Journal of Molecular Structure 482–483 (1999) 551–556

Determination of unsaturation grade and trans isomers generated

during thermal oxidation of edible oils and fats by FTIR
M.C.M. Moya Moreno a,*, D. Mendoza Olivares b, F.J. Amézquita López b,
J.V. Gimeno Adelantado a, F. Bosch Reig a
a
Department of Analytical Chemistry, Faculty of Chemistry, University of Valencia, C/Dr. Moliner 50, 46100 Burjassot, Valencia, Spain
b
Department of Chemistry, Instrumental Laboratory, Faculty of Chemistry, University of Guanajuato, Noria Alta, 36050 Guanajuato, Mexico
Received 24 August 1998; received in revised form 30 November 1998; accepted 30 November 1998

Abstract
The oxidative deterioration of culinary oils and fats during episodes of heating associated with normal usage (80 C–300 C,
20–40 min) was monitored by FTIR spectroscopy. The thermal oxidation of polyunsaturated fatty acids during heating was
studied by the determination of unsaturation percentage and trans isomers at various temperatures and heating times. Oils
frequently used in food frying such as olive oil, sunflower oil, corn oil and seeds oil (sunflower, safflower and canola seed), and
lard were studied. The Absorbance Correction Method is proposed to correct the spectral interference and allows the analytic
use of signal which would not be initially valid for quantitative analysis. The results show that there is a decrease in unsaturation
and an increase in trans isomers starting at 150 C and becomes more pronounced at temperatures around 250 C. This variation
in unsaturation grade and conformation provides evidence of the transformation of essential polyunsaturated fatty acids and
subsequent decrease in the oils’ nutritional value. 1999 Elsevier Science B.V. All rights reserved.
Keywords: Edible oils; Polyunsaturated fatty acids; Trans isomers; FTIR

1. Introduction body must be present in the diet to avoid symptoms

Oils and fats are an important part of the human diet
and more than 90% of global production is used as
food or as ingredients in food products. Oils and fats
in the diet are a rich source of energy, they contain
certain fatty acids, which are nutritionally indispen-
sable and their functional and textural characteristics
add to the flavour and acceptability of many natural
and processed foods. The polyunsaturated fatty acids
(PUFA) which cannot be synthesised by the human
* Corresponding author. Tel.: 34 6 386 4533; fax: 34 6 386
4436.
E-mail address: montse.moya@uv.es (M.C.M. Moya Moreno)
0022-2860/99/$ - see front matter 1999 Elsevier Science B.V. All rights reserved.
PII: S0 022 - 2 86 0( 98 )0 0 937 - 5
caused by shortage. They can be considered as
vitamin factors and linoleic, and linolenic acids
must be considered specifically as they are polyunsa-
turated fatty acids which are necessary for the normal
development and functioning of human tissues and
are known as essential fatty acids [1]. Oils and fats
begin to decompose from the moment they are
isolated from their natural environment when changes
occur causing a disagreeable taste and smell. Atmo-
spheric oxidation is the most important cause of dete-
rioration in fats. This oxidative rancidity is
accelerated by exposure to heat, light, humidity and
the presence of trace transition metals [2]. The intense
frying of oil causes an oxidizing thermal degradation
552 M.C.M. Moya Moreno et al. / Journal of Molecular Structure 482–483 (1999) 551–556

with the formation of decomposition products [3–6], standard by relating it with the absorbance at another
and change in physical properties [7]. Spectroscopic auxiliary frequency 2 at which only the standard
methods can be employed to evaluate deterioration in absorbs and not the analyte, i.e., knowing the absor-
oils subjected to intense heat [8]. bance arising exclusively from the standard at another
Fourier Transform Infrared (FTIR) Spectroscopy frequency 2, it is possible to predict the absorbance at
provides a quick and accurate way of evaluating the frequency of interest 1, as the relation between
thermal degradation in these lipids. This article the both is constant.
reports a methodology to evaluate the variation in Therefore, the Standard Correction Factor
composition in edible oils (sunflower oil, corn oil, F c 1 2 is defined at the frequency 1 using 2 as
olive oil and seeds oil) and lard when it is subjected auxiliary frequency from the infrared spectrum of the
to intense heat (80 C–300 C, 20–40 min). The unsa- pure standard compound,
turation percentage and trans isomers percentage was
Fc 1 2 AP 0 1 AP 0 2 2
determined by FTIR at various temperatures and
heating times, using n-valeronitrile as an internal stan- where, AP 0 1 and AP 0 2 are the measured absor-
dard, which quickened sample preparation and did not bance values at 1 and 2, respectively, in the spectrum
necessitate the determination of optical path length. of the pure standard compound.
Quantitative analysis by IR can be performed with The Standard Correction Factor multiplied by the
techniques based on the addition of a standard to the absorbance at the auxiliary frequency 2 in the
sample in order to measure absorbance quotients so prepared sample n AP n 2 , gives the absorbance at
that a variable and unknown spacer may be used as the frequency of interest 1 in the same sample which
measurements are independent of the optical path is only because of the standard, AP n 1
length. An analyte band and a standard band which
are free from interference are required. When the AP n 1 AP n 2 ·F c 1 2 3
unknown band overlaps with one of the standard Arranging the terms in Eq. (1) and substituting Eq.
bands this spectral interference may be corrected by (3) will provide the corrected absorbance at 1 which
the method developed by the authors known as the is only because of the analyte
Absorbance Correction Method (ACM).
AA n 1 An 1 AP n 1
1.1. Absorbance correction method: theoretical 4
background AA n 1 An 1 AP n 2 ·F c 1 2

