and chemical barrier, protecting the oil from
degradation that could be accelerated by heating.
Additionally, concerning the longer drying process tested (freeze-
drying), although chemical pathways of oil degradation would be in-
hibited at lower temperatures, these conditions would allow biochem-
ical pathways of oil degradation, for instance by endogenous enzyme
activities of polyphenoloxidase, peroxidase and/or lipoxygenase.
Another contributing factor to the extractability of minor avocado
components (from the unsaponifiable fraction) into the oil might have
been the avocado pulp microstructure resulting from each of the drying
technologies used, although this was not examined in the present work.
Therefore, concerning pulp processing before oil extraction, it seems
important to keep as low as possible the processing duration and its
temperature. We confirmed that microwave oven drying can be suc-
cessfully applied for drying avocados before pressing to obtain high
quality oils, as previously shown (Santana et al., 2015), but oven drying
at 60 °C should also be considered, preferably for the unpeeled fruit,
because it leads to increased contents of minor components, especially
in oils extracted from ripe avocados.
Although the GLM model confirmed the overall conclusion towards
the importance of the fruit peeling and pulp drying process used to
determine avocado oil composition, quality and stability, unexpectedly
tocopherols were not retained as independent factors in the model. In
addition, although the model was sound, and consistent with the gen-
eral conclusions revealed by the data (Fig. 1 and Table 3), the fraction
of IP variability explained by the independent factors was below 75%,
which would be the advisable threshold value. Possibly, phenolic an-
tioxidants in avocado pulp and peels, might have varied according to
the factors investigated (presence or absence of peel, ripening stage,
and pulp drying process), and this might have been a factor missing in
the GLM. It has been reported that avocado peels are rich in phenolic
compounds (Rodríguez-Carpena et al., 2011), and therefore we decided
to determine phenolic compounds profile of selected oils, as com-
plementary data. Avocado oils obtained by the seemingly best drying
technology used, which was using microwaves, were analyzed
(Table 4).
3.5. Phenolic compounds profile in avocado oils from the best pulp drying
process used: Microwave oven
Avocado ripening stage and peeling influenced the phenolic com-
position of the pressed oils (Table 4; Supplementary Fig. 1). Fruit ri-
pening increased the total phenolic content approximately 3- and 13-
fold in the oils extracted with or without the peel, respectively. Simi-
larly, unpeeling avocado increased approximately 7- and 1.4-fold the
total phenolic contents in oils from unripe and ripe avocados, respec-
tively. In the oils from unripe and ripe peeled avocados, two and five
phenolic compounds were identified, respectively. In contrast, six and
seven phenolic compounds were identified in the unripe and ripe un-
peeled samples, respectively. Taken together, these results point out to
the importance of processing unpeeled avocados to enhance the con-
tents of phenolic antioxidants and the stability of avocado oils. Inter-
estingly, although oils extracted from unpeeled avocados showed high
contents of p-coumaric, gallic and 3,4 di-OH-p-phenylacetic acids, oils
from ripe avocados presented high contents of these phenolic com-
pounds independently of peeling. Therefore, it seems that avocado ri-
pening induced accumulation of these phenolic compounds in the fruit.
4. Conclusion
Avocado peeling and pulp drying process were the main factors that
influenced the final quality of avocado oils, especially concerning oxi-
dative stability and antioxidant capacity, and content of minor com-
ponents in the oil. Oil extraction from microwave dried and unpeeled
Hass avocados resulted in high quality and stable avocado oil, with high
antioxidant activity, besides valuable nutritional properties because of

360
I. Santana, et al.
the amounts of α-tocopherol, β-carotene, and phenolic compounds,
although it reduced oil yield by roughly 20%, compared to oven-drying
unpeeled ripe avocado. Interestingly, by combining the technological
factors investigated herein (avocado ripening stage and peeling, and the
pulp drying process applied) it is possible to produce high quality
avocado oil, tailored for the desired application and depending on the
available resources and the potential market.
Declaration of interests
None.
Acknowledgements
The authors would like to thank the financial support granted by
Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio
de Janeiro (FAPERJ), Conselho Nacional de Desenvolvimento Científico
e Tecnológico (CNPq) and Coordenação de Aperfeiçoamento de Pessoal
de Nível Superior (CAPES, Finance code 001), Brazil. I.S. and L.O.S.
were recipients of CNPq scholarships, and V.O.Di-S. was a recipient of a
FAPERJ scholarship. A.G.T., L.M.C.C. and S.P.F. are recipients of CNPq
fellowships.
Appendix A. Supplementary data
Supplementary data to this article can be found online at https://
doi.org/10.1016/j.foodchem.2019.02.014.
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