LWT - Food Science and Technology 96 (2018) 560–567

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LWT - Food Science and Technology

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

Effect of deodorization method on the chemical and nutritional properties of T

fish oil during refining
Gongshuai Songa,b, Mengna Zhanga,b, Xi Penga,b, Xina Yua,b, Zhiyuan Daia,b, Qing Shena,b,∗
a
Institute of Seafood, Zhejiang Gongshang University, Hangzhou, China
b
The Joint Key Laboratory of Aquatic Products Processing of Zhejiang Province, Hangzhou, 310012, China

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

Keywords: Fish oil is a health beneficial product but contains unpleasant odoriferous substances originating from the de-
Fish oil gradation of lipids and proteins. In this study, several deodorization methods including liquid-liquid extraction
Deodorization methods (LLE), green tea polyphenol (GTP) treatment, and solid-phase adsorption (SPA) with activated clay, zeolites or
Sensory evaluation diatomite were studied in order to replace the industrially used molecular distillation (MD) and steam dis-
Fatty acids
tillation (SD). These techniques were compared in terms of physicochemical characteristics, sensory properties,
Principal component analysis
saturability and cis/trans configuration of fatty acids, and volatile compounds, and the results showed significant
difference (p value ≤ 0.05, n = 3). The alkaline ethanol based LLE was superior in reducing acidity, peroxide,
and saponification value, enriching EPA and DHA, removing unpleasant volatiles, and bleaching. Its low tem-
perature condition can well avoid the peroxidation and degradation of lipids and geometrical isomerization of
the natural cis bond. Afterwards, the method was validated and the values for intra-day repeatability (RSDs) fell
within the range of 0.22–1.96%. For the inter-day repeatability, RSDs were less than 5.37% for all analytes. In
conclusion, this method showed advantages with low cost, high efficient, and simplified procedure in the
deodorization of crude fish oil.

1. Introduction fatty acids (FFAs), volatiles, and oxidation products (Menegazzo,

Fish oil, a major source of polyunsaturated fatty acids (PUFAs), has
gained scientific recognition for many years in human nutrition and
disease prevention. The predominant PUFAs are the polyunsaturated
omega-3 acids with four to six double bonds, including eicosapentae-
noic acid (EPA C20:5 n3) and docosahexaenoic acid (DHA C22:6 n3)
(Morais, Mourente, Martinez, Gras, & Tocher, 2015). Fish oil has de-
monstrated the health-promoting effects that include reduction in car-
diovascular disease, disorder of the immune system, inflammatory,
hypertensive and depression effects (Skiba, Poławska, Sobol, Raj, &
Weremko, 2015; Van et al., 2013). Therefore, scientific researches for
obtaining high quality fish oil have been actively carried out.
The criteria for evaluating the quality of fish oil includes the acidity
index (AI), peroxide value (PV), iodine value (IV), saponification value
(SV), the saturability and cis/trans configuration of fatty acids, and the
unpleasant volatiles. Generally, crude fish oil has to undergo refining
steps before being consumed or utilized as food supplements. The re-
fining process comprises the gum conditioning, neutralization, washing,
drying, bleaching, filtration, and deodorization, among which the
deodorization is the most critical step used for primary removing free
Petenuci, & Fonseca, 2014). The traditional methods of deodorization,
molecular distillation (MD) and steam distillation (SD), requires high
temperatures (180–270 °C) and low pressure (0.1–1 kPa) (Merkle et al.,
2017). However, the heat treatment will induce a serious of chemical
transformations (oxidation reactions, cis-trans isomerization, cycliza-
tion, polymerization and/or double bond migration) due to the occur-
rence of numerous methylene-interrupted ethylenic double bonds
(Fournier et al., 2006). Therefore, alternative protocols with mild
conditions for removing odoriferous compounds in crude fish oil re-
fining are concerned.
Nowadays, there are a variety of methods used for small molecules
removing and oil refining, including liquid-liquid extraction (LLE),
solid-phase adsorption (SPA), and green tea polyphenol (GTP) treat-
ment. Alkaline ethanol based LLE is suitable for the separation of heat-
sensitive and low molecular weight compounds. Furthermore, the al-
kaline ethanol has a positive effect on the enzyme hydrolysis efficiency
of cellulose, which is important for modern biorefinery industries (Yang
et al., 2016). Solid-phase adsorption (SPA) using adsorbents, such as
activated clay, zeolites, diatomite, has been applied to remove different
undesirable compounds because of its simple operation conditions and

∗
Corresponding author. Institute of Seafood, Zhejiang Gongshang University, NO. 149, Jiaogong Road, Hangzhou, China.
E-mail addresses: leonqshen@163.com, sq@zjgsu.edu.cn (Q. Shen).

