foods

Article
Determining the Arrhenius Kinetics of Avocado Oil:
Oxidative Stability under Rancimat Test Conditions
Tugba Aktar 1, * and Eda Adal 2
1 Food Engineering Department, Faculty of Engineering, Alanya Alaaddin Keykubat University,
07450 Antalya, Turkey
2 Food Engineering Department, Faculty of Engineering, Gaziantep University, 27310 Gaziantep, Turkey
* Correspondence: [email protected]; Tel.: +90-242-510-61-20




Received: 18 June 2019; Accepted: 26 June 2019; Published: 30 June 2019


Abstract: Avocado is a highly potential functional fruit with significant health benefits which has
high demand for consumption with a preferable taste. The fruit is one of the oil sources that still
needs further examination on its probable kinetic behavior and oxidative stability as well as some
characteristic behavior to commercialize and increase the market demand as functional oil. Hence,
this study was motivated primarily for obtaining the Arrhenius kinetic information about avocado
oil to evaluate the oxidative stability and provide predictive information about the shelf life by using
the Rancimat method which is an accelerated shelf life test. Specifically, this research paper presents
the study of the physical, physicochemical, chemical, and oxidative stability tests with the shelf life
expectancy and kinetic property of avocado oil. According to the analyses, avocado oil has 210 days
of predicted shelf life at 25 ◦ C. This gives it a greater chance to be considered a good alternative to
other oils as well as its antioxidant and phenolic content. According to the findings presented in this
study, avocado oil has a very similar profile to olive oil and can be used as an alternative functional
oil source.

Keywords: avocado oil; accelerated shelf life testing; kinetic behavior; functional oil; rancimat;
oil technology

1. Introduction
Avocado (Persea americana Mill) is a fruit with high-calorie content, which is considered as a
functional food due to lipid fraction. It is one of the ancient fruits known have survived from the
Aztec times and has been used as vegetable butter since then [1]. Avocados grow at tropical and
subtropical regions and belong to the family of Lauraceae, which has about 150 species all with a similar
lipid profile to olive oil [2]. The main reason avocados have attracted the attention of researchers and
consumers is because of their functional compounds, which provide additional properties than just
oil [3]. The fruit flesh consists of 30% oil. Typical avocado oil has about 75% monounsaturated fat
that comprises oleic and palmitoleic acids, where 25% is saturated and polyunsaturated fatty acids [2].
The avocado oil has a characteristic flavor and a high smoke point that is above 250 ◦ C. The oil has a
green color and avocado flavor with mushroom flavor notes. This makes it worth promoting sensory
aspects in the future. More importantly, the main functional component is the α-tocopherol, which is
an antioxidant chemical that is usually around 79 to 190 mg/kg in avocado oil [1]. In the literature,
the avocado has been investigated as fruit and source of vegetable oil, and studies are mostly focusing
on the characterization of the oil, physicochemical properties, fatty acid composition, and comparative
studies with olive oil [4–6].

Foods 2019, 8, 236; doi:10.3390/foods8070236 www.mdpi.com/journal/foods

Foods 2019, 8, 236 2 of 13

To obtain the maximum functional components, the fruit needs to be carefully processed into the
oil with extraction methods such the soxhlet extraction, pressing, and ultrasonic extraction [7]. Not only
for avocado but also for all the vegetable–fruit oils, quality parameters are crucial for legislation as well
as consumers safety. While extracting the oil from the source, heat is generally applied for conventional
extraction methods. When applying heat, many desirable technological advantages occur, as well
as undesirable reactions. Oils are sensitive to heat and undergo quality loss caused by chemical
instability [7,8]. The most critical chemical change that represents quality loss and deterioration in
the oil is lipid oxidation. This well-studied field of oil occurs due to the chain reactions that result
in the formation of various oxidation products that are not suitable for consumption. The oxidation
procedure is the result of heat as well as environmental conditions, such as oxygen. Effects of the
lipid oxidation can be analyzed from the oxidized product; however, we can also express those effects
by mathematical relationships. Most often, the cause of the oxidation is the temperature, especially
while storing the oil/fat, and dependence of the temperature can be expressed best by the Arrhenius
model [9]. However, a researcher should always keep in mind that this mathematical model can only
be applied to a simple food system since the oxidative reactions are product dependent and often
more complex [10]. To apply the best fitting mathematical relationship to oil, it is necessary to obtain
information about the chemical structure, as well as manufacturing and processing conditions. Kinetic
data helps us to understand the oxidation reaction to calculate the best conditions to obtain better
quality with minimum deterioration. Additionally, using kinetic data, we can predict the oxidative
stability changes under various temperature applications, and storage and transporting periods [7].
Oil pressing/extraction technology is facing changes in the main manufacturing processes due
to the expected trend of cold pressing for better flavor, color, and higher functional nutrient content.
Apart from the mentioned advantages, cold pressing leads to a lower oxidation probability. For other
fruits and seeds such as olive oil, cold-pressing has become the conventional method.
Production volume of the avocado cold press oil is 2000 tonnes/year globally [11]. Despite the
increasing demand on the unsaturated functional oils, there is a limited number of studies on the
characterization of the avocado oil and the effect of production technologies on the quality. The main
reason for the cold pressing demand is due to easy and low-cost manufacturing as well as nutritional
protection. On the other hand, non-thermal applications were found to be beneficial for the physical
and chemical characteristics of the oil but disadvantaged in terms of extraction efficiency [12].
This paper presents innovative information about the mathematical relationships of the kinetics
of avocado oil as well as including some characteristic information about the oil source. The main
objective of this study was to develop mathematical models to describe the reaction rate as a function
of temperature for avocado oil to explore the shelf life and storage conditions, to provide insight
into the new generation functional oils and also to provide a step for future studies that are essential
for avocado oil. On the other hand, antioxidant properties and some characteristic properties were
determined, which might be either missing in the literature or need confirmation.

