high resolution-time of flight mass spectrometry

Accepted Manuscript Analysis of volatiles in Dezhou Braised Chicken by comprehensive two-dimensional gas chromatography / high resolution-time of flight mass spectrometry Yan Duan, Fuping Zheng, Haitao Chen, Mingquan Huang, Jianchun Xie, Feng Chen, Baoguo Sun PII:

S0023-6438(14)00561-1

DOI:

10.1016/j.lwt.2014.09.006

Reference:

YFSTL 4137

To appear in:

LWT - Food Science and Technology

Received Date: 17 June 2013 Revised Date:

30 June 2014

Accepted Date: 1 September 2014

Please cite this article as: Duan, Y., Zheng, F., Chen, H., Huang, M., Xie, J., Chen, F., Sun, B., Analysis of volatiles in Dezhou Braised Chicken by comprehensive two-dimensional gas chromatography / high resolution-time of flight mass spectrometry, LWT - Food Science and Technology (2014), doi: 10.1016/ j.lwt.2014.09.006. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

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Analysis of volatiles in Dezhou Braised Chicken by

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comprehensive two-dimensional gas chromatography /

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high resolution-time of flight mass spectrometry1

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Yan Duan1, Fuping Zheng1, 2, ∗, Haitao Chen1,2, Mingquan Huang1, 2, Jianchun Xie1,2, Feng Chen3,

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Baoguo Sun1,2

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1. Beijing Key Laboratory of Flavor Chemistry, Beijing Technology and Business University, 33

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Fucheng Road, Beijing 100048, PR China.

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2. Beijing Higher Institution Engineering Research Center of Food Additives and Ingredients,

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Beijing Technology and Business University, 33 Fucheng Road, Beijing 100048, PR China.

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3. Department of Food, Nutrition and Packaging Sciences, Clemson University, Clemson, South

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Carolina 29634, USA.

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ABSTRACT: A fast and mild extraction method and an accurate identification method to analyze

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the volatiles in non-volatile food matrices were set up. The accelerated solvent extraction

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1 The manuscript was presented in the international conference of “Food Innova-2012” Hangzhou, China, December 12-14, 2012. ∗ Corresponding author. Tel.: +86 10 6898 4857; Fax: +86 10 6898 4857 E-mail address: [email protected] (Fuping Zheng) 1

ACCEPTED MANUSCRIPT followed by solvent assisted flavor evaporation (ASE-SAFE) was used to extract the volatiles

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from Dezhou braised chicken. The extract was analyzed by comprehensive two-dimensional gas

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chromatography / high resolution-time of flight mass spectrometry (GC×GC/HR-TOFMS) and

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GC-qMS. Ninety one compounds were identified by GC×GC/HR-TOFMS with an aid of NIST08

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library and accurate mass determination, 44 compounds were identified by GC-qMS only with

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NIST08 library. The main odour-active constitutes of the Dezhou braised chicken were carbonyl

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compounds (33.04%). 2-Enals and 2,4-dineals were considered to be the most important

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odour-active constitutes in chicken. In our study, 6 very polar and unstable 2-enals or 2,4-dineals

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compounds,

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(E,E)-2,4-heptadienal and (E,E)-2,4-nonadienal, were only identified by GC×GC/HR-TOFMS.

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Three very polar and unstable furanones, including 5-acetyldihydro-2(3H)-furanone (0.04%),

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dihydro-3-hydroxy-4,4-dimethyl-2(3H)-furanone

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2,5-dimethyl-4-hydroxy-3(2H)-furanone

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GC×GC/HR-TOFMS. Some trace sulfur-containing and nitrogen-containing compounds,

(E)-2-hexenal,

(E)-2-nonenal,

(Z)-2-decenal,

(E)-2-undecenal,

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including

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(0.32%)

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(0.32%)

were

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only

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detected

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ACCEPTED MANUSCRIPT including disulfide dipropyl (0.05%), 2-acetylthiazole (0.06%), 2-pyrrolidinone (0.15%) and

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benzothiazole (0.07%) that might play important roles in the flavor of chicken, were also only

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found by GC×GC/HR-TOFMS.

