Volatile compound profiling from soybean oil in the heating process

Abstract Soybean oil heating or cooking is a very complicated process. In order to better understand the composition of the volatile compounds from soybean oil during heating process, volatile profiling was carried out through vacuum‐assisted headspace solid‐phase microextraction combined with GC‐MS. As a result, a total of 72 volatile compounds were detected and identified during this process, including aldehydes (27), alcohols (14), ketones (10), furans (6), aromatic compounds (9), acids, and esters (6). And the forming temperature of each volatile was determined. Results show most of volatile aldehydes and alcohols were formed at 120°C leading to release off‐flavor largely, which was considered as a critical temperature point for the formation of soybean oil flavor during the whole heating process. Meanwhile, ketones and furans were formed at 150°C, and acids were detected at 180°C. The content of most volatile compounds increased significantly with the temperature raised. Simultaneously, results of principal component analysis demonstrate that flavor characteristics of soybean oil have a big difference between higher and lower temperature in the heating process.

compounds in heating process, which results in forming desired and undesired flavor. Moreover, the undesired flavor causes the rancidity problems of the fried product (Katragadda, Fullana, Sidhu, & Carbonell-Barrachina, 2010;Wu & Chen, 1992). Therefore, heating process is a crucial operation process for flavor forming of the frying oil and fried food.
The heating of soybean oil causes a series of chemical reactions which destroy the structure of unsaturated fatty acids and reduce oil quality. Studies have shown that when the oil temperature reaches 250°C, the content of unsaturated fatty acids decreases, while that of transfatty acids gradually increase (Filip, Fink, Hribar, & Vidrih, 2010;Hou, Wang, Wang, Xu, & Zhang, 2012;Moreno, Olivares, López, Adelantado, & Reig, 1999). Excessive intake of transfatty acids can cause a rise in serum total cholesterol and low-density lipoprotein cholesterol (Mensink & Katan, 1990), which may increase the risk of coronary heart disease (Listed, 1995;Watts, Jackson, Burke, & Lewis, 1996;Xu et al., 2006) and cardiovascular diseases as well as diabetes (Mozaffarian, 2006).
In recent years, researchers paid much attention on studying the composition and characteristics of the off-flavor in frying oil and fried food products. They found a large number of aldehydes in the gases of cooking oil and fried foods, for example, hexanal, heptanal, and pentanal, which causes unpleasant flavor and reduces the selflife of the fried products (Fullana, Carbonell-Barrachina, & Sidhu, 2004;Katragadda et al., 2010;Zhu, Wang, Zhu, & M. K. G., 2001).
But to date, the flavor characteristics of soybean oil throughout the whole heating process has not been studied. Therefore, the purpose of this study is to analyze the volatile compositions of soybean oil generated at different temperatures during the heating process, to determine specific temperatures point at which different volatile compounds generated during heating, and to provide a theoretical basis for soybean oil flavor forming.

| Vacuum-assisted HS-SPME
Samples were heated in a step-by-step heating process. And the heating temperature point was set at 30, 60, 90, 120, 150, and 180°C, taking into consideration their smoke points of soybean oil (Katragadda et al., 2010). The experimental system is shown in Figure 1. This system consists of an oil bath and water bath (DF-101S Heat-gathering Style Magnetism Mixer) to control temperature and a vacuum pump to produce negative pressure and to remove the original gases. An extraction bottle numbered 1 containing 0.1 g soybean oil sample was heated in an oil bath at 30°C and balanced for 40 min to make each volatile substance to reach its saturated vapor pressure. An empty extraction bottle numbered 2 was placed in a water bath and vacuumized for 5 min by using a vacuum pump to −0.1 Mpa. The volatile compounds in the extraction bottle numbered 1 were introduced in the extraction bottle numbered 2 under negative pressure for 3 min. The volatile flavor compounds in the bottle numbered 2 were extracted using SPME fiber (50/30 µm DVB/CAR/PDMS; Supelco, Co.) for 1 hr. After extraction, the SPME with volatile compounds were immediately introduced into the GC-MS for detection. Then, bottle numbered 1 was vacuumized (−0.1 Mpa) for 5 min to remove the residual volatile compounds with aims to eliminate interference caused by volatile flavor compounds generated during the previous temperature treatment and heated to 60°C. Then, the procedure mentioned above was repeated for the following temperature setting points.