2. Materials and Methods

Avocado fruits (Fuerte and Hass variety) were collected from the local market of Alanya, a town
of Antalya city (Turkey) from a registered farmer. From each variety, 2 kg of fruit were purchased,
which is good enough for the analysis. All chemicals were purchased from Sigma-Aldrich Chemical
Company (St. Louis, MO, USA). Chemicals that used GC analysis were of chromatographic grade.
Other chemicals and solvents were in analytical grade.

2.1. Extraction of Avocado Oil

Avocado oil was extracted as applied previously in the literature [13]. Selected fruits were washed
and deseeded. The avocados were then mashed and transferred to a stainless steel vessel (malaxer).
An adequate amount of water was added, and the mixture was mixed for 60 min at 45 ◦ C. The mixture
Foods 2019, 8, 236 3 of 13

was then centrifuged at 5000 rpm for 20 min. (Eppendorf 5810R, Hamburg, Germany). Finally, the oil
layer was removed, placed into a dark brown bottle and stored at 4 ◦ C until analysis.

2.2. Kinetic Data Analysis

Oxidative stability of avocado oil was determined by the rancimat method, which is considered
as an accelerated determination of oxidation [14]. The rancimat method evaluates the stability
by measuring the oxidation induction time, with the Rancimat apparatus (Metrohm 743, Herisau,
Switzerland) which is capable of operating over a temperature range of 50 to 220 ◦ C. During the
analysis, rancimat vessels containing 3 g of oil samples were placed in an electric heating block.
Effluent air containing volatile organic acids from the oil sample was collected in a measuring vessel
containing distilled water (60 mL). The conductivity of water was measured automatically as oxidation
proceeded. Filtered, cleaned, and dried air was allowed to bubble through the hot oil at 20 L/h.
The oxidation induction time (OIT) of the oil samples were then automatically recorded at 100, 110,
120, 130, and 140 ◦ C. OIT is the time taken until there is a sharp increase in conductivity, which is
determined by the intersection of the baseline with the tangent to the conductivity curve [15]. Between
each run, the glassware was rigorously cleaned to avoid any contamination that would catalyze
the peroxidation. The tubes were cleaned with acetone after each run and then washed off with
washing liquid and hot water. The washed tubes were rinsed with distilled water and dried in the
oven. Measuring vessels, electrodes, and connecting tubes were cleaned several times with alcohol
and distilled water. For each parallel test, eight samples were accommodated in the equipment and
analyzed simultaneously. Kinetic parameters provide information about lipid oxidation. At moderate
temperatures, due to the high concentration of oxygen dissolved in the oil, the lipid is prone to the
autoxidation regardless of the oxygen pressure and oxidation rate [16–18]. In this study, the effect of
temperature on the rate of lipid oxidation was illustrated by means of the Arrhenius equation:

Ea
ln(k) = ln A −
RT
where k is the reaction rate constant or reciprocal oxidation induction time (OIT), A is the pre-exponential
factor or frequency factor, Ea is the activation energy (kJ mol−1 ), R is the molar gas constant
(8.314510 J K−1 mol−1 ), and T is the absolute temperature (K).
The OIT of the oil determined activation energy, and frequency factors were calculated from the
slopes and intercepts of the lines generated by regressing ln (k) vs. 1/T by use of the least squares
linear regression, respectively. By those values and calculations, the shelf life prediction of avocado
oil at 25 ◦ C was calculated by linear regression of log OIT versus T in K, which adapted from the
literature [7,19].

