CyTA - Journal of Food

ISSN: 1947-6337 (Print) 1947-6345 (Online) Journal homepage: https://www.tandfonline.com/loi/tcyt20

FTIR spectroscopy combined with chemometrics

for analysis of lard adulteration in some
vegetable oils Espectroscopia FTIR combinada con
quimiometría para el análisis de adulteración con
grasa de cerdo de aceites vegetales

A. Rohman , Yaakob B. Che Man , P. Hashim & A. Ismail

To cite this article: A. Rohman , Yaakob B. Che Man , P. Hashim & A. Ismail (2011) FTIR
spectroscopy combined with chemometrics for analysis of lard adulteration in some vegetable
oils Espectroscopia FTIR combinada con quimiometría para el análisis de adulteración
con grasa de cerdo de aceites vegetales, CyTA - Journal of Food, 9:2, 96-101, DOI:
10.1080/19476331003774639

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CyTA – Journal of Food
Vol. 9, No. 2, August 2011, 96–101

FTIR spectroscopy combined with chemometrics for analysis of lard adulteration in some vegetable
oils
Espectroscopia FTIR combinada con quimiometrı́a para el análisis de adulteración con grasa de
cerdo de aceites vegetales
A. Rohmana,b, Yaakob B. Che Mana,c*, P. Hashima and A. Ismaila
a
Halal Products Research Institute, Universiti Putra Malaysia, 43400 UPM, Serdang, Selangor, Malaysia; bDepartment of
Pharmaceutical Chemistry, Faculty of Pharmacy, Gadjah Mada University, Yogyakarta 55281, Indonesia; cDepartment of Food
Technology, Faculty of Food Science and Technology, Universiti Putra Malaysia, 43400 UPM, Serdang, Selangor, Malaysia
(Received 7 January 2010; final version received 2 March 2010)

This study was aimed to develop a fast technique of Fourier transform infrared (FTIR) spectroscopy for detection
and quantification of lard adulteration in some vegetable oils, namely canola oil (Ca O), corn oil (CO), extra virgin
olive oil (EVOO), soybean oil (SO), and sunflower oil (SFO). The FTIR spectra associated with Ca O, CO, EVOO,
SO, and SFO as well as their blends with lard were scanned, interpreted, and identified. The chemometrics of partial
least square (PLS) and discriminant analysis (DA) at fingerprint regions of 1500–1000 cm71 was used for
quantifying and classifying of lard in the mixture with vegetable oils, respectively. PLS calibration can be successfully
used for quantification of lard in the mixture with vegetable oils, either using normal spectra or its first derivatives.
Furthermore, DA based on Mahalanobis distance can classify lard in vegetable oils.
Keywords: FTIR; lard; vegetable oils; partial least square; discriminant analysis

El objetivo de este estudio fue desarrollar una técnica rápida de espectroscopia de infrarrojos por transformada
Fourier (FTIR) para la detección y cuantificación de adulteración con grasa de cerdo de algunos aceites vegetales,
principalmente aceite de colza (Ca o), aceite de maı́z (CO), aceite de oliva virgen extra (EVOO), aceite de soja (SO)
y aceite de girasol (SFO). El espectrograma FTIR asociado con Ca O, CO, EVOO, SO y SFO, ası́ como sus mezclas
con grasa de cerdo, fueron escaneadas, interpretadas e identificadas. El análisis quimiométrico de mı́nimos
cuadrados parciales (PLS) y análisis discriminantes (DA) en la región de 1500–1000 cm71 se usaron para cuantificar
y clasificar la grasa de cerdo en la mezcla con aceites vegetales, respectivamente. La calibración PLS puede ser usada
satisfactoriamente para la cuantificación de grasa de cerdo en la mezcla con aceites vegetales, bien usando un
espectro normal o sus primeras derivadas. Además, DA basados en la distancia Mahalanobis pueden clasificar la
grasa de cerdo en aceites vegetales.
Palabras clave: FTIR; grasa de cerdo; aceites vegetales; mı́nimos cuadrados parciales; análisis discriminante