| GC-MS analysis
An Agilent 6890/5975C GC-MS system equipped with HP-5 ms column (30 m × 0.25 mm i.d., 0.25 μm film thickness; J&W Scientific) was used to analyze volatile compounds that accumulated on the SPME fiber. The carrier gas was helium with splitless mode, which was delivered at a linear velocity of 1 ml/min. The desorption time was 5 min in the injection port at 250°C. The temperature was programmed to be hold at 35°C for 3 min and increased to 280°C at a rate of 5°C/min. The mass selective detector was operated in the electron impact ionization mode at 70 eV, in the scan range m/z 40-400. The interface temperature was 230°C, and the re-

| Statistical analysis
The experiments for determination of volatile flavor compounds of soybean oil in the heating process were repeated for three times. Data were presented as mean ± standard error. The statistical analysis was performed using SPSS 16.0 software (SPSS Inc.).
Differences between means were evaluated by one-way analysis of variance (Duncan's multiple-range test). Comparisons that yielded p < .05 were considered significant. PCA (principal component analysis) was performed using Unscrambler 9.7 (CAMO Software AS) in order to group the samples according to the class of volatiles.

| Compositions of volatile compounds of soybean oil in the heating process
As shown in Table 1 It was commonly accepted that aldehydes were original from the lipid oxidation (Varlet, Prost, & Serot, 2007). If the reaction begins, it will always be there until the oxygen runs out.
Alcohols are another important class of compounds that make up the volatile flavor of soybean oil during heating process. As it is showed in Table 1, most of alcohols are detected above 120°C.
Only, (E)-2-hepten-1-ol was detected throughout the whole heating process. Alcohols are also considered as the product of fatty acid oxidation (Dong et al., 2013). 1-penten-3-ol has an irritating buttery taste, 1-pentanol has a floral and resinous taste, and 1-hexanol has the smell of grass and flowers.
Most aromatic compounds were detected at <60°C. The reason for this is still unknown. Some acids and esters containing <10 carbon atoms were detected at high temperatures, such as heptanoic and nonanoic acid. Although the amount of esters detected in the process is small, they often bring pleasant, sweet, fruity odors.
Generally, it is considered that most of volatile compounds were original from the lipid oxidation, and the major precursor F I G U R E 3 The forming temperature point of different volatile compounds is lipid hydroperoxides, including alkyl hydroperoxides, allyl hydroperoxides, and fatty ester hydroperoxides. In the lipid oxidation process, these precursors were firstly formed, followed by forming corresponding free radical, which made the lipid schemes decomposed by free radical oxidation, yielding volatile aldehydes.