https://doi.org/10.1016/j.lwt.2018.06.004
Received 1 November 2017; Received in revised form 25 May 2018; Accepted 2 June 2018
Available online 04 June 2018
0023-6438/ © 2018 Elsevier Ltd. All rights reserved.
G. Song et al.
high efficiency (Jin et al., 2017; Shen, Yang, Li, & Cheung, 2014). Green
tea polyphenol (GTP) is a natural polymer consisting of six catechins
with significant antioxidant and chelating properties in removing metal
ions and organic matter (Heim, Tagliaferro, & Bobilya, 2002). These
methods were reported to be effective in oil deodorization. However,
their performance in the deodorization of fish oil has not been reported
and systematically compared.
The objectives of this work are to evaluate the effect of different
methods adopted for deodorization on the volatile components and the
fatty acids composition, as well as to provide an alternative for the
replacement of traditional high-temperature methods in the deodor-
ization process of crude fish oil.
2. Materials and methods
2.1. Materials and reagents
The oil samples (10 kg) from by-products of the processed tuna and
anchovies were purchased from Zhoushan Ocean Fish Oil Company
(Zhejiang, China), which were directly collected from the production
line compliant with enterprise standards with AI ≤ 3.0 mg KOH/g,
PV ≤ 5.0 m Eq/kg, and IV ≥ 110.0 g/100 g, insoluble impurities
≤ 0.5%. Standard mixtures of 37 fatty acid methyl esters (FAMEs) were
purchased from ANPEL Laboratory Technologies Inc. (Shanghai,
China). Phosphoric acid, sodium hydroxide, potassium hydroxide,
ethanol, boron trifluoride-methanol complex, acetone, methanol an-
hydrous and n-hexane were analytical grade and purchased from Xilong
Scientific Co. Ltd (Guangzhou, China). Activated clay, zeolites, diato-
mite, and GTP were purchased from Changzhou Dingbang Mineral
Products Technology Co. Ltd (Jiangsu, China).
2.2. Sample preparation
The crude fish oil was processed by degumming and deacidification
of the crude oil extracts according to Crexi (Crexi, Monte, de Souza
Soares, & Pinto, 2010; Li, Sun, Qian, & Liu, 2016). The crude fish oil
was deodorized using the following methods in triplicates (1) molecular
distillation, 100 g crude fish oil was fed to the molecular distillation
unit at a rate of 1.5 mL/min under 10−4 kPa of vacuum. The feeding
tank, condenser and evaporator temperatures were set at 35, 50, and
150 °C, respectively. (2) steam distillation, a total of 100 g of the crude
fish oil was heated to 180 °C with agitation at 300 rpm. The steam
distillation was carried out under vacuum (60 kPa) for 60 min (3) li-
quid-liquid extraction, the deodorizing solvent of alkaline ethanol was
prepared by dissolving 0.5 g potassium hydroxide into 100 mL aqueous
ethanol (45:55, v/v). A total of 20 mL fish oil was added to the alkaline
ethanol and heated to 70 °C with stir, then maintaining the temperature
for 15 min. Afterwards, the oil phase was separated and washed three
times with water until pH = 7. (4) solid-phase adsorption, the solid-
phase adsorption of odoriferous compounds from crude fish oil was
conducted by heating an open vessels containing a stirred slurry of oil
and adsorbent (10:1, w/w) to 70 °C and holding at that temperature for
a period of 15 min. Three kinds of sorbents, activated clay, zeolites, and
diatomite, were tested successively. The deodorized fish oil was got by
filtrating the mixture. (5) GTP treatment, 10 g GTP was accurately
Table 1
Physicochemical characterization of oils.
Physicochemical index Crude fish oil Molecular distillation Steam distillation
a
AI (mg KOH/g) 0.28 ± 0.03 0.07 ± 0.04 b 0.12 ± 0.07 c
a
PV (m Eq/kg) 3.52 ± 0.12 3.71 ± 0.02 b 3.65 ± 0.01 c
a
IV (g/100 g) 118.5 ± 0.3 132.8 ± 0.2 b 130.2 ± 0.1 b
a
SV (mg KOH/g) 221.6 ± 0.2 222.1 ± 0.1 a 223.3 ± 0.1 b
The superscript letters indicate significant levels among the oil samples tested (p < 0.05).
LWT - Food Science and Technology 96 (2018) 560–567
weighed and added to 100 g fish oil in a flask. The mixture was heated
to 60 °C, and agitated at 300 rpm under vacuum (60 kPa) for 20 min.
The deodorized fish oil was obtained after filtrating the mixture. To
ensure the effectiveness, the samples were processed by different op-
erators, and the RSD of the results were controlled to be less than 15%.
The technological process was repeated using three parallel samples.
2.3. HS-SPME-GC/MS
The volatile compounds were analyzed using a headspace solid
phase micro extraction gas chromatography mass spectrometry (HS-
SPME-GC/MS) with a divinylbenzene/carboxen/polydimethylsiloxane
(DVB/CAR/PDMS) fiber. A total of 5 g oil sample was placed into a
15 mL headspace vial and the vial was tightly sealed with a PTFE-lined
cap. The program consisted of equilibrating the sample at 0 °C for
10 min, inserting the fiber into the headspace for chemical adsorption
at 60 °C for 30 min, then transferring the fiber to the injector for des-
orption at 250 °C for 1 min.
A TR-35MS elastic capillary column (30 m × 0.25 mm I.D., 0.25 μm
film, Agilent Technologies Co. Ltd, State of Delaware, USA) was used
for the separation of volatile compounds. The analytical grade helium
gas (purity 99.999%) was used as the carrier gas at a constant flow rate
of 1 mL/min. Each injection was ran in the splitless mode for 1 min with
a split injection port (split ratio 1:10). The GC oven temperature was
programmed at an initial 40 °C for 3 min, increased to 90 °C at a rate of
5 °C/min, then heated to 230 °C at 10 °C/min for 7 min. The mass
spectrum (MS) operated in electron impact mode at 70 eV in the range
of 33–450 m/z (mass-charge ratio) with the source temperature of
250 °C. The interface temperature was 280 °C. The volatiles were
identified by comparing the mass spectra of all detected compounds
with these in NIST library 2.0. The result was accepted when both
matching and reverse matching degree were above 800. The relative
contents of volatile compounds were quantified by calculating the peak
area (Sun, Zhang, Chen, Zheng, & Fang, 2016). Each injection was re-
peated for five times in order to confirm the accuracy of the analytical
method.
2.4. Statistical analysis
Statistical analysis and the calculation of mean, standard deviation,
and level of significance were performed by using Kingsoft WPS soft-
ware and SPSS 16.0 software (SPSS Inc., Chicago, IL, USA). The data
showing significant difference was examined by analysis of variance
(ANOVA, P < 0.05).
3. Results and discussion
3.1. Physicochemical characterization
The AI, PV, IV, and SV of crude and deodorized fish oils were presented
in Table 1. AI is usually associated with free fatty acids (FFA) and un-
desirable flavor present in fats and oils (Crexi et al., 2010). The AI of crude
fish oil was 0.28 ± 0.03 mg KOH/g, and it was reduced after deodoriza-
tion. The AI of deodorized fish oil fell in the range of 0.07–0.23 mg KOH/g.
The most effective method for removing FFA and undesirable flavor was
GTP Liquid-liquid extraction Zeolites Diatomite Activated clay
treatment
0.18 ± 0.03 d 0.15 ± 0.02 e 0.21 ± 0.07 f 0.23 ± 0.03 f 0.19 ± 0.01 d
1.94 ± 0.08 d 2.93 ± 0.02 e 2.34 ± 0.26 f 2.17 ± 0.07 g 2.09 ± 0.03 h
125.3 ± 0.2 c 127.4 ± 0.5 d 126.6 ± 0.2 c 120.8 ± 0.3 e 124.1 ± 0.2 f
224.5 ± 0.2 b 223.8 ± 0.1 b 223.6 ± 0.1 b 220.5 ± 0.2 b 224.0 ± 0.1 b
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MD (0.07 ± 0.04 mg KOH/g), followed by SD (0.12 ± 0.07 mg KOH/g)

