Effects of Fe3+ and Antioxidants on Glycidyl Ester Formation in Plant

May 3, 2017 - This research investigated the effects of Fe3+ and antioxidants on the ... at 200 °C. Results show that Fe3+ can promote the formation ...
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Effects of Fe3+ and Antioxidants on Glycidyl Esters Formation in Plant Oil at High Temperature and Their Influencing Mechanisms Wei Wei Cheng, Guoqin Liu, and Xinqi Liu J. Agric. Food Chem., Just Accepted Manuscript • Publication Date (Web): 03 May 2017 Downloaded from http://pubs.acs.org on May 7, 2017

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Journal of Agricultural and Food Chemistry

Effects of Fe3+ and Antioxidants on Glycidyl Esters Formation in Plant Oil at High Temperature and Their Influencing Mechanisms

Weiwei Cheng†, Guoqin Liu*, †, ‡, §, and Xinqi Liu⊥, #



School of Food Science and Engineering and ‡Guangdong Province Key Laboratory for

Green Processing of Natural Products and Product Safety, South China University of Technology, Guangzhou 510640, China §

College of Food Science and Engineering, Henan University of Technology, Zhengzhou

450001, China ┴

Beijing Advanced Innovation Center for Food Nutrition and Human Health and #School of

Food and Chemical Engineering, Beijing Technology and Business University, Beijing 100048, China

Corresponding author *G. Liu, Tel: 8620-8711-3875. Fax: 8620-8711-4262. E-mail: [email protected].

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ABSTRACT: This research investigated the effects of Fe3+ and antioxidants on glycidyl

2

esters (GEs) formation and the free radical-mediated mechanisms involving the recognition

3

of cyclic acyloxonium free radical intermediate (CAFRI) for GEs formation in both plant oil

4

model (palm oil, camellia oil, soybean oil, and linseed oil) system and chemical model

5

(dipalmitin and methyl linoleate) system heated at 200 °C. Results show that Fe3+ can

6

promote the formation of GEs, which can be inhibited by antioxidants in plant oil during

7

high-temperature exposure. Based on the monitoring of cyclic acyloxonium and ester

8

carbonyl group by Fourier transform infrared spectroscopy, the promotion of Fe3+ and the

9

inhibition of antioxidants (tert-butylhydroquinone and α-tocopherol) for GEs formation

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occurred not only through lipid oxidation but also through directly affecting the formation of

11

cyclic acyloxonium intermediate. Additionally, a quadrupole-time of flight tandem mass

12

spectrometry measurement was conducted to identify the presence of radical adduct

13

captured by 5,5-dimethylpyrroline-N-oxide, which provided strong evidence for the

14

formation of CAFRI. Thus, one possible influencing mechanism can be that free radical

15

generated in lipid oxidation may be transferred to dipalmitin and promote CAFRI formation.

16

Fe3+ can catalyze free radical generation while antioxidants can scavenge free radical, and

17

therefore they also can directly affect CAFRI formation.

18

KEYWORDS: glycidyl esters, plant oil, iron ion, antioxidants, lipid oxidation, influencing

19

mechanisms

20 21 22 23

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Introduction

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In 2010, glycidyl esters (GEs) of fatty acids, a group of new potential food-processing

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contaminants, was first be detected in edible oils and fats due to the regular overestimation

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of 3-monochloropropane-1,2-diol (3-MCPD) esters of fatty acids analyzed by early indirect

28

methods.1 Since then, plentiful literature has reported that GEs are mainly formed in the

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deodorization step during oil refining and therefore widely existed in refined oils, fats, and

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oil-based food, including infant formula.2-6 Although there is no available evidence for the

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detrimental effect of GEs on human and animal, the hydrolysate, glycidol, has been

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determined to be genotoxic and carcinogenic to rats7 and has already been categorized as

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group 2A carcinogen by the Intl. Agency for Research on Cancer (IARC).8 Therefore, to

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develop the efficient elimination and inhibition method for reducing GE levels in edible oils,

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the further influencing factors and formation mechanisms should be investigated

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thoroughly.

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Up to now, it has been well known that deodorization temperature and time, type of oil, as

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well as oil preparation method have significant effects on the formation of GEs.9

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Furthermore, diacylglycerols (DAGs) and monoacylglycerols (MAGs) have been identified

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as the precursors of GE formation.9 Recently, our research group has published a

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comprehensive review of the formation, occurrence, analysis, and elimination method of

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GEs in refined edible oils.10 To sum up, there are four proposed reaction pathways for GE

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formation derived from DAGs and MAGs, all involving the intramolecular rearrangement via

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charge migration and differing by means of either the nature of the intermediate or the

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leaving group. Two of the proposed mechanisms involve a common reactive intermediate,

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that is, cyclic acyloxonium ion, formed by either deacidification of 1,2-DAGs or dehydration

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of MAGs (either 1-MAGs or 2-MAGs). The other 2 pathways consider a direct

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intramolecular rearrangement followed by elimination of fatty acid for DAGs (either

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1,2-DAGs or 1,3-DAGs) or of water for 1-MAGs. Our previous study confirmed the

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presence of cyclic acyloxonium intermediate (CAI) during GE formation at high temperature

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using attenuated total reflectance Fourier transform infrared spectroscopy (ATR‒FTIR),

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which was also earlier conducted to investigate the formation of CAI during 3-MCPD ester

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formation.11 Afterward, a novel free radical-mediated mechanism for 3-MCPD ester

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formation from DAGs,12 triacylglycerols (TAGs),13 and MAGs14 was proposed, which was

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involved in the formation of cyclic acyloxonium free radical intermediates but not cyclic

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acyloxonium cations. Also, based on that, mitigating 3-MCPD esters formation by

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antioxidant and radical scavenger was also investigated in detail.15,16 However, the issue

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whether the free radical-mediated mechanisms are as well responsible for the formation of

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GEs remains still unclear.