Introduction of lard in any food systems is serious problems in

Since very early time, the adulteration of fats and
oils has become serious problem in the market. The
adulteration is sometimes acted deliberately and
sometimes accidentally (Shukla, Dixit, & Singh,
2005) which involves the replacement of high price
oils with cheaper ones. Lard can be viewed from two
perspectives, economical and religion aspects. In
some countries, food producers prefer to mix
vegetable oils with lard to reduce the production
cost. Currently, lard is one the cheapest oils and
commonly available in food industries. Lard or
industrially modified lard could be effectively blended
with other vegetable oils to produce shortenings,
margarines, and other specialty food oils (Marikkar,
Ghazali, Che Man, Peiris, & Lai, 2005). The presence
view of religious concerns, because some religions
like Islam, Judaism, and Hinduism forbid their
followers to consume any foods containing porcine
and its derivatives like lard (Regenstein, Chaudry, &
Regenstein, 2003).
The adulteration of fats and oils is not easy to
detect when the oil adulterant has a composition
approaching with that of the original oil (Rossell,
King, & Downes, 1983). Most initial researches
conducted to analyze the oil adulterant involved a
time consuming and laborious processes such as wet
chemical methods and chromatography techniques.
Therefore, there is a demand to develop rapid and
sensitive method for detecting and quantifying the
adulterants.

*Corresponding author. Email: yaakobcm@gmail.com

ISSN 1947-6337 print/ISSN 1947-6345 online

Ó 2011 Taylor & Francis
DOI: 10.1080/19476331003774639
http://www.informaworld.com