and LLE (0.15 ± 0.02 mg KOH/g). In fish oil, the production of hydro-
peroxides is measured as PV, which is a widely employed index for the
measurement of oxidative degradation products in oils. The PV of crude fish
oil was 3.52 ± 0.12 m Eq/kg, and it was reduced to 1.94–3.71 m Eq/kg
after deodorization with different methods. The GTP (1.94 ± 0.08 m Eq/
kg) performed best because of its strong antioxidant and chelating power.
The solid sorbents also showed a significant decrease in PV of fish oils
after SPA, including the zeolites (2.34 ± 0.26 m Eq/kg), diatomite
(2.17 ± 0.07 m Eq/kg), activated clay (2.09 ± 0.03 m Eq/kg). They have
great potential for removing primary oxidation products because of
their favorable sorption properties. On the contrary, when using MD
(3.71 ± 0.02 m Eq/kg) and SD (3.65 ± 0.01 m Eq/kg), the PV of fish oils
increased due to the high temperatures adopted in the deodorization pro-
cess. IV is closely relevant to the proportion of unsaturated fatty acids
(UFAs) in oil (Crexi et al., 2010). No matter which deodorization method
was used, the content of deodorized oils increased significantly. The IV of
crude oil was 118.53 ± 0.25 g/100 g, and it increased to 132.75
± 0.18 g/100 g, 130.16 ± 0.12 g/100 g, 127.42 ± 0.34 g/100 g, 126.61
± 0.23 g/100 g, 125.31 ± 0.20 g/100 g, and 124.05 ± 0.19 g/100 g for
the method of MD, SD, LLE, zeolites, GTP treatment, and activated clay,
respectively. The fish oil deodorized by MD suggests a higher content of
UFAs, because the degradation of unsaturated glycerides and evaporation of
SFAs during the process. In addition, the above mentioned deodorization
techniques will increase the SV of fish oil slightly except the SPA using
diatomite, whose SV (220.51 ± 0.17 mg KOH/g) was lower than the un-
deodorized oil (221.64 ± 0.15 mg KOH/g). This reduction in SV was ex-
plained by the effectiveness of diatomite in removing phosphatides and
soap during the deodorization process (Menegazzo et al., 2014).
The results showed that all the methods can refine crude fish oil
with the indices below the tolerance levels set by the industrial stan-
dard of China (SC/T 3502-2000; AI ≤ 1.0, PV ≤ 5.0 and IV ≥ 120).
Although MD and SD can reduce the AI by removing short chain FFA,
they will create lipid peroxide accumulated in fish oil at high tem-
perature. The LLE, GTP, and SPA performed comparatively better in the
indices of AI, PV, IV, and SV.