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Lipid oxidation is one of the vital reactions resulting in the quality deterioration of edible

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oil and oil-based food and commonly occurs resulting from the oxidative cleavage of an

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electron-rich double bond in unsaturated fatty acid (UFAs). It has been well known that lipid

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oxidation involves many types of free radicals which are possible to participate in the

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formation of CAI and even influence GE formation. Pro-oxidant and antioxidant can

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significantly affect the lipid oxidation via the corresponding promotion and inhibition of free

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radicals. Currently, the free radical-mediated mechanisms of 3-MCPD ester formation have

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been identified before,12 but there is yet no report about the relationship between lipid

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oxidation/free radical and GE formation at high temperature.

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Therefore, our present study aimed to investigate the effect of Fe3+ and antioxidants on

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the formation of GEs and to further ascertain the free radical mediated mechanisms and the

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influencing mechanisms of lipid oxidation for GE formation in both model oil system and

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chemical model system. To exclude the interference of DAG and MAG discrepancy in

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different types of oils, model oil was selected as real oil system. A series of model

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experiments consisting of the addition of Fe2(SO4)3, tert-butylhydroquinone (TBHQ), and

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α-tocopherol (VE) were conducted at 200 °C with dipalmitin, diolein, or methyl linoleate as

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chemical models. FTIR was used to determine the cyclic acyloxonium and ester carbonyl

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groups, and quadrupole−time of flight mass spectrometer (Q‒TOF MS) was used to identify

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the presence of radical adduct captured by 5,5-dimethylpyrroline-N-oxide (DMPO).

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Materials and methods

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Materials

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Crude palm oil was a gift from Yihai Kerry (Tianjin) Investment Co., Ltd. (Tianjin, China),

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and it originates from Malaysia. Crude camellia oil was purchased from Yongxing Taiyu

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Camellia Oil Co., Ltd. (Chenzhou, China). Crude soybean oil was provided as a gift from

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China National Cereals, Oils and Foodstuffs Corporation (COFCO). Crude linseed oil was

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graciously provided by Gansu Abest Plant Oil Development Co., Ltd. (Jinchang, China).

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These crude oils were prepared by solvent extraction and free of GEs. Model palm oil

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(MPO), model camellia oil (MCO), model soybean oil (MSO), and model linseed oil (MLO)

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were

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sn-1,2-Dipalmitin (DPG, 98%), sn-1,2-diolein (DOG, ≥97%), 3-MCPD (98%), DMPO (≥

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97%), TBHQ (97%), hexadecane ( ≥ 99%), and VE ( ≥ 96%) were purchased from

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Sigma-Aldrich Co., Ltd. (St. Louis, MO). Methyl linoleate (ML, 95%) was purchased from

prepared

by

column

chromatography

technology

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the

present

study.

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Aladdin Reagent Co., Ltd. (Shanghai, China). d5-3-MCPD (99%) and phenylboronic acid

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(PBA) were bought from Wako Pure Chemical Industries, Ltd. (Osaka, Japan). Analytical

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grade sodium thiosulfate, iron(III)sulfate (Fe2(SO4)3), n-hexane, glacial acetic acid, diethyl

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ether, petroleum ether (bp 30−60 °C), methyl tertiary butyl ether (MTBE), and anhydrous

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sodium sulfate were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai,

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China). 100‒200 mesh silica gel was obtained from Qingdao BANKE Separation Materials

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Co., Ltd. (Qingdao, China). All other chemicals in this work were of analytical grade.

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Preparation of Model Oil

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The degumming, deacidification, and bleaching procedure of crude plant oils were

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conducted according to the same operation in our previous report,9 which aimed to obtain

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the refined-bleached (RB) plant oils. Subsequently, column chromatographic method was

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carried out to prepare the corresponding model oil with the absence of polar fraction. The

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solvent A for the column was the mixture of petroleum ether and diethyl ether (87:13, v/v),

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and solvent B was diethyl ether. The silica gel (about 80 g) activated by 5 % deionized

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water was loaded into a glass column fitted with a stopcock and a sintered disc with an aid

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of a slurry in solvent A.

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RB oil obtained above was treated with anhydrous sodium sulfate and activated carbon

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to remove the moisture and chlorinated substances, respectively. Subsequently, the RB oil

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was dissolved in 2 × 30 mL of solvent A and poured into the glass column. 150 mL of

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solvent A was used to elute the nonpolar fraction that was regarded as model oil in this

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study. Additionally, the polar fraction remained on the silica column was also eluted by 150

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mL of solvent B. Last, the solvent in the nonpolar or polar eluent was preliminarily removed

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by rotary evaporator (Zhengzhou Yarong Instruments Co., Ltd., Zhengzhou, China) at

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50 °C and then evaporated completely with a gentle nitrogen flow. Both fractions were

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stored in the nitrogen-filled bottle at ‒18 °C until analysis.

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Heating Experiment in Oil Model System

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A series of different levels of TBHQ and Fe2(SO4)3 were added into model oil prepared in

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Preparation of Model Oil section and mixed completely. The final concentrations were

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reached at 0.05, 0.12, and 0.18 mg/g for TBHQ, as well as 0.1, 0.2, and 0.4 mg/mL for

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Fe2(SO4)3. Alternatively, the reaction bottle (10.2 cm tall, I.D. 1.8 com) was sealed by an

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ampoule filling machine (Changsha Buyuan Pharmaceutical Machinery Co., Ltd.,

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Changsha, China) under nitrogen aeration or exposed in air. Model oil without any addition

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of TBHQ or Fe2(SO4)3 was set as control. The heating treatment was conducted at 200 °C

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under air or nitrogen. After that, the samples were cooled to room temperature and stored

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in a nitrogen-filled glass bottle at 4 °C prior to further analysis. All experiments were

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performed in triplicate.

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Heating Experiment in the Chemical Model System

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DPG and DOG, dissolved in hexadecane and prepared at a concentration of 1.0 mg/mL,

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were selected as the chemical model representing the precursors of GE formation in plant

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oil. The antioxidants (TBHQ and VE) were prepared at 0.005 mol/L in hexadecane. 2 mg

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aliquot of pro-oxidant (Fe2(SO4)3) or 1.0 mL of antioxidant solution, as well as 4 mL of pure

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ML, were selectively added into chemical model system to investigate their effect on GE

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formation. The formulas of model mixtures were exhibited in Table 1. After being blended

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thoroughly by vortexing, the mixture was heated at 200 °C for 1 h in a drying oven

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(Shanghai Keelrein Scientific Instruments Co., Ltd., Shanghai, China), and then stored in a

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nitrogen-filled glass bottle at 4 °C prior to GE analysis. All experiments were performed in

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triplicate.