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Key words: ASE-SAFE; GC×GC/HR-TOFMS; GC-MS; Dezhou Braised Chicken; Volatiles

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1. Introduction

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Dezhou Braised Chicken is a famous traditional dish in China, having the

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reputation of “Top 1 Braised Chicken in China” due to its attractive appearance and

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flavor. It originated from Dezhou, Shandong Province, China, in the era of Guangxu

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of the Qing Dynasty, and popularized for over 300 years. It’s also known as

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“Boneless Braised Chicken” because the bones can be easily separated from the

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chicken after cooking. The cooking procedure of Dezhou Braised Chicken is enrolled

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in China Intangible Cultural Heritage. The common cooking procedure is as follows:

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the chicken is painted with honey after shaping, then deep-fried for several minutes

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till its surface become golden yellow, finally boiled for 6-10 hours with soy sauce,

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ACCEPTED MANUSCRIPT sugar, salt and tens of spices, such as Chinese prickly ash, star anise, Chinese

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cinnamon bark, clove, ginger, fennel, etc. After cooking, the braised chicken is in

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lustrous golden yellow, tender and rich in flavor. The delicious taste and flavor

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usually consist of numerous non-volatile compounds, as well as many volatile

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compounds plus trace amounts of aroma compounds. To date, over 1000 volatile

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compounds have been identified in meat. These volatiles include alcohols, acids,

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esters, hydrocarbons, ethers, lactones, heterocyclic compounds and sulfur compounds

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(Shahidi, 1994). Aldehydes especially the 2-enals, 2,4-dineals, ketones, heterocyclic

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and sulfur compounds, are believed to play an important role in the flavor of cooked

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chicken (Gasser, & Grosch, 1989).

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Volatiles in chicken have been analyzed by solid-phase micro-extraction (SPME)

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(Dimitris, Nigel, Gerard, & Amalia, 2012), simultaneous distillation extraction (SDE)

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(Watkins, Roser, Warner, Dunshea, & Pethick, 2012), etc. SPME can only be used to

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analyze the highly volatile compounds because only the head-space compounds are

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ACCEPTED MANUSCRIPT extracted. Although the SDE method is a very effective and universal method for

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analyzing aroma compounds, a sample needs to be heated for a very long time to

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extract the flavors, which can be harmful to thermally sensitive components. Another

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disadvantage of SDE is that it usually needs a large amount of sample. Regarding

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these shortcomings, Richer et al (Richer, Ezzell, Porter, Avdalovic, & Pohl, 1995)

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introduced a new method called accelerated solvent extraction (ASE) that has been

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adopted in recently years. Under high pressure and high temperature, the organic

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solvents are able to more efficiently extract the analytes in environmental samples (Li,

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Landriault, Fingas, & Liompart, 2003), medicines (Yu, You, Wu, Wang, & Zhao,

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2010), food (Toschi, Bendini, Ricci, & Lercker, 2003) et al. This method is suitable

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for solid and/or semisolid samples, of which the target analytes are trapped in the

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cells during extraction. However, a problem of ASE is that the fat in a sample (e.g.

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chicken) may be extracted into solvent, which will cause a big problem for GC-MS

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analysis. Fortunately, we found that ASE combined with solvent-assisted flavor

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ACCEPTED MANUSCRIPT evaporation (SAFE) could resolve this problem. The SAFE is a new technique for

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aroma extraction and has been widely used to isolate aroma compounds from

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complex matrices. In high vacuum(5×10-3Pa), SAFE allows the isolation of volatiles

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from either solvent extracts, aqueous foods, such as milk or beer, aqueous food

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suspensions, such as fruit pulps, or even matrices with solvent extraction (Engel, Bahr,

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& Schieberle, 1999). In this way, volatiles and non-volatiles were separated. ASE

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combined with SAFE could not only achieve an enhanced extraction from solid

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samples, but also remove non-volatile compounds from the extraction solution.

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To identify the volatiles in chicken, both an efficient extraction method and a

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powerful analytical method are important. Even though GC-MS is commonly used to

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analyze volatiles, it’s not appropriate for profile analysis because it is very difficult to

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select one GC column suitable for all compounds, and the traditional software only

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focuses on structural analysis and not on the profile data processing. In this context

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we adopted a new technology that not only had good resolution for all compounds,

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ACCEPTED MANUSCRIPT but also maintained desirable sensitivity. The two-dimensional GC (GC×GC) can

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integrate two GC columns with different polarity into one injection port in order to

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separate polar or nonpolar compounds. The high resolution-time of flight mass

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spectrometer (HR-TOFMS) was used for this flavor analysis because of its very fast

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data collection, full mass sensitivity and high resolution that is capable of obtaining

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accurate mass values. It is a perfect tool for chemical quantitation and qualification.