3.2. Sensory evaluation

The quadratic discriminant analysis (QDA) of the aroma profiles of

Fig. 1. (A) the physical properties of the fish oil processed with different deo-
representative fish oil processed by different methods were shown in
dorization methods, the samples labeled from 1 to 8 were crude fish oil and
Fig. 1. In fish oil, sensory characterization is associated with the pre- deodorized fish oils processed by activated clay, zeolites, diatomite, liquid-li-
sence of specific fragrance compounds, and it not only can provide quid extraction, GTP treatment, molecular distillation, and steam distillation.
some indication of the overall quality of the oil, but also possibly reflect (B) QDA of organoleptic scores for fish oils deodorized by different methods.
the oxidative state of the material itself (Sandgruber & Buettner, 2012).
The crude fish oil has a strong fishy and sour odor, and in most
identified in the fish oil, with the length of fatty acyl chain distributing
cases, it is not attractive to consumers. After deodorization, the orga-
in C8-C22. There was no trans fatty acid (trans-FA) formation. The pro-
noleptic characterization varied depending on the deodorization
portions of SPA, MUFA, and PUFA were 36.93 ± 0.27%, 28.92
method used. The color of activated clay processed fish oil was yellow
± 0.29%, and 34.15 ± 0.22%, respectively. C16:0 (22.07 ± 0.06%)
and transparent, which was significantly different from the others.
is the main constituent, follow by the DHA (18.69 ± 0.03%) and C18:1
However, a grassy smell was generated and mixed with a light fishy
cis (16.62 ± 0.03%). The sum of EPA and DHA accounts for
odor, leading to an unpleasant flavor to the panelists. The color of fish
28.17 ± 0.11% of fish oil.
oil samples processed by SPA (zeolites and diatomite) and MD turned to
The identity and relative content of each SFA, MUFA and PUFA in
orange, while the rest fish oil samples were reddish. From the reflect of
the deodorized fish oil were shown in Table 3. The composition of fish
panelists, the effect of MD and LLE on the odor of fish oil was significant
oil and the relative content of each fatty acid varied depending on the
than the others, which could remove the main undesirable volatile
deodorization methods of MD, SD, GTP treatment, LLE, zeolites, dia-
compounds and improve the sensory quality of the oil effectively. The
tomite and activated clay. Oleic acid (C18:1 cis) accounted for 59.06%,
deodorized oils processed by the methods of activated clay, zeolites,
59.61%, 59.76%, 59.85%, 58.83%, 56.73% and 57.73% of the total
diatomite and molecular distillation arrived primary standard. The re-
MUFAs in the oil processed by the above methods. The PUFAs per-
sults indicated that LLE is comparable with MD and SD in terms of
centage is a useful indicator for comparing the nutritional values of fish
sensory characteristics.
oils (Elsen et al., 2013). EPA and DHA accounted for the highest per-
centage of the PUFAs in the oil processed by different methods (Özogul,
3.3. Fatty acid composition and cis/trans configuration
Özogul, Çiçek, Polat, & Kuley, 2009). EPA is the precursor of pros-
taglandins, thromboxanes and leukotrienes. DHA considered as the
The fatty acid composition in the crude oil was analyzed and the
most important PUFAs for the component of the phospholipid
results were shown in Table 2. A total of 20 fatty acid species were

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Table 2
Fatty acid profile of the crude fish oil.
Fatty acids Saturated (SFA)
C12:0 C13:0 C14:0 C15:0 C16:0 C17:0 C18:0 C20:0 ∑SFA
0.16 ± 0.03 0.10 ± 0.02 6.87 ± 0.01 1.27 ± 0.11 22.07 ± 0.06 1.31 ± 0.01 4.50 ± 0.02 0.60 ± 0.02 36.93 ± 0.27

Monounsaturated (MUFA)
C14:1 C16:1 C17:1 C18:1 cis C20:1 C22:1 ∑MUFA
0.30 ± 0.12 7.27 ± 0.08 0.88 ± 0.01 16.62 ± 0.03 2.30 ± 0.05 1.55 ± 0.01 28.92 ± 0.29
Polyunsaturated (PUFA)
C18:2 cis C18:3 n6 C18:3 n3 C20:4 EPA DHA ∑PUFA
2.22 ± 0.04 0.14 ± 0.01 1.75 ± 0.05 1.87 ± 0.01 9.48 ± 0.08 18.69 ± 0.03 34.15 ± 0.22

∑SFA: Sum of the saturated fatty acids; ∑MUFA: Sum of the monounsaturated fatty acids; ∑PUFA: Sum of the polyunsaturated fatty acids.

membrane of brain and retina cells (Zhong, Madhujith, Mahfouz, &
Shahidi, 2007). Furthermore, some studies showed that the EPA-rich
and DHA-rich diets are effective for reducing allergy symptoms. The
methods of MD, SD, GTP treatment, and LLE have positive effect on the
purification of EPA and DHA from 82.49% to 90.77%, 90.62%, 88.50%,
88.65%, respectively. There was no effect of zeolites, diatomite and
activated clay on the PUFAs of fish oil, which was related to physical
adsorption under low temperature and short time.
The trans fatty acid, C18:2 trans, was found in the deodorized fish
oil processed by MD and SD, with relative contents 0.02 ± 0.01% and
0.06 ± 0.09%, respectively. It could be attributed to that the tem-
perature and heating time used are sufficient to induce geometrical
isomerization of the natural cis double bonds of PUFAs (e.g. α-linolenic
acid) to the more thermo dynamically stable trans configuration (Li, Ha,
Wang, Li, & Li, 2012). The trans-FAs intake has been suggested to in-
fluence systemic inflammation, and may increase insulin resistance
(Micha & Mozaffarian, 2009; Kien et al., 2013). The World Health Or-
ganization recommended that trans-fat intake be limited to less than 1%
of the overall energy intake. Therefore, it was imperative to optimize
deodorization process conditions, and employ the optimal method for
avoiding causing essential fatty acids probably undergo a variety of
chemical and physical changes due to thermal decomposition, and
temperature is regarded as the determining parameter to be controlled
during the deodorization of fish oil (Fournier et al., 2007). Other
methods could well avoid the generation of trans-FA, because the fish
oil was processed under low temperature and short time.
3.4. Volatile components analysis
The volatile components in fish oil are mainly originated from the
microbial spoilage and autoxidation of proteins, amino acid and lipids.
A refining process must be employed to remove the unpleasant com-
ponents, especially the aldehydes and ketones, which are responsible
for the characteristic odor of fish oil (Oliveira & Miller, 2014). In this
study, the composition and proportion of the volatile compounds were
identified and shown in Table 4. There were 35 compounds of identified
volatile compounds in the deodorized oils processed by different
methods, including 8 kinds of alcohols, 7 kinds of aldehydes, 5 kinds of
ketones, and 15 kinds of hydrocarbons. Their relative contents were
calculated by the area normalization method of the compounds and