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Analytical Methods. Peroxide value (POV). POV was determined according to a modified

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iodometric method.17 Briefly, 5 g of oil sample was dissolved completely in 50 mL of solvent

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mixture consisting of acetic acid and isooctane (3:2, v/v), in which 0.5 mL of saturated KI

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solution was then added. After 1 min at ambient temperature with an occasional shake, 30

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mL of deionized water was poured into the reaction cube, shaking vigorously to release all

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iodine from isooctane layer, and then titration was performed against 0.1 mol/L sodium

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thiosulfate using 1% starch indicator until the blue-grey color just disappeared. All samples

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were determined in triplicate unless otherwise mentioned. Results were presented as

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mequiv peroxide/kg of oil.

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p-Anisidine Value (p-AV). A measure of p-AV was done based on AOCS official Method

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Cd 18-9018 with some modifications. Briefly, 2.0 g aliquot of oil sample was weighed into a

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25 mL volumetric flask and dissolved by iso-octane, and then diluted to 25 mL. A Genesys

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10S spectrophotometer (Thermo Scientific, Waltham, MA) was used to determine the

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absorbance of this above solution (Ab) at 350 nm, against isooctane as blank.

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Subsequently, 5 mL of oil-isooctane solution was mixed with 1 mL 2.5 g/L p-anisidine

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reagent. After 8 min at 25 °C, the absorbance of this reaction solution (As) at 350 nm was

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measured, and the mixture containing 5 mL isooctane and 1 mL p-anisidine reagent was

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used as blank. p-AV was calculated based on the following formula: p-AV=25 × (1.2As‒

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Ab)/m, where As and Ab denote the absorbance of the reaction mixture and oil-isooctane

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solution, respectively, and m is the mass of oil. Additionally, according to the previous

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report,19 total oxidation value (TOTOX) was calculated as (2 × POV) + p-AV. All measures

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were performed in triplicate.

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Total Polar Component (TPC). The amount of TPC was determined by a gravimetric

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method. The polar fraction was isolated from model oil by column chromatography

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separation, as described in Preparation of Model oil section. After removing the elution

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solvent, TPC was then dried in an oven at 105 ± 2 °C and weighed accurately. All

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measures were performed in triplicate. TPC level was presented as the percentage of

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model oil.

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TPC composition. High performance size exclusion chromatography (HPSEC) was

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utilized to analyze the composition of TPC based on the method reported by Cao et al.20

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with a minor adjustment of sample preparation. TPC sample was prepared based on the

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procedure in Preparation of Model Oil and Total Polar Component (TPC) sections. Then,

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this sample was dissolved in a mixture of n-hexane and isopropanol (10:1, v/v) to a

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concentration of 10 mg/mL. After being filtered through a 0.45 µm nylon membrane, this

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solution was transferred into the 1.5 mL test tubes and stored at ‒18 °C until further

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analysis. TPCs, consisting of oxidized triacylglycerols (oxTAGs), triacylglycerol polymers

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(TGPs), triacylglycerol dimers (TGDs), DAGs, MAGs, and free fatty acids (FFAs), were

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determined by high-performance liquid chromatography using Waters 2695 system

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equipped with a refractive index detector and two tandem Ultrastyragel columns (300 mm ×

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4.6 mm, 5 µm; 100 and 500 Å), connected to a guard column (50 mm × 4.6 mm) (Waters

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Co., Milford, MA). A 10 µL aliquot of TPC sample was injected and eluted with

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tetrahydrofuran at a flow rate of 0.7 mL/min at the column temperature of 35 °C. oxTAGs,

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TGPs, TGDs, DAGs, MAGs, and FFAs in oil samples were quantified by area normalization

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method. All measures were performed in triplicate. The results were presented as the

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percentage of model oil.

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GE analysis. An indirect method for GE determination in oil sample using gas

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chromatography−mass spectrometry (GC‒MS) was developed base on the AOCS Official

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Method (AOCS Cd 29c-13)21 and original DGF method C-III 18 (09)22 with some

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modifications. Briefly, two 100 mg of oil samples were weighed into test tubes containing

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100 µL MTBE and 0.5 g d5-3-MCPD, which was labeled as assay A and assay B. Then,

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200 µL of 25 g/L sodium methoxide-methanol solution was added into the test tubes to

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liberate free 3-MCPD and glycidol from their bound forms, which is stop by the addition of

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600 µL of 200 g/L acidic sodium chloride solution (assay A) and 600 µL of 600 g/L acidic

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sodium bromide solution (assay B), respectively. After 600 µL n-hexane was added into

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both tubes and kept at room temperature with vigorous shaking for about 5 min, both tubes

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were centrifuged at 4000 r/min for 5 min. Subsequently, 200 µL of saturated PBA-diethyl

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ether solution was added to both obtained organic phase above and stood at 30 °C for 20

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min. these both assays were dried using a soft nitrogen flow and then reconstituted in 500

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µL isooctane, which was stored at ‒18 °C until GC-MS analysis.

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3-MCPD derivatives were separated and quantified in a Shimadzu QP2010Plus GC-MS

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system (Shimadzu, Tokio, Japan) equipped with a 30 m × 0.25 mm × 0.25 µm Agilent

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TG-5MS column. 1 µL of sample was injected in a splitless mode with high purity helium as

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the carrier gas at 1.18 mL/min. The oven temperature program and EI-MS conditions were

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depicted as our previous report.9 The quantitation of GEs was conducted according to the

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original DGF method C-III 18 (09), GE level was calculated as (Assay A ‒ Assay B) × 0.67.

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All measures were conducted in triplicate.

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Determination of Cyclic Acyloxonium and Ester Carbonyl Groups by FTIR. The

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variation of cyclic acyloxonium and ester carbonyl groups was monitored using ATR-FTIR.