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In addition a new data processing software (GC-image V2.3) can be used to explore

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the spectra in 3D.The 3D mode is very useful for chemical profile analysis.

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Comprehensive two-dimensional gas chromatography is a brand-new technique

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developed in the 1990s for the separation of complex mixtures (Wang, Weng, Zhang,

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Zhu, Chen, & Wei, 2010). The 2D GC column system has two orthometric columns

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with different polarity and distinct separation mechanisms. It depends upon a

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modulator to accomplish orthometric columns with different polarities and distinct

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separation mechanisms. It depends upon a modulator to transfer orthogonally

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ACCEPTED MANUSCRIPT between the two capillary columns. Compared to one-dimensional GC, the 2D GC

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has a better resolution, higher sensitivity, larger peak capacity, shorter analysis time

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and more well-regulated qualification. Besides the modulator, the GC×GC technique

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has another advantage relying on its time-of-flight spectrometry which has a high

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acquisition rate up to 100 full-range mass spectra/s. The GC×GC analysis is well

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suited to analyze complex matrices, such as petroleum samples (John, & Frank, 2005),

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environment samples (Kallio et al., 2003), organic pesticides (Dallüge, Rijn, Beens,

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Vreuls, & Brinkman, 2002) and essential oil (Zhu et al., 2005) etc. However, there are

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no reports cocerning the application of GC×GC/HR-TOFMS on the analysis of

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flavors in chicken meat. This research investigated a novel extraction procedure and

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an analytical method for volatile compounds in chicken meat which was a solid or

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semisolid complex mixture.

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2. Material and methods

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2.1. Material

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ACCEPTED MANUSCRIPT Dezhou Braised Chicken was purchased from the chain store of Shandong

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Dezhou Braised Chicken Co. Ltd at Beijing Chengxiang Supermarket, Beijing, China.

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Methylene dichloride(≥99.5%)[,] and sodium sulfate(≥99.5%) were supplied by

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Sinopharm Chemical Reagent Beijing Co., Ltd., Beijing, China.

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2.2. Sample preparation

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The chicken was cut into one cubic centimeter pieces, grounded by a pulverizer

in liquid nitrogen and kept at -20℃ until further analyzed.

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2.3. Extraction of flavor compounds

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2.3.1. Accelerated Solvent Extraction (ASE)

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Extraction of the chicken flavor was carried out using accelerated solvent

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extractor (ASE 100, Sunnyvale,ºCalifornia, USA) with dichloromethane as the

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extraction solvent. Thirty grams of the ground chicken meat were weighted and

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placed in the extraction cell of the instrument. Extraction was carried out at 100℃.

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After the injection of the solvent into the cell, a pressurized static extraction phase

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ACCEPTED MANUSCRIPT lasting 5 min was carried out at 1600 psi, followed by a flow of fresh

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dichloromethane equal to 120% of the cell volume. The extraction cycle was repeated

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by three times.

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2.3.2. Solvent-assisted flavor evaporation (SAFE)

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The aforementioned extraction liquid (2.3.1) was evaporated by a high-vacuum

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(5×10-3Pa) distillation to get the volatiles using the SAFE (Kimble Bomex (Beijng)

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Labware Co. Ltd., Beijing, China) at 50 ºC. After the evaporating, the solution was

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dried over anhydrous sodium sulfate. The dichloromethane with the volatiles mixtures

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was stirred for 15min by the rotary evaporator and concentrated to 0.5 mL under a

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stream of nitrogen in the end.

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2.4. GC-qMS analysis

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GC-qMS analysis of ASE-SAFE extracts was performed on an HP 6890 GC

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coupled with an HP 5973 mass spectrometer (Agilent Technologies, Palo Alto,

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California, USA). It was equipped with a DB-Wax column (Agilent) (30m length,

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250µm I.D., and0.25µm phase thickness). The sample (2 µL) was injected at 250 ºC

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in a splitless mode. The solvent delay time was set at 5 min. The initial oven

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temperature was maintained at 40 ºC for 2 min, increased to 120 ºC at 5

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increased to 150 ºC at 3 ºC /min, increased to 200 ºC at 6 ºC /min, then increased to

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250 ºC at 8 ºC /min, and then held for 1min at that temperature. The ionization

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potential of MS was 70 eV and its MS scan ranged from m/z 40-500amu. The MS

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source and transfer line temperatures were 230 and 250 ºC, respectively.