2.3. Determination of Basic Physical and Physicochemical Properties of Avocado Oil

2.3.1. Density of Avocado Oil

The density of the avocado oil was measured using pychnometric methods according to the
International Organization of Standardizations (“Methods for determining the density of non-cellular
plastics, Part 1: Immersion method, liquid pyknometer” (ISO 1183-1:2012)). Measurements were taken
of the volume of the oil displaced by a known mass of the samples at 25 ◦ C. According to that reading,
the density value of the oil samples was calculated by taking the ratio of mass to volume.

2.3.2. Refractive Index

Refractive indices (optical measurements) of the oil samples were measured by using an Abbe
Refractometer (ATAGO NAR 1T-LIQUID, Tokyo, Japan). A halogen lamp was used for the high bright
white light source.
Foods 2019, 8, 236 4 of 13

2.3.3. Free Fatty Acid

The free fatty acid of the avocado oil was measured as one of the physicochemical properties, and
the method of measurement was used for sample titration [20]. For the measurement, diethyl ether
was used, and the indicator was phenolphthalein. Neutralization was done by NaOH, and the total
volume of the NaOH was measured for the percentage of free fatty acid (oleic acid) content as follows:

V × 28.2 × N
% Free fatty acids =
m
where V = volume of 0.1 N NaOH used for the sample, N = normality of NaOH used for titration, and
m = mass of the sample in grams

2.4. Determining the Chemical Properties of Avocado Oil

2.4.1. Determination of Fatty Acid Composition of Avocado Oil

The fatty acid composition of avocado oil was analyzed using Agilent GC7890A system (Agilent
Technologies, Wilmington, DE, USA) gas chromatography equipped with a flame ionization detector
and HP-88 capillary column (88% Cianopropylaryl; 100 m × 0.250 mm ID × 0.20 µm film thickness).
The flow rate of the carrier Helium gas was set to 1 mL per minute, where detector temperature
and injector temperature was 250 ◦ C, respectively. The oven temperature was set to start with
175 ◦ C for 10 minutes and increase 5 ◦ C per minute up to 230 ◦ C. Injection volume was 1 µL. These
analytical conditions were used for fatty acid methyl esters (FAME) extraction with n-heptane after
cold methylation with 2N KOH in methanol. Specifically, fatty acids were converted into FAME by
0.1 g of the oil sample being mixed with 2 mL n-heptane, which followed by addition of 0.2 mL (2N)
methanolic KOH addition. After the phase separation GC analysis was initiated.
The specific standard for each fatty acid type was used for the identification of the fatty acids by
comparing the retention times for standards and samples. The area was expressed as percentages of
the areas of the total fatty acids [21].

2.4.2. Determining the Total Phenolic Content of Avocado Oil

The total phenolic contents of the avocado oil were assessed with the Folin–Ciocalteu method [22].
Total phenolic content was calculated using a standard curve of gallic acid and reported as milligram
gallic acid equivalent milligram per grams of oil (mg of GAE g−1 ). Briefly, the reaction mixture
contained 0.1 mL of oil extracts and 0.5 mL of the Folin–Ciocalteu reagent freshly prepared in the
laboratory and 1.5 mL of 20% sodium carbonate and was completed to 10 mL volume with pure water.
After 2 h of reaction at ambient temperature, the absorbance at 765 nm was measured, and the obtained
value was used for the calculation of the phenolic contents in oil using gallic acid as a standard.

2.4.3. Determination of 2,2-Diphenyl-1-picrylhydrazyl (DPPH) Radical Scavenging Activity

DPPH scavenging capacity of the avocado oil was determined according to the previously reported
method applied previously [23]. During the experiments, a stable 2,2-diphenyl-1-picrylhydrazyl
(DPPH) radical was used, which was purchased from Sigma-Aldrich (St. Louis, MO, USA). Freshly
made 100 mM DPPH-MeOH solution was mixed into the oil extract at different concentration of mg oil
equivalents/mL to start the radical–antioxidant reaction. The absorbance at 517 nm was measured
against a blank of pure methanol after 30 min of the reaction.

2.4.4. Component Analysis of Avocado Oil

Fourier transforms infrared radiation (FTIR) is one of the trending analytical tool for edible
oils [24]. FTIR is a quantitative tool that carries out component analyses. In this study, the spectral
data were collected on a Perkin–Elmer Spectrum 100 spectrophotometer (Spectrum Two) (Shelton, CT,
Foods 2019, 8, 236 5 of 13

USA) fitted with a universal attenuated total reflectance (UTAR) sampling device. A drop of each oil
sample was placed directly onto the Universal diamond ATR crystal, and all spectra were measured
at room temperature against a background spectrum of air in the wavenumber range from 4000 to
600 cm−1 . Between each sampling procedure, the cell was thoroughly cleaned and dried by aspirating
hexane through the cell using a vacuum and its cleanliness verified spectrally. Spectra were examined
using the instrument's software Spectrum 10 STD (Perkin-Elmer, Shelton, CT, USA) with peak heights
and areas computed from the raw spectra.