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The mixtures of DPG with Fe2(SO4)3 and VE, respectively, as well as single DPG, were

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heated at 200 °C for 15 min. After that, Fe2(SO4)3 or VE was removed by solvent extraction

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(n-hexane and methanol, respectively), centrifugation, and nitrogen blowing. The dried

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samples were separately pressed onto the KBr pellet and scanned immediately at

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wavenumbers ranging from 1500 to 1900 cm-1 in a VERTEX 70 spectrometer (Bruker

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Optics, Ettlingen, Germany). DSG at 25 °C was used as negative control. Data were

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processed using OPUS 7.2 software (Bruker Optics GmbH, Ettlingen, Germany).

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Determination of DMPO Radical Adducts by Q‒TOF MS/MS. The determination of

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DMPO radical adducts was conducted according to the previous report by Zhang et al.12

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with a minor modification. 2 mL of 1.0 mg/mL DPG in toluene was mixed with 2 mL of 5.0

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mg/mL DMPO, which was then heated at 200 °C for 15 min. Likewise, 4 mL of 0.5 mg/mL

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DPG-toluene solution was also heated at the same conditions, which was set as control.

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After both cooling immediately to room temperature in the freezing chamber, the DPG

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solution was diluted 50-fold with toluene and then injected into the Bruker Maxis Impact Q‒

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TOF mass spectrometer (Bruker Co., Bremen, Germany) combined with tandem mass

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spectrometry (MS/MS) for analysis. Electrospray ionization positive mode (ESI+) was

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selected as the LockSpray source. High purity nitrogen was used for drying (4.0 L/min), and

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nebulization (0.6 Bar). The drying heater was set as 180 °C, and the voltage of capillary

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was 3.5 kV. MS spectra were recorded at the range of 50‒1000 mass/charge (m/z) and

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processed using a Bruker Compass DataAnalysis 4.1 software (Bruker Co., Bremen,

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Germany).

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Statistical analysis. All tests were carried out in triplicate independently. Data were

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reported as mean ± standard deviation (SD). Statistical significance was measured by

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Duncan's multiple range tests using SPSS 22.0 software (IBM Corp. Armonk, NY, USA),

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and the difference between data was considered to be significant at P < 0.05.

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RESULTS AND DISCUSSION

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The relationship between GE formation and lipid oxidation and radical scavenging in

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plant oil was investigated and demonstrated in the present work. Previously, we have get a

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result that 200 °C is a turning point for the formation rate of GEs in laboratory-scale

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deodorization, and thus it was used as the heating temperature. To eliminate the

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interference of polar fractions containing the precursors of GE formation, that is, DAGs and

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MAGs, model oil without polar fractions was prepared and used as the research subject.

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Based on the fatty acid composition of plant oil that is essentially and closely linked to lipid

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oxidation, palm oil, camellia oil, soybean oil, and linseed oil were selected in this study. The

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fatty acid composition of their corresponding model oil was shown in Figure 1. The effects

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of Fe3+, TBHQ, and VE on GE formation were investigated in both model oil and chemical

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model systems via GC‒MS and FTIR. At last, in order to ascertain their influencing

244

mechanisms, the free radical in model systems was captured by DMPO and identified by

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Q‒TOF MS/MS. The results presented in this work will provide evidence for the free

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radical-mediated GE formation mechanism and show that lipid oxidation/free radical

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promotes for the formation of GEs in plant oil. Further, this work will lay a foundation for the

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development of efficient elimination methods of GEs.

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Evolution of GE Level, POV, TOTOX, and TPC Level in Plant Oil Model at High

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Temperature. It is well known that high temperature is not only largely responsible for the

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formation of GEs,23,24 but also can easily lead to lipid oxidation. According to our previous

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work,9 the experimental temperature was set at 200 °C in this study. When MPO, MCO,

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MSO, and MLO were heated directly at this temperature, GE levels were rather low and

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varied insignificantly during heating (data not shown). This observation is explained based

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on the previous finding that TAG is not the precursor of GE formation.24 Therefore, 4% DPG

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was added into the model oil to ensure the feasible formation of GEs in model oil. GE level,

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POV, TOTOX, and TPC level were monitored every 30 or 60 min during heating and the

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results were shown in Figure 2. From Figure 2A, GE levels in all tested model oils

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increased significantly with time. Interestingly, GE level in MLO was rather higher than that

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in other model oils. Based on the fact that the highest GE content was detected in palm oil

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and even palm oil-based fat.1,25-27, it was interesting to find that the formation of GEs in

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MPO was lowest in the tested model oils, which suggested the great contribution of TPC

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involving precursors to the formation of GEs. To our knowledge, this is the first report that

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when plant oil possesses the same amount of precursors, such as DAG, the type of plant

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oil seems responsible for the difference of GE levels in different tested oils. Lipid oxidation

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is one of the most common thermal reaction at high temperature, which is largely related to

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the degree of unsaturation of plant oil. It was well expected that MLO had the highest POV

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and TOTOX, followed by MSO, then MCO and MPO (Figure 2B and Figure 2C), which

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accounted for the difference of unsaturated fatty acid in tested oils (Figure 1). These results

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seemingly coincided with GE formation in tested oils (Figure 2A). However, it has also been

271

reported that conventional heating enabled the increasing DAG level in edible oil.28 In the

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present study, as shown in Figure 2D, it was indeed found that TPC contents increased

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linearly with time up to 3 h, regardless of the type of oil. this finding was in agreement with

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the results reported by Correia et al.29 Additionally, TPC content in tested oil was ranked as

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MLO > MSO > MCO > MPO, which seemed also to be positively related to GE formation.

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Accordingly, these facts raised a question: what led to the discrepancy of GE levels among

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tested oils, lipid oxidation or TPC formation?