The ACM permits the analytic use of signals which Once the corrected absorbance value is obtained,
interfere with the signals from the standard. The basis the chosen quantitative method can then be applied.
for this method is as follows:
Let AA n 1 , be the absorbance of the analyte at
2. Experimental
frequency 1 in a prepared sample n (sample stan-
dard) and AP n 1 be the absorbance of the standard at
2.1. Instrumentation
the same frequency, thus giving rise to a spectral inter-
ference in the measurement of the analytical signal.
Perkin-Elmer 1600, Model 1605 FTIR Spectro-
The total measured absorbance at 1 is denoted here as meter with deuterated triglycine sulphate (DTGS)
An 1 , so that
detector. Number of the co-added scans:16; Resolu-
An 1 AA n 1 AP n 1 1 tion: 4 cm 1. Data acquisition and processing soft-
ware Spectrum for Windows, Perkin-Elmer.
Note that in the infrared spectrum the absorbance
bands owing to the same compound show a constant 2.2. Reagents
relation even when the concentration of this
compound or sample thickness is varied, it is possible n-Valeronitrile Aldrich analysis grade; nujol
to predict the absorbance at 1 resulting from the Aldrich IR grade; methyl linoleate Aldrich analysis
 M.C.M. Moya Moreno et al. / Journal of Molecular Structure 482–483 (1999) 551–556
Fig. 1. Infrared spectra of sunflower oil unheated (…) and heated at 300 C during 40 min (—).
grade; methyl elaidate Aldrich analysis grade; and
chloroform Baker analysis grade.
2.3. Samples
Oils from the Mexican and Spanish market were
analysed: Virgin Olive oil (Domeq), Corn oil
(Mazola), Sunflower (Cristal), Seeds oil (Sunflower,
Safflower and Canola, As), and lard.
2.4. Procedure
Samples were heated in a porcelain capsule and
were subjected to increasing heat on an electric
device, up to a temperature of 300 C, during this
process heating was maintained for 20 and 40 min at
the following temperatures: 80 C, 150 C, 200 C,
250 C and 300 C 5 C. The samples and internal
standard n-valeronitrile were mixed in a test tube,
weighed on an analytical scale and the sample was
shaken manually to ensure total homogenisation.
The proportion in weight of n-valeronitrile in the
prepared samples was between 35% and 65%. To
obtain the infrared spectra one or two drops of the
prepared samples were placed, using a Pasteur pipette,
between two circular pieces of well polished potas-
sium bromide crystals without using a spacer so
sample thickness was not controlled but varied in
553
each prepared sample. In the IR spectra of the
prepared samples absorbance was measured as the
peak height at 3008 cm 1 with respect to the base
line tangent at 3116–2718 cm 1, at 970 cm 1 with
respect to the baseline 990–944 cm 1 and the peak
2246 cm 1, corresponding to n-valeronitrile, with
respect to the base line tangent at 2408–2144 cm 1.
Eight mixtures of known composition of n-valero-
nitrile, methyl linoleate and nujol were prepared to
obtain the calibration straight line for the determina-
tion of unsaturation grade. In these mixtures, the
absorbance quotient 3008 cm 1/2246 cm 1 was
measured with respect to the earlier mentioned base-
lines. In the same way eight mixtures of known
composition of n-valeronitrile, methyl elaidate and
nujol were prepared to obtain the calibration straight
line for the determination of trans isomers and the
absorbance quotient (970 cm 1/2246 cm 1) was
measured.
3. Results and discussion
The infrared spectra show how the intense frying
produces a variation in composition of the oils’
nature, causing a decrease in unsaturated components
and an increase in trans isomers (Fig. 1). The
554 M.C.M. Moya Moreno et al. / Journal of Molecular Structure 482–483 (1999) 551–556

Fig. 2. Calibration straight line to obtain the trans isomers percentage as methyl elaidate, using ACM.