2.5. Statistical Analyses

All experiments and measurements were made in triplicate for each replicate. The coefficient
of variation for quadruplicate determinations was typically less than 5%. All data were subjected to
ANOVA and regression analyses using the SPSS statistical package (Version 22, SPSS Inc., Chicago, IL,
USA). Duncan’s multiple range tests were applied to measure specific differences between pairs of
means. General statistical values, such as mean value, correlation of determinations (R2 ), p-values, and
standard deviation, were calculated. Additionally, the analysis made on the avocado oil were grouped
as Kinetic data analysis (Rancimat test), physical and physicochemical analyses (density, refractive
index, free fatty acid content), chemical analyses (accelerated oxidation, fatty acid composition, total
phenolic content, DPPH radical scavenging), and component analyses (FTIR). Each experiment was
done in triplicate, and the mean data were calculated

3. Results and Discussion

The main aim of this study was to characterize and investigate the kinetic analysis of avocado
oil. The rancimat method was used as a reflection of the expected shelf life determination as well
as determining the kinetic behavior of the avocado oil. Rancimat measurements for the oxidation
induction period (OIT, time in hours) were done isothermally at five different temperatures (100 ◦ C,
110 ◦ C, 120 ◦ C, 130 ◦ C, and 140 ◦ C) and those values were selected according to the study done
previously [25]. OIT of the avocado oil can be seen in Table 1. Results illustrate that OIT values are
decreasing by the increasing temperature. This result has been previously validated [26] According
to the experiments and calculation, a linear regression model was obtained, which was used for
calculating the expected shelf life of the avocado oil.

Table 1. Oxidation induction time (OIT) values and reaction rate constants (kOIT = 1/OIT) at five
different temperatures T for avocado oil.

T (◦ C) OIT (h) * kOIT (×10−3 h−1 )

100 15.02 ± 2.03 a 66.6
110 7.06 ± 3.15 b 141.6
120 2.96 ± 0.25 c 337.8
130 1.44 ± 0.03 d 694.4
140 0.68 ± 0.15 e 1470.6
* Values printed in one column, with the different letters (a–e) in superscript are statistically different at the p < 0.05
level, 95% confidence limit, according to Duncan’s Multiple Range Test. OIT, oxidation induction time.

As mentioned earlier, the k value (reaction rate constant) was determined by the reciprocal OIT
for each temperature and represents lipid oxidation of avocado oil, which can be seen for different
temperature operations in Table 1. By using the k value rate of lipid oxidation as a function of
temperature, a direct relationship can be observed.
As shown in Figure 1, ln (k) vs. 1/T was obtained from the measurements. Those values were
used to obtain activation energy and pre-exponential factor. The slope was used to calculate Ea while
intercept used for A (frequency factor) as explained in (R2 > 0.99). More specifically, k and T values
have a semi-logarithmic relationship with a linear dependency with good correlation of determination
Foods 2019, 8, 236 6 of 13

(R2 > 0.99). Duncan’s multiple range tests were calculated for OIT assay and statistically significant
(p < 0.05) difference was found at all temperatures. The results indicate that the reaction rate constant
increase with increasing temperature, which means that the oil oxidation is faster at higher temperatures
as authors expected. Noteworthy, the kinetic rate constant prediction, especially at lower temperatures,
Foods 2019, 8, x FOR PEER REVIEW 6 of 13
has some limitations which cause some uncertainties and errors. Those limitations are linked with the
test. Specifically, the oil follows different pathways of lipid oxidation at lower and higher temperatures
at lower and higher temperatures depending on the metal ions and antioxidants activity. On the other
depending on the metal ions and antioxidants activity. On the other hand, the degree of oxygen solubility
hand, the degree of oxygen solubility is different in varying temperatures, where literature reporting
is different in varying temperatures, where literature reporting to see an increase of 25% for each 10 ◦ C.
to see an increase of 25% for each 10 °C.