Table 3
The composition and relative contents of fatty acids in different kinds of fish oil processed by different methods(%).
Fatty acid composition Molecular distillation Steam distillation GTP Liquid-liquid extraction Zeolites Diatomite Activated clay
treatment

C12:0 0.13 ± 0.04 a 0.12 ± 0.07 b 0.14 ± 0.03 c 0.14 ± 0.02 c 0.13 ± 0.02 a 0.13 ± 0.01 a 0.15 ± 0.04 d
C14:0 5.67 ± 0.01 a 5.75 ± 0.02 b 5.99 ± 0.01 c 6.05 ± 0.04 d 6.02 ± 0.01 dc 6.18 ± 0.01 e 6.13 ± 0.04 f
C16:0 20.04 ± 0.05 a 20.44 ± 0.04 b 20.78 ± 0.04 c 21.13 ± 0.08 d 20.66 ± 0.07 c 20.97 ± 0.07 e 20.66 ± 0.09 c
C17:0 0.85 ± 0.02 a 0.99 ± 0.01 b 1.54 ± 0.08 c 0.93 ± 0.02 d 1.85 ± 0.11 e 1.78 ± 0.01 f 1.43 ± 0.14 g
C18:0 4.26 ± 0.00 a 4.01 ± 0.02 b n.d. n.d. 4.52 ± 0.04 c 4.50 ± 0.06 c 4.43 ± 0.09 d
∑SFA 32.04 ± 0.18 a 32.38 ± 0.27 a 29.57 ± 0.21 b 29.47 ± 0.25 b 34.64 ± 0.37 cd 34.77 ± 0.28 c 34.20 ± 0.54 d
C14:1 0.33 ± 0.01 a 0.27 ± 0.03 b 0.19 ± 0.07 c 0.30 ± 0.10 d 0.10 ± 0.10 e 0.26 ± 0.03 b 0.28 ± 0.10 f
C16:1 7.21 ± 0.12 a 7.23 ± 0.07 ab 7.24 ± 0.02 b 7.17 ± 0.04 c 6.85 ± 0.07 d 6.96 ± 0.00 e 6.87 ± 0.06 d
C17:1 1.29 ± 0.08 a 1.19 ± 0.06 b 1.32 ± 0.01 c 1.31 ± 0.01 c 0.92 ± 0.04 d 0.95 ± 0.12 e 0.87 ± 0.05 f
C18:1 cis 18.77 ± 0.05 a 19.01 ± 0.01 b 19.25 ± 0.11 c 19.29 ± 0.06 c 17.22 ± 0.03 d 16.73 ± 0.13 e 16.88 ± 0.03 e
C20:1 2.56 ± 0.03 ab 2.56 ± 0.02 ab 2.57 ± 0.02 a 2.55 ± 0.01 b 2.90 ± 0.04 c 2.99 ± 0.08 d 2.77 ± 0.03 e
C22:1 1.62 ± 0.02 a 1.63 ± 0.01 ab 1.64 ± 0.04 b 1.61 ± 0.10 ac 1.48 ± 0.06 d 1.60 ± 0.02 ac 1.57 ± 0.07 e
∑MUFA 31.78 ± 0.31 a 31.89 ± 0.20 a 32.21 ± 0.27 b 32.23 ± 0.32 b 29.27 ± 0.34 c 29.49 ± 0.38 c 29.24 ± 0.34 c
C18:2 trans 0.02 ± 0.01 a 0.06 ± 0.09 b n.d. n.d. n.d. n.d. n.d.

C18:2 cis 2.00 ± 0.02 a 2.00 ± 0.06 a 2.03 ± 0.04 b 2.01 ± 0.05 a 2.03 ± 0.03 b 2.06 ± 0.02 c 2.18 ± 0.05 d
C18:3 n6 n.d. n.d. n.d. n.d. 0.12 ± 0.01 a 0.13 ± 0.00 ab 0.13 ± 0.02 ab
C18:3 n3 1.34 ± 0.11 a 1.40 ± 0.02 b 2.37 ± 0.01 c 2.34 ± 0.02 c 1.59 ± 0.02 d 1.68 ± 0.04 e 1.58 ± 0.02 d
C20:4 n.d. n.d. n.d. n.d. 2.39 ± 0.01 a 2.27 ± 0.01 b 2.41 ± 0.02 c
EPA 10.54 ± 0.00 a 10.58 ± 0.04 a 10.88 ± 0.06 b 10.89 ± 0.07 b 9.61 ± 0.02 c 9.46 ± 0.02 d 9.69 ± 0.01 c
DHA 22.30 ± 0.04 a 22.27 ± 0.01 a 22.97 ± 0.04 b 23.07 ± 0.05 b 19.97 ± 0.06 c 20.06 ± 0.01 c 20.15 ± 0.02 d
∑EPA + DHA 32.84 ± 0.04 a 32.85 ± 0.05 b 33.85 ± 0.10 c 33.96 ± 0.12 c 29.58 ± 0.08 d 29.52 ± 0.03 d 29.84 ± 0.03 e
∑PUFA 36.18 ± 0.18 a 36.31 ± 0.22 a 38.25 ± 0.15 b 38.31 ± 0.19 b 35.79 ± 0.18 c 35.66 ± 0.10 c 36.10 ± 0.14 d
∑UFA 67.96 ± 0.51 a 68.20 ± 0.42 a 70.46 ± 0.42 b 70.54 ± 0.51 b 65.06 ± 0.52 c 65.15 ± 0.48 c 65.34 ± 0.48 c