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Evaluation of Contribution of formed TPC, Pro-Oxidant, and Antioxidant to the

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Formation of GEs in Plant Oil Model at High Temperature. As depicted above, both

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TPC content and degree of lipid oxidation seemingly increased with the same trend with GE

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formation in tested oils. To further explain this phenomenon, first, TPC, consisting of oxTAG,

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TGP, TGD, FFA, MAG, and DAG, was analyzed every hour for a period of 3 h, and the

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results were presented in Table 2. In spite of the linear increase of TPC contents in all

284

tested oils (Figure 2D), DAG and MAG levels were relatively low compared to

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oxidized/polymerized products, that is, oxTAG, TGP, and TGD, suggesting that oxidative

286

polymerization was the main chemical reaction in heated oil exposed in air, as reported by

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Choe and Min.30 Previously, Craft et al.31 have reported that GE content increases

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exponentially when DAG level exceeds 3‒4% of oil but varied slightly with below 3% of

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DAG level. Thus, DAG and MAG (altogether 0.042‒0.139% of oil) from pyrolyzation of TAG

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contributed mildly to the formation of GEs during heating. In addition, as shown in Figure 3,

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the total levels of DAG and MAG did almost not present significant difference (P > 0.05) in

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tested oils heated for the same time. These findings seemingly indicated that the

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remarkable increase of TPC content did not necessary contribute to GE formation and the

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discrepancy of GE levels in different tested oils (Figure 2A) compared with our previous

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report.9

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Subsequently, the second experiment was to confirm the effects of pro-oxidants and

297

antioxidants on the formation of GEs in four tested oils. For this sake, Fe2(SO4)3 and TBHQ

298

were added into the model oil to promote or inhibit the lipid oxidation evaluated by POV and

299

TOTOX levels (data not shown). Meanwhile, GE level was detected in these model oils

300

heated at 200 °C for 1 h. The results were shown in Figure 4. It is well known that Fe3+ can

301

initiate oil peroxidation by catalyzing the generation of alkyl radical.32,33 From Figure 4A, GE

302

levels in tested model oils heated at 200 °C for 1 h increased significantly with the addition

303

of Fe2(SO4)3 (P < 0.05). However, when EDTA-2Na, a chelating agent, together with

304

Fe2(SO4)3 was added to model oil, GE level was significantly lower than that without

305

EDTA-2Na (P VE. These results were consistent with that in the presence of ML and

355

therefore indicated that Fe3+, TBHQ, and VE might also directly acting on the formation of

356

GEs, which could involve the free radical mechanisms based on the promotion or inhibition

357

mechanisms of these substances for lipid oxidation as depicted above.

358

Monitoring of Cyclic Acyloxonium and Ester Carbonyl Groups in Chemical Model

359

System Using ATR-FTIR. As depicted above, pro-oxidants and antioxidants could

360

influence the formation of GEs not only through lipid oxidation but also through the direct

361

effect on its formation pathway (for example, accelerated or inhibited the formation of CAI)

362

in plant oil. Accordingly, to further investigate whether and how the pro-oxidant and

363

antioxidant affect the formation reaction of GEs, cyclic acyloxonium and ester carbonyl

364

groups were monitored in DPG system using ATR-FTIR. Due to the absence of ester

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365

carbonyl group in VE and without formation of cyclic acyloxonium group at high

366

temperature,16 VE was just selected as the antioxidant in FTIR test. From Figure 6, two

367

different carbonyl absorbance bands centered at 1709 and 1733 cm-1, representing two

368

different chemical environments for the ester carbonyl groups (at sn-2 and sn-1 position,

369

respectively),11 were observed in the spectra of DPG at 25 °C. Upon heating to 200 °C, just

370

a carbonyl band centered at 1736 cm-1 was observed and the absorbance band of cyclic

371

acyloxonium group centered at 1640 cm-1 appeared. When Fe2(SO4)3 and VE were

372

respectively added into two same DPG model systems and heated at 200 °C for 1 h.

373

Compared with the bands of carbonyl band and cyclic acyloxonium group of heated DPG

374

system without any addition, for the former, the carbonyl band at 1736 cm-1 was weaker but

375

the band of cyclic acyloxonium group was stronger; for the latter, on the contrary, DPG

376

system heated together with VE still exhibited two carbonyl peaks and stronger one at 1733

377

cm-1 as well as weaker band of cyclic acyloxonium group. These findings suggested that

378

Fe3+ could accelerate the formation of cyclic acyloxonium intermediate, which was

379

nevertheless slowed by VE. Previously, it has been reported that free radical, determined

380

using electron spin resonance (ESR) spectrometry, was formed in sn-1,2-stearoylglycerol

381

(DSG) system during heat exposure and a higher temperature seemingly resulted in an

382

increasing ESR signal.12, 16 As a consequence, these observations led to an interesting

383

hypothesis that the formation of GEs involved free radical-mediated mechanism, and more

384

likely, occurred via a free radical intermediate.

385

Monitoring of DMPO Radical Adducts in Chemical Model Using Q‒TOF MS/MS. To

386

confirm the occurrence of a free radical intermediate during GE formation at high

387

temperature, Q‒TOF MS and MS/MS were carried out to determine the DMPO radical

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adducts. The mixtures of DPG and DMPO heated at 200 °C for 15 min were injected into

389

Q‒TOF MS/MS system. The MS and MS/MS spectra were recorded at the range of m/z

390

100‒800 and shown in Figure 7 and Figure 8, respectively. As exhibited in Figure 7A, m/z

391

314.5035 and 553.9145, attributed to [CAI’]+ (calcd 314.5017) and [CAI’’]+ (calcd 553.9172),

392

respectively, shown in Figure 9A, was observed in the heated DPG system without DMPO,

393

suggesting the formation of two different CAI. Glycidyl palmitate (PGE) (m/z 312.4838,

394

calcd 312.4870) was also detected, which was consistent with FTIR measurement.9

395

Analysis of the mixture of DPG and DMPO under the same conditions as above found two

396

pairs of sodiated adduct ions, that is, at m/z 466.6428 most likely attributed to [CAI’ +

397

DMPO + O + Na]+ (calcd 466.6489) and m/z 706.0681 for [CAI’’ + DMPO + O + Na]+ (calcd

398

706.0644), as well as their corresponding hydrogenated ions (Figure 7B). However, neither

399

[CAI’]+ nor [CAI’’]+ were observed in Figure 7B due to their complete reaction with DMPO,

400

suggesting that the CAI existed in a free radical form, readily captured by DMPO. In

401

addition, [PGE]+ was detected in the heated mixture of DPG and DMPO, which indicated

402

the absence of GE radical intermediate formation at high temperature. This observation

403

was not consistent with the speculative description of Zhang et al.13 that GEs might be

404

formed through epoxide ring free radical intermediate. These findings provided support for

405

that the DMPO radical adducts were involved in the formation of GEs derived from DPG in

406

this model system at high temperature.