0.5 y = -11985x + 29.389

R² = 0.9992

0
0.0024 0.00245 0.0025 0.00255 0.0026 0.00265 0.0027

-0.5
lnkOIT

-1

-1.5

-2

-2.5

-3
1/T

Figure 1.
Figure 1. ln
ln(k)
(k)vs.
vs.1/T
1/Twhich
which illustrates
illustrates thethe activation
activation energy
energy andand pre-exponential
pre-exponential factorfactor
wherewhere the
the slope
slope is used for the calculation of E a while the intercept is used for the calculation of A (frequency
is used for the calculation of Ea while the intercept is used for the calculation of A (frequency factor).
factor).
Calculated Arrhenius activation energies (99.6 ± 2 kJ·mol−1 ) and pre-exponential factor
1012 h−1 ) are
(5.8 ×Calculated Arrhenius
listed inactivation
Table 2. energies
According (99.6to±the
2 kJ·mol −1) and pre-exponential factor (5.8 × 1012
literature, the stability of avocado oil was
h −1) are listed in Table 2. According to the literature, the stability of avocado oil was similar to that of
similar to that of olive oil because of similar lipid profile and stability results, which means our
olive oil
results arebecause of similar
in accordance with lipid profile
literature and stability
findings results,
[11]. Another which means
supporting ourtheresults
data for are of
similarity in
accordance
avocado oil with literature
and olive oil wasfindings [11]. Another
investigated supporting
on different kinds ofdata olivefor the
oils similarity
under of avocado
rancimat oil
conditions
and illustrated
olive oil that
was the investigated on different
activation energies for thekinds of olive
oxidation of theoils under
olive rancimat
oils were between conditions −1
and
97.7 kJ·mol
illustrated
and 101.9 kJ·mol −1
that the, activation
which is veryenergies
similar fortothe
theoxidation
findings of the olive
in this studyoils were
[25]. betweeninvestigation
A previous 97.7 kJ·mol−1
and 101.9
showed kJ·mol
that −1 , whichofisdifferent
the kinetics very similar
type to of the findings
vegetable in as
oils, this study
well as E[25]. A previous
a values investigation
of selected vegetable
showed that the kinetics of different −1
type of vegetable −1
oils, as well as
oils, was found to be between 79 mol and 104 kJ·mol which have a different fatty acid profile E a values of selected vegetable
[7].
oils, was found
Alternative oils,to
suchbe between 79 mol
as vegetable −1 and olive
oils and 104 kJ·mol −1
oil, werewhich
found have a different
to have similarfatty acid profile
activation [7].
energies.
Alternative
The oils whichoils,have
suchlower
as vegetable
activationoilsenergy
and olive oil, were to
are expected found to have
require similar
a higher activationto
temperature energies.
induce
aThe oils which
certain change have
in thelower
rateactivation energy
of oxidation. are expected
Therefore accordingto require
to ourafindings,
higher temperature
avocado oiltoisinduce
stable
a certain change
compared to otherinoilthe rate ofasoxidation.
sources well as oliveTherefore
oil. according to our findings, avocado oil is stable
compared to other oil sources as well as olive oil.
Foods 2019, 8, x FOR PEER REVIEW 7 of 13
Foods 2019, 8, 236 7 of 13
Table 2. Estimated Arrhenius parameters for avocado oil oxidation on isothermal conditions (lnkOIT
vs. 1/T).
Table 2. Estimated Arrhenius parameters for avocado oil oxidation on isothermal conditions (lnkOIT vs. 1/T).
Parameters Avocado Oil
Parameters
b (intercept) Avocado
29.4 ± Oil
0.83
ba(intercept)
(slope) 29.4 ± 0.83
−11985 ± 311
a (slope)
R2 −119850.99
± 311
R2 0.99
Ea (kJ/mol) 99.6 ± 2
Ea (kJ/mol) 99.6 ± 2
AOIT
A OIT (×10
(×101212h−1h)−1 ) 5.85.8
Relationshipbetween
Relationship between lnk
lnk vs.1/T
vs.
OITOIT 1/Tgives
givesaalinear
linearrelationship
relationshipasasy=y =−11985x
−11985x+ +29.389
29.389where
where coefficient
coefficient of
2
1/T is represented
of 1/T with letter
is represented withaletter
and the additive
a and the constant
additiveofconstant
the relation is shown
of the withisletter
relation shown with letter Rb.
b. Meanwhile;
represents regression coefficient, E a represents calculated Arrhenius activation energy, and A OIT represents the
Meanwhile; R2factor.
pre-exponential represents regression coefficient, Ea represents calculated Arrhenius activation energy,
and AOIT represents the pre-exponential factor.

Kinetic properties calculations were based on the measurements done by the Rancimat method to
Kinetic properties calculations were based on the measurements done by the Rancimat method
investigate the expected shelf life of avocado oil. A linear relationship between the natural logarithm
to investigate the expected shelf life of avocado oil. A linear relationship between the natural
of the OIT and the temperature was used to calculate the predicted shelf life of avocado oil at 25 ◦ C,
logarithm of the OIT and the temperature was used to calculate the predicted shelf life of avocado oil
as shown in Figure 2.
at 25 °C, as shown in Figure 2.