Fourier transform infrared (FTIR) spectroscopy
combined with partial least square (PLS) model has
received a great attention to be used in the quantitative
analysis of lard in cake formulation (Syahariza, Che
Man, Selamat, & Bakar, 2005), chocolate products
(Che Man, Syahariza, Mirghani, Jinap, & Bakar,
2005), and to analyze lard in the mixture with other
animal fats (Che Man & Mirghani, 2001). In authenti-
cation, FTIR combined with chemometrics has been
explored to detect adulterants such as lard in cod-liver
oil (Rohman & Che Man, 2009a), palm oil (Rohman &
Che Man, 2009b), and palm kernel oil (Manaf, Che
Man, Hamid, Ismail, & Syahariza, 2007) in virgin
coconut oil, sunflower, corn, soybean, and hazelnut
oils (Lerma-Garcia, Ramis-Ramos, Herrero-Martinez,
& Simo-Alfonso, 2010), corn oil (Gurdeniz, & Ozen,
2009), and sunflower oil (Downey, Mc Intyre, &
Davies, 2002) in extra virgin olive oil (EVOO), soybean
oil in camellia oil (Wang, Lee, Wang, & He, 2006), and
some vegetable oils in olive oil (Tay, Singh, Krishnan,
& Gore, 2002). However, there is no reported study
related to the application of FTIR spectroscopy for
detection and quantification of lard in some vegetable
oils. Therefore, this study is directed to analyze the
presence of lard as an adulterant in some vegetable oils,
namely corn oil (CO), canola oil (CaO), EVOO,
soybean oil (SO), and sunflower oil (SFO)
using chemometrics of PLS and discriminant analysis
(DA) for quantification and classification of lard,
respectively.
Materials and methods
Sample preparation
Vegetable oils (CaO, CO, EVOO, SO, and SFO) were
purchased from the local market in Serdang, Selangor,
Malaysia with the same batch. Lard was prepared
according to Rohman and Che Man (2009a). Before
being used for FTIR measurement, the lard was
thawed at 50 8C in a water bath.
Quantification of lard in vegetable oils
The calibration model was prepared by making a
series of data set of lard mixed with CaO, CO,
EVOO, SFO, and SO in concentrations of 1–50%
v/v in the neat forms. To complete the homogeniza-
tion, these mixtures were shaken vigorously. For
validation, a number of independent samples of
vegetable oils containing lard were prepared. All
samples were analyzed using FTIR spectrometer. The
spectral regions where the variations were observed
between vegetable oils and those adulterated with
lard were chosen for developing PLS model. The
selection of frequency regions was optimized such a
way that gives the highest values of coefficient of
determination (R2) and the lowest values of root
mean standard error of calibration (RMSEC).
CyTA – Journal of Food 97
Discriminant analysis
For DA, pure vegetable oils (CaO, CO, EVOO,
SFO, and SO) were dissolved in chloroform in
concentration range of 1–50% v/v. Furthermore,
these vegetable oils and lard were mixed to obtain a
series of standard or trained samples and adulterated
samples containing lard of 1–50% v/v in chloroform.
The samples containing lard were assigned as
adulterated, while a series of pure vegetable oils
was marked with pure of corresponding vegetable
oils and classified using DA.
FTIR spectra measurement
FTIR spectra of analyzed oil samples were recorded
using Nicolet 6700 FTIR spectrometer (Thermo
Nicolet Corp, Madison, WI) equipped with deuterated
triglycine sulphate (DTGS) as a detector and potas-
sium bromide (KBr)/Germanium as beam splitter, and
connected to software of the OMNIC operating system
(Version 7.0 Thermo Nicolet). The sampling compart-
ment was Smart Attenuated Total Reflectance kit
(ARK) (Thermo Electron Corp.) with dimension of
10 6 60 mm. The Smart ARK is an advanced multi-
bounce horizontal attenuated reflectance accessory,
producing 12 internal reflections with a penetration
depth (infrared beam) of 2.0 mm. The accessory was
composed of zinc selenide (ZnSe) crystal with an
aperture angle of 458 and refractive index of 2.4 at
1000 cm71. FTIR spectra were collected from 32 scans
at a resolution of 4 cm71 with strong apodization
throughout the mid infrared region (4000–650 cm71).
These spectra were subtracted against background air
spectrum. After every scan, a background of new
reference air spectrum was taken. The ATR plate was
carefully cleaned in situ by scrubbing with hexane twice
followed by acetone and dried with special soft tissue
designed for ATR cleaning before filling in with the
next sample. Cleanliness was verified by collecting a
background spectrum and compare to the previous
one. These spectra were recorded as absorbance values
at each data point in triplicate.
Chemometrics
The software TQ AnalystTM version 6 (Thermo
electron Corporation, Madison, WI) was used for
chemometrics analysis, including PLSand DA. The
optimum number of PLS factors was determined by
cross validation, employing cancellation one standard
at a time by plotting the number of factors against
the root mean standard error of cross validation
(RMSECV) and determining the minimum factor.
The predictability of the models was tested
by computing root mean square error of
prediction (RMSEP) as described by Gurdeniz and
Ozen (2009).
98 A. Rohman et al.