The superscript letters indicate significant levels among the oil samples tested (p < 0.05).
∑SFA, Sum of saturated fatty acids; ∑MUFA, Sum of monounsaturated fatty acids; ∑PUFA, Sum of the polyunsaturated fatty acids; ∑UFA, Sum of unsaturated fatty
acids.
n.d., not detected.

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Table 4
The identities and contents of volatile compounds in deodorized fish oils processed by different methods.
Volatile compounds Relative content of volatile compounds/%

Crude fish oil Molecular distillation Steam distillation GTP treatment Liquid-liquid extraction Zeolites Diatomite Activated clay

a b c d e f
Alcohol 1-Penten-3-ol 0.83 ± 0.03 0.33 ± 0.03 3.17 ± 0.08 0.50 ± 0.04 n.d. 0.47 ± 0.05 0.75 ± 0.04 0.52 ± 0.04 d
a
Cyclopentanol 1.21 ± 0.06 n.d. n.d. 0.68 ± 0.06 b n.d. n.d. n.d. 0.97 ± 0.05 c
a
1-Pentyn-3-ol 0.96 ± 0.04 n.d. 4.49 ± 0.13 b 1.14 ± 0.05 c n.d. 0.54 ± 0.05 d 0.43 ± 0.05 e 1.74 ± 0.06 f
a
1-Hexanol 0.92 ± 0.13 n.d. n.d. 0.86 ± 0.07 b n.d. 0.57 ± 0.06 0.67 ± 0.04 d 0.67 ± 0.04 d
a c
2-Ethylhexanol 0.87 ± 0.06 n.d. n.d. 0.22 ± 0.02 b 0.16 ± 0.02 0.81 ± 0.07 d 1.10 ± 0.04 e 0.19 ± 0.02 c
a c
4-ethylBenzenemethanol 3.35 ± 0.10 n.d. n.d. 3.71 ± 0.12 b 3.06 ± 0.06 n.d. 3.14 ± 0.05 d 0.90 ± 0.08 e
a
Cyclopentadecanol 0.11 ± 0.02 n.d. n.d. n.d. n.d. n.d. n.d. n.d.
a
(Z)-9-Hexadecen-1-ol 0.15 ± 0.03 n.d. n.d. n.d. n.d. 0.17 ± 0.03 b n.d. 0.09 ± 0.01 c
a c e
Total 8.43 ± 0.15 0.33 ± 0.03 b 7.66 ± 0.20 7.12 ± 0.28 d 3.22 ± 0.06 2.56 ± 0.12 f 6.08 ± 0.11 g 5.07 ± 0.24 h
a
Aldehyde Butanal 0.58 ± 0.05 n.d. n.d. n.d. n.d. n.d. n.d. n.d.
a e c
Hexanal 2.29 ± 0.15 1.07 ± 0.07 b 1.28 ± 0.04 c 1.75 ± 0.05 d 0.66 ± 0.04 1.48 ± 0.11 f 1.26 ± 0.04 2.19 ± 0.03 g
a e a
Nonanal 2.67 ± 0.08 0.79 ± 0.05 b 3.25 ± 0.09 c 2.20 ± 0.08 d 1.89 ± 0.09 3.33 ± 0.08 f 2.72 ± 0.07 2.36 ± 0.04 g
a e
2-Ethylcrotonaldehyde 1.73 ± 0.06 n.d. 0.41 ± 0.03 b 0.15 ± 0.02 c n.d. 0.80 ± 0.03 d 0.34 ± 0.04 2.04 ± 0.06 f

c e
Undecanal 1.88 ± 0.07 a 0.22 ± 0.02 b n.d. 1.90 ± 0.10 a 1.15 ± 0.05 n.d. 2.07 ± 0.06 d 2.63 ± 0.07
e
Dodecanal 1.13 ± 0.03 a 0.53 ± 0.04 b 1.53 ± 0.06 c 1.03 ± 0.06 d 0.62 ± 0.04 3.26 ± 0.07 f 1.09 ± 0.05 g n.d.
c
Benzaldehyde 0.53 ± 0.04 a n.d. 1.47 ± 0.07 b 0.51 ± 0.05 c n.d. 0.83 ± 0.05 d 0.75 ± 0.08 e 0.51 ± 0.02
e f
Total 10.81 ± 0.38 a 2.62 ± 0.07 b 7.95 ± 0.19 c 7.54 ± 0.18 d 4.32 ± 0.09 9.71 ± 0.22 f 8.23 ± 0.25 g 9.73 ± 0.11
Ketone 2-Butanone 0.75 ± 0.04 a n.d. n.d. n.d. n.d. n.d. n.d. n.d.
c