407

To further confirm the formation of DMPO radical adducts, Q‒TOF MS/MS of the

408

selected parent ion at m/z 706.0681 was also conducted. From Figure 8, the fragment ions

409

at m/z 553.9152 corresponding to [CAI’’]+, as well as m/z 685.0949 and 686.0984

410

consistent with [CAI’’ + DMPO + O + H]+ (calcd 685.0904) and [CAI’’ + DMPO + OH + H]+

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(calcd 685.0983), respectively, were detected upon collisionally activated dissociation of

412

parent oil. Furthermore, [DMPO + Na]+ and [DMPO + O + Na]+ were also found in the

413

fragment ions, which suggested that the parent ion at m/z 706.0681 was indeed the

414

sodiated adduct of CAI and DMPO with one oxygen atom bound. Hence, these above data

415

adequately demonstrated that GEs were formed through the free radical intermediate at

416

high temperature.

417

Proposed Influence Mechanism of Lipid oxidation on the Formation of GEs at High

418

Temperature. According to the above data, a free radical-mediated mechanism for the

419

formation of GEs from DAGs at high temperature was demonstrated in the present study

420

(Figure 9A). The reaction process involved in the formation of cyclic acyloxonium free

421

radical intermediates (CAI’ and CAI’’), which might be initiated by the removal of either fatty

422

acid radical (R•) at the sn-2 position or hydroxyl radical at the sn-3 position combining with

423

the thermally resulting radical (such as ROO•) that most likely originated from oil

424

peroxidation, or the alternatively direct fracture of covalent bond under high-temperature

425

condition. The ester carbonyl group at the sn-1 position might be immediately attacked by

426

the sn-2 carbon-centered radical to form CAI’ free radical. Also, the ester carbonyl group at

427

the sn-2 position might also be attacked by the sn-3 carbon-centered radical to form CAI’’

428

free radical. As shown in Figure 9A, subsequently, CAI free radical could perform a further

429

intramolecular rearrangement to form GEs in which the free radical might participate in the

430

termination stage of oil autoxidation.36 As a consequence, the generated free radical during

431

lipid oxidation could be in favor of the formation of cyclic acyloxonium free radical

432

intermediates, which explained why lipid oxidation promoted GE formation (Figure 9B).

433

Thus, the effects of Fe3+, TBHQ, and VE on the formation of GEs, as shown above, was well

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434

understood. More importantly, Fe3+ facilitated the formation of free radical, and therefore

435

also positively (+) affected CAI formation. On contrary, antioxidant, such as TBHQ, VE, et

436

al., could negatively (‒) affect CAI formation by scavenging free radical. In brief, the

437

relationship between GE formation and lipid oxidation was exhibited in Figure 9B.

438

In summary, the significant effect of Fe3+, TBHQ, and VE on GE formation at high

439

temperature was demonstrated in both model oil system and chemical model system. The

440

formation of GEs involved the free radical-mediated mechanisms and the formation of

441

cyclic acyloxonium free radical intermediates was confirmed, which explained why the

442

radical-generated lipid oxidation was in favor of GE formation. Therefore, it was well

443

understood that Fe3+ promoted the formation of GEs, which was also inhibited by TBHQ

444

and VE, by not only indirectly regulating lipid oxidation/free radical, but also by directly

445

affecting CAI formation. These results will enlighten the development of effective

446

elimination and inhibition methods for GE formation in the production of refined edible oil.

447

AUTHOR INFORMATION

448

Corresponding Author

449

*Phone: +86 20 87113875. Fax: +86 20 87114262. E-mail: [email protected].

450

ORCIDID

451

Guoqin Liu: 0000-0001-9513-1115

452

Funding

453

The work was supported by the National Hi-tech Research and Development Project of

454

China (Project No. 2013AA102103), the National Key Research and Development Program

455

of China (Project No. 2016YFD0400401‒5), the National Natural Science Fund of China

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(Project No. 31471677), the Public Welfare (Agriculture) Research Project (Project No.

457

201303072), and the Fundamental Research Funds for the Central Universities of China.

458

Notes

459 460 461

The authors declare no competing financial interest.

ABBREVIATIONS USED GEs, glycidyl esters; 3-MCPD, 3-chloropropane-1,2-diol; MPO, model palm oil; MCO,

462

model camellia oil; MSO, model soybean oil; MLO, model linseed oil; DAGs, diacylglycerols;

463

MAGs, monoacylglycerols; CAI, cyclic acyloxonium intermediate; TPC, total polar

464

component; FFAs, free fatty acids; PGE, glycidyl palmitate; PBA, phenylboronic acid;

465

TBHQ, tert-butylhydroquinone; VE, α-tocopherol; DMPO, 5,5-dimethylpyrroline-N-oxide;

466

oxTAGs, oxidized triacylglycerols; TGPs, triacylglycerol polymers, TGDs, triacylglycerol

467

dimers; POV, peroxide value; p-AV, p-anisidine value; TOTOX, total oxidation value; SFA,

468

saturated fatty acid; MUFA, monounsaturated fatty acid; PUFA, polyunsaturated fatty acid;

469

ATR‒FTIR, attenuated total reflectance Fourier transform Infrared; GC‒MS, gas

470

chromatography-mass

471

chromatography; Q‒TOF, quadrupole−time of flight; ESI, electrospray ionization. DGF,

472

German Society for Fat Science; AOCS, American Oil Chemists’ Society.

spectrometry;

HPSEC,

high

473

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performance

size

exclusion

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References (1) Weißhaar, R.; Perz, R. Fatty acid esters of glycidol in refined fats and oils. Eur. J. Lipid Sci. Technol. 2010, 112, 158–165.

477

(2) Wong, Y. H.; Muhamad, H.; Abas, F.; Lai, O. M.; Nyam, K. L.; Tan, C. P. Effects of temperature

478

and NaCl on the formation of 3-MCPD esters and glycidyl esters in refined, bleached and

479

deodorized palm olein during deep-fat frying of potato chips. Food Chem. 2017, 219, 126–130.