1.4

1.2 y = -0.0338x + 4.552

R² = 0.9991
1

0.8
log (OIT)

0.6

0.4

0.2

0
80 100 120 140 160
-0.2

-0.4
T (°C)

−
Figure 2. The linear relationship between the natural logarithm of the oxidation induction time (OIT)
temperature.
and the temperature.

It can be observed that avocado oil obeys the Arrhenius relationship over the temperature range
of 100 ◦ C to 140 ◦ C (R22 > 0.99). The data calculated from this relationship are shown in Table
100 °C to 140 °C (R > 0.99). The data calculated from this relationship are shown in Table 3.
−2 ◦ C−1
According
According to to those
thosecalculations,
calculations,the
the temperature
temperature coefficient
coefficient of avocado
of avocado −3.4 ×−3.4
oil was
oil was 10−2 ×
°C10
−1 (logOIT

= a(T) + =b).
(logOIT a(T) + b). behavior
Kinetic Kinetic behavior of the avocado
of the avocado oil hasoil has
not not studied
been been studied previously
previously according
according to
to our
our literature knowledge. Therefore, the kinetic behavior of the avocado oil is still missing,
literature knowledge. Therefore, the kinetic behavior of the avocado oil is still missing, and this study and this
study will trigger
will trigger the further
the further research
research ideas ofideas of the avocado
the avocado oil. Previously
oil. Previously other oilother oil sources,
sources, such as
such as soybean
−2 ◦ −1
soybean
oil, wereoil, wereand
tested, tested, and the temperature
the temperature coefficient
coefficient wasas
was found −3.12as× −3.12
found 10−2 °C×−110 C [27]. study
[27]. Another Anotheron
study on the vegetable oil temperature coefficient was calculated as −2.78 × 10 −2 and −3.15–2× 10 −2 ◦ C−1
the vegetable oil temperature coefficient was calculated as −2.78 × 10 and −3.15 × 10 °C (mean
−2 –1
Foods 2019, 8, 236 8 of 13

(mean value −3.01 × 10−2 ◦ C−1 ) [28]. Even though those findings are not necessarily comparable with
the avocado oil, they seem to be fitting with our findings and calculations as a general profile.

Table 3. Calculated results of the linear relationship between the natural logarithm of the OIT assessed
by the Rancimat test and the temperature for the treatment combinations in.

Parameters Avocado Oil

a (slope) −0.034 ± 0.4
b (intercept) 13.8 ± 1
R2 0.99
OIT25 ◦ C (day) 211 ± 10

Noteworthy, rancimat tests of the ambient storage were declared to lead either over prediction
or under prediction of the actual shelf life depending on the type of oil [27]. Additionally, the lipid
oxidation at low and high temperatures may go through different reaction pathways, depending on
the reactivity of metal ions and antioxidants at different temperatures [19]. Therefore, the rancimat
method can be claimed as a remarkable kind of a test as a fast shelf life prediction, but it would be
suitable to compare fast and regular shelf life tests in future researches.
According to our calculations, avocado oil has 210 days of shelf life according to the model.
The usual shelf life of the vegetable oils was presented to be between 12 and 15 months at 25 ◦ C when
good manufacturing and good storing conditions are followed [26]. Additionally, it is also necessary to
highlight that once the oxygen interaction initiates, shelf life starts to decrease. However, the addition
of the alternative antioxidants, minor protective/antioxidative components may increase the shelf life,
which is still in need of further investigation to obtain the best component and best practice.
Results of the physical and physicochemical properties, specifically density, refractive index,
and free fatty acid measurements are shown in Table 4. The density of the avocado oil was 0.91 g/mL
(±0.0001). Meanwhile, the refractive index value was 1.4680 (±0.0002). For the density and refractive
index values, literature has reported similar results [6,29]. Those properties are similar to those of olive
oil. In the literature, virgin olive oil was presented to have 0.8639 g/mL density while the refractive
index was between 1.480 and 1.465 according to varying wavelength values of experiments [30].
The free fatty acid value was found to be 1.065% (±0.040) oleic acid. An earlier study illustrated
that free fatty acid value of the virgin olive oil and extra virgin olive oil was measured as 0.26% and
0.36%, respectively [31]. It can be seen that the acid value of the avocado oil is higher than the olive oil.
Those values imply that avocado oil has similar characteristic properties to olive oil, which has also
been supported by previous researches [11,32,33]. Noteworthy, oil processing technique is critical and
varies the free fatty acid values [12]. Therefore, the effect of different oil pressing techniques on the free
fatty acid value can be a further research question.

Table 4. Physical and physicochemical properties of the avocado oil.