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2.5. GC×GC/HR-TOFMS analysis

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The Fas TOF GC×GC/HR-TOFMS system consisted of an Agilent 7890

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/min,

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(Agilent Technologies, Palo Alto, California, USA) gas chromatography and

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cold-jet modulator and a high resolution time-of-flight mass spectrometry (Zoex

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Corp., Nebraska, USA), which has high scan rate up to 500Hz, 4000-7000 resolution

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and high sensitivity. The data processing software is GC-image HRMS 2.3. The first

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column was DB-1 (10m×0.25mm i.d. × 0.25µm film thickness) and the second

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column was BPX-50 (1m×0.1mm i.d. × 0.1µm film thickness), both were from

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Agilent. A volume of 1µL of sample was injected into the GC injector with a split

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ratio of 10:1 at 250

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were used with the cryogenic trap cooled to -90

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separation was performed using the following temperature program: initial

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temperature 40

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C. A modulation period of 7s and the hot jet widths of 300ms

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C and held for 2 min. The transfer line into the TOF-MS source was

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heated at 280

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collision energy of 70 eV. The data acquisition rate was 100 Hz over a mass range of

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40-500 amu. The identification of volatiles of the Dezhou braised chicken was based

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on NIST08 library search, accurate mass calculation of the compound fragment ions

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and the standard compounds.

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3. Results and discussion

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3.1. GC-qMS analysis

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ACCEPTED MANUSCRIPT Fig.1 shows the total ion chromatogram on a DB-wax capillary column. Volatiles

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isolated from the Dzhou braised chicken by ASE-SAFE-GC-MS are shown in Table

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In Table 1, a total of 44 flavors were listed, which were identified by comparison

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with chemical standard, the NIST 08 database and retention index. Those compounds

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included 9 aldehydes, 3 ketones, 9 nitrogen containing, oxygen containing or

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sulfur-containing compounds, 9 alcohols, 5 acids, 1 esters, 3 ethers, 1 phenol and 4

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hydrocarbons, which have been reported in other references (He, & Xu, 2001; Xiang,

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Hou, Zhang, & Li, 2010).

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Aldehydes which are mainly from lipid oxidation, usually have low thresholds

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and often play an important role in meat flavor (Mottram, 1991). Due to the

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advantage of the ASE-SAFE method, 9 aldehydes and 3 ketones were extracted and

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identified regardless of the volatility of the aldehydes and ketones.

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3.2. GC×GC/HR-TOFMS analysis

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ACCEPTED MANUSCRIPT Fig.2 shows GC×GC image in the comprehensive 2D GC. Fig.3 shows the 3D

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image of the compounds separated by the GC×GC. X axis is the retention time of the

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compounds (dependent on carbon numbers) and the Y axis is the polarity. It is clear

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that the 2D GC has better resolution compared to the overlapping peaks in

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one-dimensional gas chromatography.

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A total of 91 volatiles were identified from the Dezhou braised chicken after

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comparison with the standard compounds, the NIST08 library, and accurate mass

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determination. Compounds with matching factors and reverse factors above 800 can

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be identified directly. Those suspected volatiles that had lower matching values than

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800 but higher than 700 were further identified by an accurate mass determination

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with the accuracy of ±0.002amu. Some compounds such as 2-enals and 2,4-dienals

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were identified by authentic standard compounds. Other compounds with low

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matching factors also possibly existed in the chicken based on the accurate mass

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determination. Table 2 also shows fragment ions of some volatiles identified by the

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accuracy mass determination.

A critical step to find the aroma-impact volatiles is to establish an appropriate

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method to extract the volatile flavor compounds from the foodstuff. As mentioned

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above, an ASE-SAFE method was used to extract the volatiles from the Dezhou

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braised chicken by the solvent dichloromethane. Macneil and Dimick (MacNeil, &

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Dimick, 1970) reported that 2-enals and 2,4-dienals played an important role in

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poultry flavor. Enals and dienals arise from the autooxidation of the appropriate

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polyunsaturated fatty acids. 2,4-Decadienal (trans, cis) is characteristic of the odour

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of cooked chicken fat and 2,4-heptadienal (trans, trans) is a major constituent of the

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volatile aldehydes isolated from chicken fat (Pippen, Nonaka, & Jones, 1958). Harkes

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and Begemann (Harkes, & Begemann, 1974) suggested that 2,6-dodecadienal (trans,

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cis) played an important role in chicken flavor via the browning reaction.