based on data compression and inverse calibration

Results and discussions
FTIR spectral analysis
Mid infrared (MIR) spectroscopy is a rapid and non
destructive technique. This technique was employed to
analyze lard as an adulterant in some vegetable oils,
namely CaO, CO, EVOO, SFO, and SO. The
importance of IR spectroscopy in the elucidation of
molecular structures originates from the much infor-
mation content obtained and the possibility to assign
certain absorption bands related to functional groups.
In fats and oils, most of the peaks and shoulders of the
spectrum are attributable to specific functional groups
(Bendini et al. 2007). The spectrum of lard was
compared with spectra of these vegetable oils. Supple-
mentary Figure 1 shows the typical spectra of lard,
CaO, CO, EVOO, SFO, and SO. The functional
groups together with vibration modes responsible for
infrared absorptions are compiled in Table 1.
FTIR spectra of lard and vegetable oils look very
similar because the main component composed of these
oils are triglycerides. However, due to the fingerprint
technique, meaning that there is no two compounds or
samples having the same spectra in terms of amount
and intensity of peaks, FTIR spectroscopy can be used
to extract the differences among these oils. Upon closer
scrutiny, the minor differences (peak heights) between
lard and vegetable oils at 1117 and 1097 cm71 (l and
m) corresponding to C H bending vibration and
CH deformation vibrations of fatty acids, are
observed. Furthermore, these frequencies, where the
FTIR spectra variations were observed, are used as the
basis for choosing the spectral regions in quantification
and classification of lard in vegetable oils.
Quantification
Quantification of lard in vegetable oils was performed
using multivariate calibration of PLS model. PLS is
(Paradkar, Sivakesava, & Irudayaraj, 2002). In PLS
model, the vegetable oil samples adulterated with 1.0–
50.0% of lard were divided into calibration and
validation sets. Supplementary Figure 1 shows that
lard has similar intensity (peak height) at 1117 and
1097 cm71. Meanwhile, in canola oil (CaO), the peak
height at 1097 cm71 was higher than that at
1117 cm71. Consequently, increasing lard concentra-
tions in CaO will result in increasing of peak height at
1097cm71, approaching the height that matches to the
lard spectrum (Supplementary Figure 2). These differ-
ences were exploited for quantification of lard in CaO
and other vegetable oils.
The spectral region selection is one of analytical
aspects must be taken into account during performing
multivariate calibration because the frequency regions
must be chosen such a way that the analyst describes
the most characteristics constituents of interest to be
determined and to provide non interfered data to
analytes (Cadet & de la Guerdia, 2001). PLS is also
called as full spectrum method, therefore it can be
applied for analysis of lard in some vegetable oils at
whole FTIR spectral regions rather than specific
regions (Faber & Rajko, 2007). For these reasons,
two FTIR spectral regions, namely at whole spectra
(3300–700 cm71) and at selected frequency regions
(1500–1000 cm71), were used for developing PLS
model. Table 2 showed the performance of PLS
calibration model for analysis of lard in some vegetable
oils using different spectral regions.
Because of its ability to give the highest values of R2
and the lowest values of RMSEC compared with other
spectral regions or their combinations, FTIR spectra at
selected fingerprint regions (1500–1000 cm71) were
used for PLS models for quantification of lard in
some vegetable oils. The higher value of R2 and the
lower value of RMSEC indicated the better calibration
model. In order to validate PLS models, the cross

Table 1. Functional groups and modes of vibration in lard and vegetable oilsa.
Tabla 1. Grupos funcionales y modelos de vibración en grasa de cerdo y aceites vegetales.

Assignment Frequency (cm71) Functional group vibration

(a) 3005 cis C¼CH stretching
(b) 2954 Asymmetric stretching vibration of methyl (CH3) group
(c) and (d) 2924 and 2852 Asymmetric and symmetric stretching vibration of methylene ( CH2) group
(e) 1743 Carbonyl (C¼O) from the ester linkage of triacylglycerol
(f) 1654 cis C¼C
(g) 1465 Bending vibrations of the CH2 and CH3 aliphatic groups
(h) 1417 Rocking vibrations of CH bonds of cis-disubstituted alkenes
(i) 1377 Symmetric bending vibrations of CH3 groups
(j) and (k) 1228 and 1155 vibrations of stretching mode from the CO group in esters
(l) and (m) 1111 and 1097 
CH bending and  CH deformation vibrations of fatty acids
(n) 1033 CO stretching
(o) 962 bending vibration of CH functional groups of isolated trans-olefin
(p) 914 Bending vibration of cis  HC CH 
(q) 721 Overlapping of the methylene ( CH2) rocking vibration and to the out of plane
vibration of cis-disubstituted olefins
a
(Source: Guillen & Cabo, 1997; Pavia, Lampman, & Kriz 2001; Vlachos et al. 2006).
 CyTA – Journal of Food 99