percentage of unsaturation expressed as a percentage n-valeronitrile/methyl linoleate/nujol, the calibration

in weight of methyl linoleate (l) and trans isomers
expressed as a percentage in weight of methyl elaidate
(e) was obtained for each type of oil at different
heating. The internal standard method was used with
n-valeronitrile (v) as internal standard. The infrared
spectrum of n-valeronitrile shows an absorbance
band at 2246 cm 1, owing to the stretching vibration
of the triple C N link which can be used as reference
band as it does not interfere with any of the sample
bands. The analytical signal used to determine the
unsaturation was at the peak 3008 cm 1, caused by
the stretching vibration of the C–H link adjoining
the double C C link. From eight mixtures of
straight line was obtained representing the quotient
[A(3008)/A(2246)] against the Cl Cv concentration
quotient, where Cl is methyl linoleate concentration
and Cv the n-valeronitrile concentration both
expressed as %(w/w). Linear regression provided
the following behaviour with a correlation index
0.998: A 3008 A 2246 0 3888 2 0326 Cl Cv .
For determination of the trans isomers, n-valeroni-
trile gives rise to a spectral interference in the
measurement of the peak at 970 cm 1, which can be
corrected by the proposed ACM. The Standard
Correction Factor F c(970, 2246), was obtained as
0.0175, providing the corrected absorbance value

Fig. 3. Infrared spectrum of a prepared sample of the unheated corn oil.

 M.C.M. Moya Moreno et al. / Journal of Molecular Structure 482–483 (1999) 551–556 555

Table 1
Unsaturation percentage (%Un), expressed as methyl linoleate (%w/w) and trans isomers percentage (%t-I), expressed as methyl elaidate
(%w/w) in oils and lard, obtained by internal standard method in FTIR spectroscopy

Sample Heating ( C/min) Olive oil Sunflower oil Corn oil Seeds oil Lard

%Un %t-I %Un %t-I %Un %t-I %Un %t-I %Un %t-I
1 Tamb 18.91 0.56 29.59 0.23 27.43 0.35 23.23 0.47 11.21 0.79
2 80-20 18.83 0.29 29.34 0.21 27.84 0.70 23.32 0.43 10.99 0.62
3 80–40 18.24 0.41 29.71 0.25 27.01 0.85 22.51 0.45 10.86 0.92
4 150–20 18.30 0.42 29.21 0.23 27.10 0.92 22.04 0.46 11.53 1.14
5 150–40 18.53 0.53 29.22 0.29 26.39 0.62 21.17 0.84 11.17 1.29
6 200–20 16.71 0.62 28.34 1.05 24.60 0.87 20.71 0.86 11.07 1.59
7 200–40 12.41 0.64 26.86 1.10 24.12 0.96 19.65 0.83 10.90 1.55
8 250–20 11.85 1.08 26.34 2.01 24.00 0.87 18.83 2.48 8.73 1.26
9 250–40 11.54 1.73 24.89 3.20 23.20 1.24 18.09 2.13 8.85 2.23
10 300–20 9.81 4.23 19.03 7.19 18.67 3.19 16.41 5.57 8.78 2.79
11 300–40 5.65 4.37 11.89 11.45 18.29 5.32 10.94 5.41 8.59 3.62

A C(970 cm 1). In the same way, from eight mixtures
of n-valeronitrile/methyl elaidate/nujol, the calibra-
tion equation obtained was: A C(970)/A(2246)
0.0161 1.3423 Ce/Cv, r 0.998, where Ce is the
methyl elaidate concentration (Fig. 2).
The unsaturation percentage and trans isomers
were calculated from the spectra of the samples
prepared with n-valeronitrile (Fig. 3) using the corre-
sponding calibration equations, the values obtained
are shown in Table 1. The results obtained show
how the intense heating of oils causes a decrease in
unsaturation and increase in trans isomers. It is
possible to establish that in all the oils studied this
degradation starts becoming considerable after
150 C and at higher temperatures, a more significant
variation is apparent. The peroxide index obtained for
each sample by the iodimetric method [9] reveals that
these changes coincide with the decomposition of
hydroperoxides (Fig. 4).
4. Conclusions
FTIR Spectroscopy provides a very useful

Fig. 4. Variation in composition (unsaturation grade, trans isomers and peroxide value) of sunflower oil at different thermal treatment.
556 M.C.M. Moya Moreno et al. / Journal of Molecular Structure 482–483 (1999) 551–556
information on the composition and the extent of
thermal self-oxidation in oils and lard subjected to
intense heat, equivalent to that used in the ordinary
preparation of food. The methodology developed in
this study permitted determination of the percentage
of unsaturated components and trans isomers in the
samples at different heating temperatures, with no
prior treatment. The use of absorbance quotient
measurements permitted quantitative analysis without
requiring a constant and known cell thickness thus
facilitating sample handling and reducing analysis
time. Further it permits quantification in cases where
it is impossible to know the optic path length accu-
rately. ACM, permits the analytical use of signals
which would initially not be valid because of the spec-
tral interference and in this particular case permits the
measurements of signals which are significant for the
information provided on the state of decomposition of
triglycerides.
It can be said that when an edible oil is subjected to
a heating process or frying, the initial nature of the
trigliceride changes significantly and the decrease in
unsaturation and increase in trans isomers is evidence
of the transformation occurring in the essential poly-
unsaturated fatty acids which the oil initially contains,
thus indicating a decrease in the oils’ nutritional
value.
Acknowledgements
Authors are very grateful to the Department of
Foreign Affairs in Mexico for a research grant for
one of the authors (Moya Moreno, M.C.M.).
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