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2-Isopropyl-2-butenal and 4-methyl-2-pentenal were identified in fried chicken and

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were suggested to be formed during frying by aldol-type condensations. A total of 9

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ACCEPTED MANUSCRIPT volatile compounds in the form of either 2-enals or 2,4-dienals have been detected by

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GC×GC/HR-TOFMS (shown in Table 1). Three furanones which had low volatility

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and high polarity have been detected only by the GC×GC/HR-TOFMS, including

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5-acetyldihydro-2(3H)-furanone

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dihydr-o-3-hydroxy-4,4-dimethyl-2(3H)-furanone

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2,5-dimethyl-4-hydroxy-3(2H)- furanone (0.32%). It is well known that poultry is

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enriched in sulfur-containing or nitrogen-containing compounds. In this research, a

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low content of disulfide dipropyl (0.05%), 2- pyrrolidinone (0.15%) 2-acetylthiazole

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(0.06%) and benzothiazole (0.07%) were identified by the GC×GC/HR-TOFMS.

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3.3. Comparison of GC-qMS and GC×GC/HR-TOFMS

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As shown in Table 3, it is clear that GC×GC/HR-TOFMS is a more powerful

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tool in identifying compounds than GC-qMS. More polar compounds were identified

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by GC×GC/HR-TOFMS than non polar volatiles because a NP (nonpolarity+polarity)

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column was installed. Many volatiles that overlapped seriously in the GC-qMS

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analysis were focused and re-injected by the modulator as a sharp injection pulse into

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the second dimension GC column by the GC×GC/HR-TOFMS. Fig.4A1 shows the

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3D plot

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Fig.4B1-4C1exhibit the chromatograms and blobs of decanal and 2,4-nonadienal

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(also listed in Table 4), which can be clearly separated by GC×GC system, but not

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separated by GC-MS. Another similar case was presented in Fig.5 to further explain

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the co-eluting problems in GC-qMS, which can be resolved by GC×GC. The

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compounds of (E,E)-2,4-heptadienal and 2-acetylthiazole in Table 5 could not be

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identified in GC-qMS, but they were identified by 2D GC. These compounds are also

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important contributors to chicken flavor.

2,4-nonadienal,

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of the selective blobs

In addition to the advantages mentioned above, several other characteristics of

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the “GC×GC/HR-TOFMS advantage” are apparent. Firstly, better resolution is

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obtained in the 2D GC for those overlapped peaks in the GC-qMS. Secondly,

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GC×GC/HR-TOFMS analysis provides better mass spectra and accurate mass values

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ACCEPTED MANUSCRIPT (±0.002amu). Since it is not always possible or reliable to identify chemicals by a

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library search, the GC×GC/HR-TOFMS method was used to overcome that limitation

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by determining the accurate mass of the compounds. Furthermore, the NIST search is

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not always right when the library match factor is too low. It is worthwhile to mention

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that benzothiazole, o-anisaldehyde and 3-methyl-1,2-cyclopentanedione (Table 2) did

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not have the highest matching factors in the primary library search. Owing to the high

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resolution and accurate mass determination, these volatiles were identified. Thirdly,

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the availability of a completely automated GC×GC system, makes quick qualitative

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analyses possible. The GC can automatically conduct the baseline calibration,

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compensate for background interference and the column bleed, and complete the

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library search.

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4. Conclusions

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A new method (ASE-SAFE) has been developed to extract and isolate the

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volatile compounds from complex foodstuff and then to analyze them by

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ACCEPTED MANUSCRIPT GC×GC/HR-TOFMS. ASE was shown to be more powerful and efficient than a direct

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solvent extraction in terms of the extraction time, extractability, solvent consumption

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and labour cost. SAFE provided an efficient and gentle method to separate the

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volatiles from the non-volatile materials, such as lipids and natural colours.

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The application of GC×GC/HR-TOFMS to detect volatiles in processed chicken

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meat was successfully achieved. This was largely due to the high resolution and

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sensitivity of the comprehensive 2D GC coupled to TOFMS. Because of its high

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resolution and power to conduct an accurate mass determination, the qualitative

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analysis of flavors has become more reliable and credible, and will facilitate the

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identification of other volatile compounds present in trace amounts. This research

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demonstrated that ASE-SAFE combined with GC×GC/HR-TOFMS has good

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potential to become an important method for flavor analysis.

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Acknowledgement

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We thank Polytech Instrument Ltd, in particularly Bin Wang for their technical

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ACCEPTED MANUSCRIPT assistance with GC×GC/HR-TOFMS. This work was supported by the National

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Nature Science Foundation of China (31171646).