validation was carried out using ‘‘leave-one-out’’
technique and the accuracy of each model was assessed
according to the residual mean standard error of
prediction (RMSEP) and R2 value.
The linearity of PLS model was made by plotting
the relationship between the actual value (x-axis) and
FTIR-calculated value (y-axis) of lard. The normal
spectra and those treated with first and second
derivatives were investigated for the optimization of
PLS calibration models. The number of factors or
principal components (PCs) used in the model was
chosen to optimize the model performance and to
minimize errors in the model caused by under-fitting
and over-fitting the data by looking at the residual
variance (Gayo, Hale, & Blanchard, 2006). Supple-
mentary Figure 3 exhibits the scatter plots for the
relationship between actual value of lard in some
vegetable oils and FTIR-calculated value using normal
spectra, either for calibration or for validation. On the
basis of the PLS calibration models, it can be stated
that the presence of lard in some vegetable oils with
level of 1% v/v was possible to detect. In addition,
FTIR method has good method precision. The relative
standard deviation (RSD) obtained was in the range of
2.32–5.68%.
The summary of number of factors, equation
obtained using PLS regression either for calibration
or for validation, and the values of R2, RMSEC, and
RMSEP for analysis of lard in vegetable oils, is
compiled in Table 3. It can be stated that normal
spectra and first derivative give the better results in
term of high value of R2 (4 0.99) and low value of
RMSEC and RMSEP for analysis of lard in Ca O
compared with second derivative treatments. This
trend was also observed for quantification of lard in
CO, SO, and SFO. Meanwhile, the presence of lard in
EVOO is best quantified using first derivative treat-
ment (Table 3).

Table 2. R2 and RMSEC values for analysis of lard in some

Classification
vegetable oilsa.
The classification of pure vegetable oils (Ca O, CO,
Tabla 2. R2 y valores RMSEC del análisis de grasa de cerdo
en aceites vegetales. EVOO, SO, and SFO) and those adulterated with lard
was performed using DA. DA can be used to determine
PLS
the class of vegetable oils and those adulterated by
performance
Frequency computing the distance from each class center in
Analyte regions (cm71) R2 RMSEC Mahalanobis distance units (Tay, Singh, Krishnan, &
Lard in CaO 3300–700 0.880 7.47 Gore, 2002). The results obtained from DA are best
1500–1000 0.999 0.745 visualized using the Coomans plot obtained from
Lard in CO 3300–700 0.927 5.60 Mahalanobis distance of FTIR spectra of standard
1500–1000 0.995 0.964 vegetable oils (Hashim et al. 2010).
Lard in EVOO 3300–700 0.489 13.7
1500–1000 0.997 0.784 The frequencies used for quantification were used for
Lard in SO 3300–700 0.995 0.819 classification (1500–1000 cm71). The samples were
1500–1000 0.997 0.781 classified into two groups: pure vegetable oils and
Lard in SFO 3300–700 0.968 3.85 adulterated samples. Both classes were subjected to
1500–1000 0.999 0.022
DA and the Coomans plot was formed. The Coomans
a
Frequency regions chosen for analysis of lard are italicised. plot is useful for visualizing the classification results. It is