Measured Property Value

Density (g/mL) 0.91 ± 0.001
Refractive index 1.4680 ± 0.0002
Free fatty acid (% oleic acid) 1.065 ± 0.040

Results of fatty acid compositions of avocado oil were illustrated in Table 5. The profile shows
that obtained fatty acids are mostly unsaturated except for the palmitic acid. Specifically, avocado oil
contains (in descending order) oleic acid (18:1 ω-9), palmitic acid (16:0), linoleic acid (18:3 ω-3), and
palmitoleic acid (16:1). The high content of unsaturated fatty acid gives a unique property to avocado
oil with delivering functionality to the product. Obtained fatty acid profile agrees with previous
reports [34–37]. However, the differences in the cultivars of the fruit might change the concentration of
the fatty acids presented, but it is likely to have a similar order of the fatty acid concentration [38].
Foods 2019, 8, 236 9 of 13

Table 5. Fatty acid profile of avocado oil.

Fatty Acids %
C14:0 (Myristic acid) Not detected
C16:0 (Palmitic acid) 18.29 ± 0.2
C16:1 (Palmitoleic acid) 8.36 ± 0.05
C18:0 (Stearic acid) 0.69 ± 0.01
C18:1 (Oleic acid) 54.33 ± 0.4
C18:2n-6 (Linoleic acid) 11.54 ± 0.2
C18:3n-3 (Linolenic acid) 0.78 ± 0.02

Due to the potential impact on human health and critical importance for the daily diet, the total
phenolic content of the avocado oil was measured. The daily intake of the total phenolic content
was estimated to be sourced by the most common 34 fruits and vegetables where avocado was one
of them [39]. Therefore, in this perspective, avocado is one of our phenolic source where it worth
doing examination about its total phenolic content and the methods to increase the usage of it. In our
study, the total phenolic content was found as 25.73 (±2.1) mg GAE per g of oil (Table 6). The previous
study done by [39] presented that avocado fruit contains 33.62 mg GAE per g of oil. With a similar
approach [40] different cultivars of avocados found that they between 6 and 49 mg GAE per g of oil.
The total phenolic content can get affected by the maturity state of the avocado, and the more mature
fruit was found to have higher phenolic content [41]. On the other hand, olive oil has shown similar
results to those of avocado oil, and this finding supports the idea of avocado oil being an alternative
oil [42,43].

Table 6. Total phenolic content and antioxidant activity of avocado oil.

Measured Property Value

Total phenolic content (mg GAE/g oil) 25.73 ± 2.1
DPPH (IC50 , mg/mL) 32.4 ± 1.3

Total antioxidant capacity varies from one kind of fruit/seed to another. Additionally, the total
antioxidant capacity is easily affected by the processing methods as well as the different parts of the
fruits [43]. It should be highlighted that for functional oil (which is likely to contain antioxidants)
resistance to oxidation stress is critical. There are several methods for measuring the total antioxidant
capacity, and these methods generate different radical/target groups [44,45]. In this study, the DPPH
method was used to evaluate the antioxidant capacity, which is also a very common method for the
assessment of antioxidants [46,47]. According to the analysis done, the antioxidant capacity of avocado
oil was found to be 32.4 mg/mL (±1.3) as IC50 value, required to lower the initial DPPH concentration
by 50% (Table 6). A previous research illustrated the antioxidant capacities of the extra virgin olive
oil, olive oil, corn oil, sunflower oil, and soybean oil as 15 mg/mL, 22 mg/mL, 52 mg/mL, 48 mg/mL,
45 mg/mL, respectively [47]. The results showed that the radical scavenging capacity of avocado oil is
higher than those of corn, sunflower, and soybean oil.
FTIR results are shown in Figure 3. The spectral regions were chosen for developing the
regressions with including the fingerprint regions were selected according to the previous researcher's
observations [48]. The spectra in the FTIR region have well-resolved bands that can be assigned to
presented functional groups of the avocado oil. The spectra are dominated by some peaks at 3006,
2922, 2853, 2162, 1991, 1744, 1464, 1418, 1378, 1237, 1160, 1118, 1096, 874, and 722 cm−1 . According to
that, the absorbance results for the region of 3006 to 2853 cm−1 are due to the bands of CH2 stretching
vibrations, asymmetric and symmetric, respectively. The high value of the frequency of this band
indicates its richness in polyunsaturated acyl groups. According to the literature, only linseed oil
shows a band frequency as high as this; for example, olive oil shows values near 3005.4, rapeseed oil
near 3007.5, and corn oil near 3008.8 cm−1 [49]. So, the frequency value of these bands in avocado oil
Foods 2019, 8, 236 10 of 13

is similar to those of olive oil. The major peak at 1744 cm−1 arises from C=O stretching vibrations
of aldehydes and ketones where the peaks at 1464, 1418, and 1378 cm−1 arise from CH2 and CH3
scissoring vibration of ethers. Meanwhile, 1237, 1160, 1118, and 1096 cm−1 peaks are associated with
C–O stretching vibration. The last peak at 722 cm−1 associates with the CH2 rocking mode. Those
Foods 2019, 8, x FOR PEER REVIEW 10 of 13
observations are supported by the results of the other researchers performed with oils [48,50].