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References

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SC

294

Pippen, E. L., Nonaka, M. & Jones, F. T. (1958). Volatile carbonyl compounds of cooked chicken.

299

. Compounds obtained by air entrainment. Journal of Food Science, 23(1): 103-113.

300

Richter, B. E., Jones, B. A., Ezzell, J. L., Porter, N. L., Avdalovic, N. & Pohl, C. (1995).

301

Accelerated Solvent Extraction: a Technique for Sample Preparation. Analytical Chimical

302

Acta, 68(6), 1033-1039.

EP

TE D

M AN U

298

Shahidi, F. (1994). Flavor of meat and meat products. Food Science & Technology, 28(2), 251.

304

Toschi, G. T., Bendini, A., & Ricci, A. & Lercker, G. (2003). Pressurized solvent extraction of

305

AC C

303

total lipids in poultry meat. Food chemistry, 83(4), 551-555.

306

Wang, H.-T., Weng, N., Zhang, S.-C. Zhu, G.-Y., Chen, J.-P. & Wei, C.-Y. (2010). Identification of

307

petroleum aromatic fraction by comprehensive two-dimensional gas chromatography with

22

ACCEPTED MANUSCRIPT

308

time-of-flight mass spectrometry. Chinese Science Bulletin, 55(19), 2039-2045.

Watkins, P. J., Roser, G., Warner, R. D., Dunshea, F. R. & Pethick, D. W. (2012). A comparison of

310

solid-phase micro-extraction (SPME) with simultaneous distillation-extraction (SDE) for the

311

analysis of volatile compounds in heated beef and sheep fats. Meat Science, 91(2), 99-107.

313

SC

Xiang, Q., Hou, J.-N., Zhang X. & Li, D.-P. (2010). Flavor study on Dezhou Braised Chicken by

M AN U

312

RI PT

309

SPME. MEAT RESEARCH, 11, 61-64.

Yu, H., You, H., & Wu Y.-W., Wang, Y. & Zhao, P. (2010). Determination of Residual

315

Sulfonamides in Animal Originated Food by High Performance Liquid Chromatography with

316

Accelerated Solvent Extraction. Modern Scientific Instruments, 6, 93-96.

318 319 320

EP

Zhu, S.-K., Lu, X., Xing, J., Zhang, S.-W., Kong, H.-W., Xu, G.-W. & Wu, C.-Y. (2005).

AC C

317

TE D

314

Comparison of comprehensive two-dimensional gas chromatography/time-of-flight mass

spectrometry and gas chromatography–mass spectrometry for the analysis of tobacco

essential oils. Analytical Chimical Acta, 545(2), 224-231.

23

ACCEPTED MANUSCRIPT 2200000

2000000

1800000

1600000

1400000

RI PT

1200000

1000000

800000

600000

200000

15.00

20.00

25.00

30.00

M AN U

10.00

SC

400000

-->

35.00

40.00

AC C

EP

TE D

Fig.1 Total ion chromatogram (on DB-Wax) of volatiles in Dezhou Braised Chicken

24

RI PT

ACCEPTED MANUSCRIPT

AC C

EP

TE D

M AN U

SC

Fig.2. GC×GC/TOF-MS chromatogram of volatiles from the Braised Chicken by ASE-SAFE

25

SC

RI PT

ACCEPTED MANUSCRIPT

AC C

EP

TE D

M AN U

Fig.3 The image of 3D in Dezhou Braised Chicken by GC×GC/HR-TOFMS

26

ACCEPTED MANUSCRIPT

b

b

RI PT

a

a

AC C

EP

TE D

M AN U

SC

Fig. 4 GC×GC plot corresponding to the chromatography in 2nd dimension (Ⅰ)

27

ACCEPTED MANUSCRIPT

c

c

d

d

SC

b

M AN U

b

RI PT

a

a

AC C

EP

TE D

Fig. 5 GC×GC plot corresponding to the chromatography in 2nd dimension (Ⅱ)

28

ACCEPTED MANUSCRIPT

Figure legends

(a) 2,4-nonadienal

(b) decanal

RI PT

Fig. 4 GC×GC plot corresponding to the chromatography in 2nd dimension ( ).