564
2-Heptanone 0.93 ± 0.04 a n.d. n.d. 0.37 ± 0.04 b n.d. 0.63 ± 0.06 0.55 ± 0.04 d n.d.
e e
2-Nonanone 1.23 ± 0.10 a n.d. 0.69 ± 0.03 b 0.90 ± 0.06 c 0.57 ± 0.04 d 1.76 ± 0.07 1.13 ± 0.08 f 1.75 ± 0.03
a
2-Decanone 0.27 ± 0.04 a n.d. n.d. n.d. n.d. 0.26 ± 0.04 0.25 ± 0.04 ac n.d.
e
2-Undecanone 0.89 ± 0.05 a 0.10 ± 0.01 b 1.37 ± 0.06 c 0.89 ± 0.06 a 0.70 ± 0.06 d 0.96 ± 0.06 0.81 ± 0.07 f 1.06 ± 0.04 g
f
Total 4.07 ± 0.19 a 0.10 ± 0.01 b 2.07 ± 0.08 c 2.16 ± 0.08 d 1.27 ± 0.06 e 3.60 ± 0.18 2.75 ± 0.17 g 2.81 ± 0.06 h
Hydrocarbon Octane 1.22 ± 0.08 a n.d. 1.36 ± 0.10 b 1.14 ± 0.07 c n.d. n.d. n.d. 1.21 ± 0.06 a
Nonane 1.29 ± 0.03 a n.d. 0.67 ± 0.05 b 1.40 ± 0.12 c 1.01 ± 0.08 d 0.63 ± 0.06 b 0.79 ± 0.09 e 1.32 ± 0.04 f
Naphthalene 0.79 ± 0.03 a 0.15 ± 0.02 b 0.93 ± 0.04 c n.d. n.d. 0.40 ± 0.05 d 0.38 ± 0.05 d 0.45 ± 0.05 e
1-Methylnaphthalene 1.34 ± 0.13 a n.d. n.d. n.d. n.d. n.d. n.d. n.d.
2,4,5-Trimethylphenol 0.42 ± 0.03 a n.d. n.d. n.d. n.d. 0.49 ± 0.06 b n.d. n.d.
a ad
Undecane 0.28 ± 0.02 a n.d. 0.86 ± 0.05 b 0.27 ± 0.04 0.28 ± 0.04 a 0.23 ± 0.02 c 0.26 ± 0.04 0.46 ± 0.04 e
a c
Dodecane 0.25 ± 0.03 a n.d. 0.26 ± 0.04 a 0.25 ± 0.03 0.37 ± 0.06 b 0.22 ± 0.02 c 0.21 ± 0.03 0.28 ± 0.04 d

Tetradecane 1.44 ± 0.11 a 0.34 ± 0.04 b 3.27 ± 0.12 c n.d. 2.05 ± 0.06 d 1.92 ± 0.03 e 2.22 ± 0.06 f 1.75 ± 0.05 g
Pentadecane 36.65 ± 0.24 a 5.88 ± 0.11 b 37.26 ± 0.16 c 36.30 ± 0.20 c 33.40 ± 0.10 d 42.77 ± 0.13 e 34.28 ± 0.09 f 38.86 ± 0.14 g
Hexadecane 1.06 ± 0.06 a 0.28 ± 0.02 b 1.44 ± 0.06 c 1.28 ± 0.05 d 0.94 ± 0.05 e 1.11 ± 0.05 f 2.05 ± 0.05 g 1.24 ± 0.09 d
Heptadecane 12.42 ± 0.13 a 1.24 ± 0.07 b 13.41 ± 0.08 c 13.41 ± 0.16 c 12.66 ± 0.13 d 15.40 ± 0.10 e 12.30 ± 0.10 b 13.79 ± 0.12 f
1-Acetyl-1-cyclohexene 0.20 ± 0.03 a n.d. 3.31 ± 0.06 b 0.22 ± 0.03 c n.d. 0.14 ± 0.01 d n.d. n.d.
2,6,10,14-tetramethylpentadecane 11.72 ± 0.13 a 0.72 ± 0.05 b 12.53 ± 0.11 c 12.28 ± 0.06 c 11.46 ± 0.08 d 13.66 ± 0.06 e 11.44 ± 0.11 d 12.52 ± 0.09 c
1-Tridecene 0.38 ± 0.04 a n.d. 1.16 ± 0.07 b 1.07 ± 0.04 c 1.53 ± 0.07 d 1.08 ± 0.04 c 0.92 ± 0.05 e 0.61 ± 0.03 f
1-Pentadecene 0.43 ± 0.03 a n.d. n.d. n.d. n.d. n.d. 0.47 ± 0.07 b n.d.
Total 69.91 ± 0.59 a 8.60 ± 0.40 b 76.46 ± 0.45 c 67.65 ± 0.43 a 63.71 ± 0.35 d 78.04 ± 0.38 c 65.32 ± 0.39 d 72.48 ± 0.50 e