120.8

115
110
105
100 1417.95

95 2162.13 1990.84
3006.20 873.96
90
85 1377.55

80 1463.76
%T 1236.77 1095.56 722.26
75
1118.08
70
65
2853.37
60 1159.87
55
50 2922.30

45 1743.88
40
36.7
4000.0 3000 2000 1500 1000 600.0
cm−1

Figure 3. FTIR spectra of avocado oil in the region of 4000 to 600 cm−1 .
Figure 3. FTIR spectra of avocado oil in the region of 4000 to 600 cm−1.
4. Conclusions
4. Conclusions
According to the analyses and literature comparison of the characteristic values of avocado oil,
it canAccording
be mentioned to the analyses
that avocado andoilliterature
has similar comparison of the
characteristic characteristic
behavior values
to the olive oilofwhich
avocado oil,
makes
it acan be potent
high mentioned that avocado
functional oil. Onoil thehas similar
other characteristic
hand, the shelf lifebehavior to the olive
was determined to oil
be awhich
little makes
above
it a high
200 days.potent functional
However, oil. On
200 days theaother
is still hand,
feasible the shelf
duration forlife was determined
storage to be a littleoil.
time for a functional above 200
Kinetic
days. However,
behavior 200 daysby
was determined is the
stilltemperature
a feasible duration forand
coefficient, storage
it wastime
found for as
a −3.4 × 10−2 oil.
functional ◦ C−1
Kinetic
. This
behavior
data wasbeen
has not determined by theand
studied earlier temperature
accordingcoefficient, and itfor
to other studies was
thefound as −3.4 coefficient
temperature × 10−2 °C−1value
. This
data found
was has nottobeen studied
be −3.12 × 10 −2 ◦
earlierCand−1 according
, −2.78 −2
× 10 to other studies
and −3.15 −2 ◦ −1
for theCtemperature
× 10 for soybean coefficient
oil and value
other
was foundoils,
vegetable to berelatively
−3.12 × 10[19,27,28].
−2 °C−1, –2.78 Those× 10findings
–2 and –3.15
are×similar
10–2 °C–1 tofor soybean
those oil andoil,
of avocado other
which vegetable
could
oils, relatively
mean [19,27,28].
that avocado oil canThose
be an findings
alternative areoil
similar
source.to those of avocado oil, which could mean that
avocado oil can be an
In conclusion, alternative
cold oil source.
pressed avocado oil which has already been previously referred as a healthy
In conclusion,
alternative cold pressed
oil and according to theavocado
illustratedoil which
resultshas
andalready been previously
calculations; it can be anreferred as anot
alternative healthy
only
alternative
to oil and
other edible according
oils but to theoil.
also to olive illustrated results and calculations; it can be an alternative not
only to other edible oils but also to olive oil.
Author Contributions: Conceptualization, T.A. and E.A.; Data curation, T.A. and E.A.; Formal analysis, T.A.
and E.A.; Investigation, T.A. and E.A.; Methodology, T.A. and E.A.; Project administration, T.A. and E.A.;
Resources, T.A. and E.A.; Software, T.A. and E.A.; Supervision, T.A. and E.A.; Validation, T.A. and E.A.;
Visualization, T.A. and E.A.; Writing—original draft, T.A. and E.A.; Writing—review and editing, T.A. and E.A.

Funding: This research received no external funding

Foods 2019, 8, 236 11 of 13

Author Contributions: Conceptualization, T.A. and E.A.; Data curation, T.A. and E.A.; Formal analysis, T.A. and
E.A.; Investigation, T.A. and E.A.; Methodology, T.A. and E.A.; Project administration, T.A. and E.A.; Resources,
T.A. and E.A.; Software, T.A. and E.A.; Supervision, T.A. and E.A.; Validation, T.A. and E.A.; Visualization, T.A. and
E.A.; Writing—original draft, T.A. and E.A.; Writing—review and editing, T.A. and E.A.
Funding: This research received no external funding.
Acknowledgments: Authors declare that there are no present or potential conflicts of interest among them and
other people or organizations that could inappropriately bias their work. Authors would also like to thank Asim
Samli Agricultural Products Import, Export & Trading Inc. for their help and support during rancimat experiments.
Conflicts of Interest: The authors declare no conflict of interest.

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