Fig. 5 GC×GC plot corresponding to the chromatography in 2nd dimension ( ). (c) octanal

TE D EP AC C 29

(d) 2-pentyl-furan

SC

(b) (E,E)-2,4-heptadienal

M AN U

(a) 2-acetylthiazole

ACCEPTED MANUSCRIPT Table 2 Compounds qualified by accuracy mass

2,6-dimethoxy-phenol

Mass (m/z)

Difference

154.06430*

0.001856

139.03833

0.000636

C7H7O3+

111.04558

0.001527

C6H7O2+

93.03244

-0.001054

C6H5O+

-0.001449

C7H5NS+

C8H10O3

M AN U

135.01227*

benzothiazole

C7H5NS

C8H10O3+

108.00152

0.001299

C6H4S+

68.98071

0.001367

C3HS+

136.05129*

-0.000588

C8H8O2+

77.03532

-0.003258

C6H5+

112.05038*

-0.001496

C6H8O2+

69.03285

-0.000641

C4H5O+

55.05269

-0.001535

C4H7+

41.03797

0.000610

C3H5+

150.04654*

-0.006600

C6H14S2+

108.00472

-0.001473

C3H8S2+

43.05267

-0.001560

C3H7+

C8H8O2

EP

TE D

2-methoxy-benzaldehyde

Fragment Ions

RI PT

Formula

SC

Compounds

3-methyl-1,2-Cyclopenta

C6H8O2

AC C

nedione

disulfide dipropyl

C6H14S2

“*”: molecular ion peak 30

ACCEPTED MANUSCRIPT Table 3 Kinds of components in GC-MS and GC×GC/HR-TOFMS GC-MS

GC×GC/HR-TOFMS

Aldehydes

9

21

Ketones

3

7

Nitrogen containing, Oxygen containing or

9

21

Sulfur-containing compounds

RI PT

Group

Alcohols

9

15

Acids

5

Esters

1

2

Ethers

3

3

M AN U

SC

9

Phenol Hydrocarbons

AC C

EP

TE D

Total

31

1

3

4

14

44

95

ACCEPTED MANUSCRIPT Table 4 The peaks overlapping in the first chromatography column ( )

Name

Odour

Volume Percent

fatty, floral, green, orange peel

0.13%

2,4-nonadienal

fatty, green

0.10%

(s)

1.78

AC C

EP

TE D

M AN U

SC

RI PT

decanal

Peak

32

2.27

ACCEPTED MANUSCRIPT Table 5 The p eaks overlapping in the first chromatography column ( )

Odour

Volume Percent

Peak

2-pentyl furan

fruity, green, pungent, sweet

0.15%

1.54

octanal

fatty, fruity, orange peel

0.86%

1.75

(E,E)-2,4-heptadienal

fatty, nutty,

0.06%

2.41

2-acetylthiazole

bread, burnt milk, caramel

0.06%

3.42

SC M AN U TE D EP AC C 33

(s)

RI PT

Name

ACCEPTED MANUSCRIPT

Compound Name

GC-qMS RT

Library

(min)

Match

GC×GC/HR-TOFMS

a

RI

b

RI*

Peak I

Peak II

Library Match/

Library

Percent

(min)

(s)