Table 3. PLS calibration and validation models for determination of lard in vegetable oils at frequencies 1500–1000 cm71.
Tabla 3. Calibración PLS y modelos de validación para la determinación de grasa de cerdo en aceites vegetales a frecuencias
1500–1000 cm71.
Equation R2
Vegetable oils Spectra Factor Calibration Validation Calibration Validation RMSEC RMSEP
Canola Normal 2 y ¼ 0.995x þ 0.121 y ¼ 0.980x þ 0.476 0.999 0.998 0.745 0.736
1st derivative 3 y ¼ 1.001x þ 0.029 y ¼ 0.958x þ 0.593 0.998 0.997 0.662 0.960
2nd Derivative 5 y ¼ 1.000x 70.009 y ¼ 0.898x þ 1.547 0.999 0.986 0.096 2.09
Corn Normal 3 y ¼ 0.995x þ 0.070 y ¼ 0.969x 70.025 0.995 0.993 0.964 1.38
1st derivative 3 y ¼ 0.996x þ 0.062 y ¼ 0.963x 7 0.193 0.996 0.990 0.894 1.78
2nd Derivative 5 y ¼ 0.999x þ 0.002 y ¼ 0.924x – 0.585 0.999 0.973 0.202 3.27
EVOO Normal 4 y ¼ 1.003x 7 0.134 y ¼ 1.105x 70.028 0.997 0.992 0.784 1.41
1st derivative 7 y ¼ 0.999x 7 0.001 y ¼ 1.009x þ 0.017 0.999 0.987 0.070 1.99
2nd Derivative 6 y ¼ 0.999x þ 0.009 y ¼ 0.918x þ 2.569 0.999 0.887 0.370 4.70
Soybean Normal 3 y ¼ 0.997x þ 0.048 y ¼ 0.998x þ 0.059 0.997 0.997 0.781 0.818
1st derivative 7 y ¼ 0.998x þ 0.036 y ¼ 0.978x þ 0.726 0.998 0.996 0.686 0.948
2nd Derivative 6 y ¼ 0.999x þ 0.004 y ¼ 0.934x þ 2.329 0.999 0.990 0.233 1.33
Sunflower Normal 8 y ¼ 0.999x 7 0.0001 y ¼ 0.999x þ 0.090 0.999 0.999 0.022 0.677
1st derivative 8 y ¼ 0.999x 7 0.0001 y ¼ 0.981x þ 0.886 0.999 0.993 0.004 1.39
2nd derivative 5 y ¼ 0.999x þ 0.005 y ¼ 0.931x þ 1.993 0.999 0.982 0.289 2.36
100 A. Rohman et al.
created by calculating two independent principal com-
ponents (PCs) models and plotting the residual distances
of samples of each two classes (Gurdeniz, Tokatli, &
Ozen, 2007). The Coomans plot for the classification of
vegetable oils and those adulterated samples with
addition of 1–50% of lard was demonstrated in
Supplementary Figure 3 A–E using 8 PCs for CO and
9 PCs for Ca O, EVOO, SO, and SFO. The x-axis
demonstrates the Mahalanobis distance to the class of
vegetable oils, while the y-axis demonstrates the distance
to the class of the adulterated vegetable oils.
Supplementary Figure 4 clearly demonstrated the
separated group of pure vegetable oils and those
adulterated with lard as visualized by the Coomans
plot. In this study, DA classified 100% of all samples
accurately according to its classes, meaning that no
samples were mistakenly grouped into the incorrect
group unless for soybean oil, which exhibited one
misclassified standards. Sometimes, misclassification
could take place due to the close similarities in
chemical composition among group samples (Manaf,
Che Man, Hamid, Ismail, & Syahariza, 2007).
Conclusion
The adulteration of lard in some vegetable oils
(CaO, CO, EVOO, SO, and SFO) can be
monitored by means of FTIR spectroscopy in
combination with chemometrics of PLS and DA.
PLS using normal and first-derivative spectra at
selected fingerprint region (1500–1000 cm71) was
well suited for determination of lard in some
vegetable oils. DA can classify pure vegetable oils
and those adulterated with lard using eight principal
components (PCs) for CO and nine PCs for Ca O,
EVOO, SO, and SFO.
Supplementary material
The supplementary material for this article is available
online at http://dx.doi.org/10.1080/19476331003774639.
Acknowledgements
A. Rohman is grateful to The Ministry of National
Education, Republic of Indonesia for its scholarship to
pursue Ph.D program in Halal Products Research Institute,
Universiti Putra Malaysia (UPM), Malaysia.
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