The superscript letters indicate significant levels among the oil samples tested (p < 0.05).
n.d., not detected.
LWT - Food Science and Technology 96 (2018) 560–567
G. Song et al.
Fig. 2. A comparison between the deodorization methods in terms of the alcohols, hydrocarbons, aldehydes, and ketones.
depicted in Fig. 2. The frequently occurred toxic and bioaccumulating
contaminants, such as dibenzo-p-dioxins, dibenzofurans, dioxin-like
polychlorinated biphenyls, and non-dioxin-like polychlorinated biphe-
nyls were not found. The difference on the sum of alcohols, aldehydes,
ketones and hydrocarbons in the oil processed by different methods
were compared, and the performance of each method for removing the
undesirable volatile components varied greatly. A representative mass
spectrum of volatile compounds was shown in Fig. 3.
Aldehydes are the major component of fishy smell, which have a
highly intense odor and an overwhelming impact on overall aroma due
to their lower odor thresholds. In this study, the aldehydes accounted
for 10.92% of the total volatile compounds in the crude fish oil, in-
cluding 6 alkenals (butanal, hexanal, nonanal, undecanal, dodecanal),
and 2 aromatic aldehydes (2-ethylcrotonaldehyde, benzaldehyde).
These volatile compounds were mostly generated by the autoxidation of
lipids and associated with the fish species, the conditions of fish oil
refinement, especially temperature, oxygen, chemical reagents, and
light or metal content. During the deodorization, the effect of aldehydes
removal is highly depending on the above mentioned methods. For
quality control, the fish oil was regarded as well deodorized when it
total content of aldehydes was ≤5%. In detail, the MD can remove most
of the aldehydes and reduce its relative content to 2.62%. LLE is also an
ideal method, which can reduce the relative content of aldehydes to
3.32%. SD and SPA are not suggested because some aldehydes were
generated and their contents are even higher than those of the original
crude fish oil sample, such as the nonanal and dodecanal. Besides, the
GTP treatment has a limited effect on the aldehydes removal.
Ketones are usually produced by lipid autoxidation of fatty acids or
the autoxidation of unsaturated fatty acids via hydroperoxides (Giri,
Osako, & Ohshima, 2010). They accounted for the lowest percentage of
the all volatile components in the crude fish oil (4.07 ± 0.19%).
However, ketones have low odor thresholds, the effect of which on the
overall flavor in the oils was significant. The results indicated that the
MD (0.10 ± 0.01%) and LLE (0.17 ± 0.02%) can remove almost all
the ketones. The effectiveness of the rest methods are not significant,
such as SD (2.07 ± 0.08%), GTP treatment (2.16 ± 0.08%), zeolites
565
LWT - Food Science and Technology 96 (2018) 560–567
(3.60 ± 0.18%), diatomite (2.75 ± 0.17%), and activated clay
(2.81 ± 0.06%).
Alcohols are also formed by secondary decomposition of hydro-
peroxides of fatty acids, attributing to the enzymatic peroxidation of n-
3 and n-6 polyunsaturated fatty acids (Giri et al., 2010). The alcohols in
crude oil accounted for 8.43 ± 0.15% of the total volatile compounds
in the crude fish oil. MD (0.33 ± 0.03%) can remove almost all the
alcohols, followed by the LLE (0.52 ± 0.04%). The alcohols do not
exert much impact on the overall aroma because they have relatively
high odor threshold values except the unsaturated alcohols such as 1-
penten-3-ol. In fact, a significant decrease of 1-pentyn-3-ol and 1-
penten-3-ol was observed in the deodorized oils processed by the
methods of MD and LLE. The methods of SD and active clay were not
suggested because they will increase the content of unsaturated alco-
hols by 2–4 times. The rest methods have a limited effect on removing
alcohols. The proportion of hydrocarbons was the highest in the fish oil.
However, due to the high odor thresholds of hydrocarbons, these
compounds have little effect on the overall aroma of fish oil.
4. Conclusion
The odor of fish oil is an important criteria for quality evaluation.
The industrially used MD is effective in deodorization but the high
temperature condition will induce geometrical isomerization of the
natural cis double bonds of PUFAs. In this study, several deodorization
methods were compared in terms of physicochemical characteristics,
sensory properties, saturability and cis/trans configuration of fatty
acids, and volatile compounds for the replacement of the traditional
methods. The results indicated that the alkaline ethanol based LLE was
superior in all aspects, and it performance is comparable with MD
without inducing the generation of trans FFA. The major component of
fishy smell, aldehydes, can be reduced to 3.32% by LLE. Besides, the
scale of this technique can be easily magnified industrially by blending
more sample and alkaline ethanol in a reaction tank, and the solvent
can be well recovered by rectification.
G. Song et al. LWT - Food Science and Technology 96 (2018) 560–567

Fig. 3. (a) A representative gas chromatographic profile of FAMEs in deodorized fish oil, the peaks were 1. C12:0; 2. C14:0; 3. C14:1; 4. C16:0; 5. C16:1; 6. C17:0; 7.
C17:1; 8. C18:0; 9. C18:1 cis; 10. C18:2 cis; 11. C18:3; 12. C20:1; 13. C22:1; 14. C20:5; and 15. C22:6, (b) the mass spectrum of volatiles in deodorized fish oil.

5. Competing financial interest
The authors declare no competing financial interest.
Acknowledgments
The authors thank the Zhejiang Provincial R&D Program
(2017C03041), the Natural Science Fund for Young Scholars of China
(No. 31601542), and the Public Welfare Research Program of Zhejiang
Province (LGN18C200001).
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://dx.
doi.org/10.1016/j.lwt.2018.06.004.
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