480 481

(3) MacMahon, S.; Begley, T. H.; Diachenko, G. W. Occurrence of 3-MCPD and glycidyl esters in edible oils in the United States. Food Addit Contam A 2013, 30, 2081–2092.

482

(4) Pudel, F.; Benecke, P.; Fehling, P.; Freudenstein, A.; Matthäus, B.; Schwaf, A. On the

483

necessity of edible oil refining and possible sources of 3-MCPD and glycidyl esters. Eur. J. Lipid Sci.

484

Technol. 2011, 113, 368–373.

485 486 487 488

(5) Inagaki, R.; Hirai, C. Formation of Glycidol Fatty Acid Esters in Meat Samples Cooked by Various Methods. J Food Process Technol 2016, 7, 557–562. (6) Wöhrlin, F.; Fry, H.; Lahrssen-Wiederholt, M.; Preiß-Weigert, A. Occurrence of fatty acid esters of 3-MCPD, 2-MCPD and glycidol in infant formula. Food Addit. Contam. A. 2015, 32, 1810–1822.

489

(7) Program, N. T. NTP Toxicology and Carcinogenesis Studies of Glycidol (CAS No. 556-52-5) In

490

F344/N Rats and B6C3F1 Mice (Gavage Studies). National Toxicology Program technical report

491

series 1990, 374, 1–229.

492

(8) IARC, Some industrial chemicals. IARC monographs on the evaluation of carcinogenic risk of

493

chemicals to humans. VOLUME 77. International Agency for Research on Cancer: Lyon, France,

494

2000; Vol. 77.

ACS Paragon Plus Environment

Journal of Agricultural and Food Chemistry

495

(9) Cheng, W.; Liu, G.; Liu, X. Formation of Glycidyl Fatty Acid Esters Both in Real Edible Oils

496

during Laboratory-Scale Refining and in Chemical Model during High Temperature Exposure. J.

497

Agric. Food Chem. 2016, 64, 5919–5927.

498

(10)

Cheng, W.; Liu, G.; Wang, L.; Liu, Z. Glycidyl Fatty Acid Esters in Refined Edible Oils: A

499

Review on Formation, Occurrence, Analysis, and Elimination Methods. Comprehensive Reviews in

500

Food Science and Food Safety 2017, 16, 263–281.

501

(11)

Rahn, A. K. K.; Yaylayan, V. A. Monitoring cyclic acyloxonium ion formation in palmitin

502

systems using infrared spectroscopy and isotope labelling technique. Eur. J. Lipid Sci. Technol.

503

2011, 113, 330–334.

504

(12)

Zhang, X.; Gao, B.; Qin, F.; Shi, H.; Jiang, Y.; Xu, X.; Yu, L. L. Free radical mediated

505

formation of 3-monochloropropanediol (3-MCPD) fatty acid diesters. J. Agric. Food Chem. 2013, 61,

506

2548–2555.

507

(13)

Zhang, Z.; Gao, B.; Zhang, X.; Jiang, Y.; Xu, X.; Yu, L. L. Formation of

508

3-monochloro-1,2-propanediol (3-MCPD) di- and monoesters from tristearoylglycerol (TSG) and the

509

potential catalytic effect of Fe2+ and Fe3+. J. Agric. Food Chem. 2015, 63, 1839–1848.

510

(14)

Zhao, Y.; Zhang, Y.; Zhang, Z.; Liu, J.; Wang, Y. L.; Gao, B.; Niu, Y.; Sun, X.; Yu, L.

511

Formation of 3-MCPD Fatty Acid Esters from Monostearoyl Glycerol and the Thermal Stability of

512

3-MCPD Monoesters. J. Agric. Food Chem. 2016, 64, 8918–8926.

513

(15)

Li, C.; Jia, H.; Shen, M.; Wang, Y.; Nie, S.; Chen, Y.; Zhou, Y.; Wang, Y.; Xie, M.

514

Antioxidants Inhibit Formation of 3-Monochloropropane-1,2-diol Esters in Model Reactions. J. Agric.

515

Food Chem. 2015, 63, 9850–9854.

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Page 24 of 41

Page 25 of 41

516

Journal of Agricultural and Food Chemistry

(16)

Zhang, H.; Jin, P.; Zhang, M.; Cheong, L. Z.; Hu, P.; Zhao, Y.; Yu, L.; Wang, Y.; Jiang, Y.;

517

Xu, X. Mitigation of 3-Monochloro-1,2-propanediol Ester Formation by Radical Scavengers. J. Agric.

518

Food Chem. 2016, 64, 5887–5892.

519

(17)

AOCS, AOCS Official Method Cd 8b-90. Peroxide Value, Acetic Acid-Isooctane Method. In

520

Official Methods and Recommended Practices of the American Oil Chemists’ Society, AOCS Press:

521

Champaign, IL: 1996.

522

(18)

AOCS, AOCS Official Method Cd 18-90. p-Anisidine Value. In Official Methods and

523

Recommended Practices of the American Oil Chemists’ Society, AOCS Press: Champaign, IL:

524

1996.

525

(19)

526 527

Shahidi, F.; Wanasundara, U. Methods for measuring oxidative stability in edible oils. Food

Lipids: Chemistry, Nutrition and Biotechnology, CRC Press: New York. 2008, 387–388. (20)

Cao, W. M.; Zhang, K. Y.; Xue, B.; Chen, F. X.; Jin, Q. Z.; Wang, X. G. Determination of

528

oxidized triacylglycerol polymers by preparative flash chromatography and high-performance

529

size-exclusion Chromatography. Asian J. Chem. 2013, 25, 9189–9194.

530

(21)

AOCS/JOCS,

Joint

AOCS/JOCS

Official

Method

Cd

29c-13:

Fatty-acid-bound

531

3-chloropropane-1,2,diol (3-MCPD) and 2,3-epoxi-propane-1-ol (glycidol), Determination in Oils and

532

Fats by GC/MS (Differential Measurement). Official Methods and Recommended Practices of the

533

AOCS, 3rd printing, 2013–2014. Additions and Revisions 6th ed.. Urbana, IL: AOCS 2013.