| Characteristics of volatile compounds composition of soybean oil at different heating temperature
To visualize the formation temperature of different volatiles, the volatile composition of soybean oil at different heating temperature point was shown in Figure 3. The general trends indicated that aromatic compounds and a small amount of aldehydes formed at lower temperatures; while a large number of aldehydes, alcohols, ketones, F I G U R E 4 Heat map of volatile compounds of soybean oil in the heating process. (a) Aldehydes, (b) alcohols and ketones (c) furans, aromatic compounds, acids, and esters and acid ester compounds formed at higher temperatures. A total of 11 volatiles were identified at 30°C during heating, including aldehydes (4), ketones (1), alcohols (1), aromatics (4), acid, and ester (1).
However, it does not mean that these compounds were generated at this temperature point, but the basic volatile ingredients of the soybean oil.
Fishy and beany odor were moderately correlated with (E,E)-2-4decadienal. Grassy, rancid, painty, and acrolein odors of oil were positively correlated with hexanal, (E)-2-hexenal, heptanal, (Z)-2heptenal, and 2-pentylfuran. And buttery and rancid odors were F I G U R E 5 The principal component analysis (PCA) based on the relative content of volatile substances formed in the soybean oil heating process. (a) The scores plot, (b) the loading plot considered as the best indicators of overall odor quality (Brewer, Vega, & Perkins, 1999). Moreover, hexanal, a known lipid oxidation breakdown product, has been used as an indicator of rancidity in a variety of foods; the height of the hexanal peak in a gas chromatogram has been well-correlated with sensory evaluation for rancid (Brewer et al. 1999). Meanwhile, the concentration of hexanal, heptanal, (Z)-2-heptenal, and 2-pentylfuran also increased dramatically at this temperature point (Figure 4). Consequently, 120°C is considered as a critical temperature point for the formation of soybean oil volatile compounds in the whole heating process, and the off-flavor began to release largely. Above this temperature, there are 13 and nine volatile compounds generated at 150 and 180°C, respectively.
Most of ketones are formed at 150°C, and acids are detected at 180°C.
In order to better understand the volatile flavor characteristics of soybean oil at different temperatures, PCA was carried out on the 72 volatile compounds of the heated samples. The peak areas of each compound were normalized, and the first and second principal components (PC1 and PC2) were chosen ultimately. The score scatter and loading scatter plots are shown in Figure 5a,b, respectively.
The scores of soybean oil heated at lower temperature (30, 60, and 90°C) were close to each other and even overlapped, mainly concentrated in the positive loading region of PC1, indicating that the flavor composition below 90°C was similar. Corresponding to the loadings scatter plots (Figure 5b), 2-ethyl-1-hexanol, (E)-6,10dimethyl-5, 9-undecadien-2-one, p-xylene, 1-methyl-naphthalene, and 2,4-di-tert-butylphenol formed a cluster at lower temperatures, indicating that these compounds contribute greatly to the flavor composition of soybean oils at lower temperatures. It also shows that these compounds are detected below 90°C.
It also can be seen from Figure 5a that the scoring points at the higher temperatures (150 and 180°C) and the lower temperature (60, 90, and 120°C) can be clearly separated. They located in the positive and negative region of PC2, respectively. This means that flavor composition of the soybean oil has a big difference between higher and lower temperature. Corresponding to the loadings scatter plots (Figure 5b), the majority of volatile compounds located in this region, which further identify that most of volatile compounds are formed at higher temperatures.
Generally, volatile compounds from soybean oil come from two main lipid oxidation reactions, including photo oxidation and thermal oxidation. Photo oxidation is a slow oxidation process, in which the forming of volatile compounds are both temperature-dependent and time-dependent. Inversely, thermal oxidation is a fast oxidation process. There is no possibility for volatiles forming to be time-dependent because of the rapid raised temperature. And based on the data in this paper, the formation of volatiles shows a certain temperature-dependent trend. As a result, authors consider that volatiles forming of soybean oil are temperature-dependent in the heating process.

| CON CLUS ION
Oil heating or cooking process is a very quick and complex network, during which lots of sharp oxidative reactions occurred rapidly.
Temperature played an important role in the soybean oil flavor forming process, which shows a temperature-dependent trend during heating, and more evidences were needed to draw this conclusion in the future. Meanwhile, the forming temperature of each volatile was finally determined. Subsequently, this research provides a theoretical possibility to modulate flavor production by controlling the temperature in the heating process.

ACK N OWLED G M ENT
We thank the support of the National Natural Science Foundation

CO N FLI C T O F I NTE R E S T
We declare that we have no conflicts of interest.

E TH I C A L A PPROVA L
This study does not involve any human or animal testing.