534

(22)

DGF In Determination of ester-bound 3-chloropropane-1,2-diol (3-MCPD esters) and

535

3-MCPD forming substances in fats and oils by means of GC-MS, Deutsche Einheitsmethoden zur

536

Untersuchung von Fetten, Stuttgart, Germany, 2009; Fettprodukten, Ed. Tensiden und verwandten

537

Stoffen: Stuttgart, Germany, 2009.

ACS Paragon Plus Environment

Journal of Agricultural and Food Chemistry

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(23)

Page 26 of 41

Shimizu, M.; Weitkamp, P.; Vosmann, K.; Matthäus, B. Temperature dependency when

generating glycidyl and 3-MCPD esters from diolein. J. Am. Oil Chem. Soc. 2013, 90, 1449–1454. (24)

Destaillats, F.; Craft, B. D.; Dubois, M.; Nagy, K. Glycidyl esters in refined palm (Elaeis

541

guineensis) oil and related fractions. Part I: Formation mechanism. Food Chem. 2012, 131, 1391–

542

1398.

543

(25)

Becalski,

A.;

Feng,

S.;

Lau,

B.

P.;

Zhao,

T.

A

pilot

survey

of

2-and

544

3-monochloropropanediol and glycidol fatty acid esters in foods on the Canadian market 2011–

545

2013. J. Food Compos. Anal. 2015, 37, 58–66.

546

(26)

Vasilios, G.; Samaras, A. G.; Zuzana Zelinkova; Karasek, L.; Buttinger, G.; Wenzl, T.

547

Analytical method for the trace determination of esterified 3- and 2-monochloropropanediol and

548

glycidyl fatty acid esters in various food matrices. J. Chromatogr. A 2016, 1466, 136–147.

549

(27)

Marc, C.; Drouard-Pascarel, V.; Retho, C.; Janvion, P.; Saltron, F. Determination of MCPD

550

Esters and Glycidyl Esters by Microwave Extraction in different Foodstuffs. J. Agric. Food Chem.

551

2016, 64, 4353–4361.

552

(28)

Lucas-Torres, C.; Pérez, Á.; Cabañas, B.; Moreno, A. Study by

31

P NMR spectroscopy of

553

the triacylglycerol degradation processes in olive oil with different heat-transfer mechanisms. Food

554

Chem. 2014, 165, 21–28.

555

(29)

Correia, A. C.; Dubreucq, E.; Ferreira-Dias, S.; Lecomte, J. Rapid quantification of polar

556

compounds in thermo-oxidized oils by HPTLC‐densitometry. Eur. J. Lipid Sci. Technol. 2015, 117,

557

311–319.

558

(30)

Choe, E.; Min, D. Chemistry of deep-fat frying oils. J. Food Sci. 2007, 72, R77–R86.

ACS Paragon Plus Environment

Page 27 of 41

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Journal of Agricultural and Food Chemistry

(31)

Craft, B. D.; Nagy, K.; Seefelder, W.; Dubois, M.; Destaillats, F. Glycidyl esters in refined

560

palm (Elaeis guineensis) oil and related fractions. Part II: Practical recommendations for effective

561

mitigation. Food Chem. 2012, 132, 73–79.

562 563 564

(32)

Schafer, F. Q.; Qian, S. Y.; Buettner, G. R. Iron and free radical oxidations in cell

membranes. Cellular and molecular biology (Noisy-le-Grand, France) 2000, 46, 657–662. (33)

Chen, B.; Mcclements, D. J.; Decker, E. A. Minor components in food oils: a critical review

565

of their roles on lipid oxidation chemistry in bulk oils and emulsions. Crit. Rev. Food Sci. Nutr. 2011,

566

51, 901–916.

567

(34)

Berdeaux, O.; Fontagné, S.; Sémon, E.; Velasco, J.; Sébédio, J. L.; Dobarganes, C. A

568

detailed identification study on high-temperature degradation products of oleic and linoleic acid

569

methyl esters by GC–MS and GC–FTIR. Chem. Phys. Lipids 2012, 165, 338–347.

570

(35)

Alamed, J.; Chaiyasit, W.; Mcclements, D. J.; Decker, E. A. Relationships between free

571

radical scavenging and antioxidant activity in foods. Journal of Agricultural & Food Chemistry 2009,

572

57, 2969–2976.

573 574

(36)

St. Angelo, A. J., Vercellotti, J.; Jacks, T.; Legendre, M. Lipid oxidation in foods. Crit. Rev.

Food Sci. Nutr. 1996, 36, 175–224.

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Figure captions

577

Figure 1. Fatty acid composition of model palm oil (MPO), model camellia oil (MCO) ,

578

model soybean oil (MSO), and model linseed oil (LO). SFA, saturated fatty acid; MUFA,

579

monounsaturated fatty acid; PUFA: polyunsaturated fatty acid.

580

Figure 2. Evolution of GE levels (A), POV (B), and TOTOX (C), as a function of time (0.5‒

581

3.0 h) in tested model oils with the addition of 4% DPG heated at 200 °C, and evolution of

582

TPC contents (D) in tested model oils heated at 200 °C. Data are mean ± standard

583

deviation (SD) (n=3).

584

Figure 3. Changes of total contents of DAGs and MAGs in MPO, MCO, MSO, and MLO

585

during heating at 200 °C up to 3 h. All data are mean ± SD (n=3). Different letters at the

586

same time represents significant difference between total levels of DAG and MAG in

587

different types of model oil (P < 0.05)

588

Figure 4. Effects of Fe2(SO4)3 (A), TBHQ (B), isolating oxygen (C) on GE formation in

589

tested model oils with addition of 4% DPG heated at 200 °C for 1 h. aequimolar quantities of

590

Fe3+ and EDTA-2Na were added into model oils. All data are mean ± SD (n=3). Values

591

labeled by a different letter are significantly different for the same model oil (P < 0.05).

592

Figure 5. GE levels in different chemical model systems heated at 200 °C for 1 h. All data

593

are mean ± SD (n=3). Values labeled by different letters are significantly different (P