Reverse Match

Probability

Response

Aldehydes 3-methyl-butanal

5.74

2

Hexanal

6.55

909

1069

1064

10.25

3

Heptanal

9.09

887

1170

1182

4

Octanal

5

Nonanal

6

Decanal

7

(E)-2-hexenal

8

(Z)-2-heptenal

12.72

899

9

(E)-2-octenal

15.50

886

10

(E)-2-nonenal

11

(Z)-2-decenal

12

(E)-2-undecenal

13

(E,E)-2,4-heptadienal

14

(E,E)-2,4-nonadienal

15

(E,E)-2,4-decadienal

16

(E)-cinnamaldehyde

1378

1392

TE D

910

AC C

852/867

60.03

1.59

MS

1.5

897/897

77.08

14.02

MS

16.55

1.67

855/882

73.7

0.71

MS

23.92

1.75

815/878

67.42

0.73

MS

31.53

1.78

893/899

60.62

1.74

MS S

38.9

1.78

828/867

21.87

0.12

MS

13.11

1.85

802/829

22.18

0.14

MS

1305

1320

20.12

1.98

920/953

58.13

0.61

MS

1411

1425

27.73

2.01

855/857

31.93

0.75

MS S

35.34

1.98

865/933

59.59

0.34

MS S

42.59

1.96

879/940

64.23

0.25

MS

49.48

1.93

803/876

22.76

0.11

MS

23.92

2.41

873/890

48.76

0.06

MS AM S

39.02

2.27

853/923

57.97

0.09

MS S

46.15

2.21

815/863

42.34

0.14

MS S

41.99

3.73

952/956

68.37

2.05

MS

EP

14.64

c

I method

1.16

M AN U

1

SC

Number

RI PT

Table 1 The volatiles in Dezhou Braised Chicken by GC-qMS and GC×GC/HR-TOFMS

25.57

651

1785

31.47

844

1999

1807

34

ACCEPTED MANUSCRIPT

18

Benzeneacetaldehyde

19

Anisaldehyde

20 21

17.64

20

2.92

917/929

71.83

1.74

MS S

25.95

3.18

819/854

27.8

0.2

MS

40.8

3.85

878/901

52.45

0.54

MS

o-anisaldehyde

40.21

3.52

791/842

78.84

0.04

MS AM

Isovanillin

50.08

4.32

801/871

38.95

0.03

MS

31.12

846

1495

919

1496

1985

Ketones

RI PT

Benzaldehyde

SC

17

2-heptanone

15.84

1.69

863/866

32.32

0.11

MS

23

3-heptanone

24

3-hydroxy-2-butanone

11.57

844

1263

25

2-methyl-3-octanone

12.79

742

1308

26

3-methyl-1,2-cyclopentanedione

27

Piperitone

28

Acetophenone

841

15.72

1.62

851/872

18.72

0.07

MS

1286

5.98

1.43

839/841

62.36

5.03

MS

1323

22.5

1.75

812/825

33.54

1.26

MS

24.64

3.07

789/830

29.62

0.07

MS AM

41.75

2.6

806/812

26.28

0.07

MS

27.85

3.09

919/929

55.34

0.43

MS

1703

TE D

23.19

M AN U

22

Niacinamide

30

dimethyl sulfone

31

disulfide, dipropyl

32

3-(methylthio)-propanal

33

2-(methylthio)-ethanol

34

3-thiophenemethanol

35

Maltol

27.81

624

AC C

29

EP

Nitrogen containing, Oxygen containing or Sulfur-containing compounds

15.95

29.74

814

929

1861

1429

1930

1968

35

48.53

6.76

862/903

55.02

0.32

MS

14.89

6.84

921/924

92.72

3.05

MS AM

32.01

1.97

768/830

47.15

0.05

MS

15.6

3.03

855/876

93.49

0.29

MS

12.16

2.41

862/862

94.01

0.06

MS

25.23

3.28

859/862

72.35

0.07

MS

30.7

3.71

912/915

92.43

1.31

MS

Isomaltol

21.78

2.55

832/905

59.65

0.1

MS

37

2-acetylthiazole

23.92

3.42

728/826

97.88

0.06

MS AM

38

2-pyrrolidinone

26.42

6.07

827/862

74.8

0.15

MS

39

Benzothiazole

39.13

4.00

793/866

56.27

0.07

MS AM

40

1-methylimidazole-5-carboxalde

28.44

5.65

838/900

81.88

0.12

MS

732

1936

hyde 41

3,5-dihydroxy-2-methyl-4H-pyr an-4-one

42

2,3-dihydro-3,5-dihydroxy-6-me

35.91

821

2225

2H-1-benzopyran-2-one

38.67

925

44

4,5-dimethyl-1,3-dioxol-2-one

33.16

775

45

2-furanmethanol

21.19

947

46

2-pentyl furan

10.33

834

47

5-acetyldihydro-2(3H)-furanone

48

dihydro-3-hydroxy-4,4-dimethyl

49

2,5-dimethyl-4-hydroxy-3(2H)-

2451

2079

36.41

3.10

823/847

86.09

0.08

MS

33.55

3.12

881/929

95.35

0.62

MS

52.33

5.2

867/901

82.19

0.47

MS

25.71

4.1

806/823

59.21

4.31

MS

1639

1659

13.35

2.19

920/923

77.03

1.53

MS

1217

1231

23.81

1.54

890/890

47.7

0.13

MS S

29.75

6.26

781/851

39.26

0.04

MS AM

24.4

3.82

854/892

26.25

0.32

MS

27.13

3.09

852/868

81.76

0.32

MS

AC C

-2(3H)-furanone

2429

EP

43

TE D

thyl-4H-pyran-4-one

SC

29.88

RI PT

36

M AN U

ACCEPTED MANUSCRIPT

furanone

36

ACCEPTED MANUSCRIPT

51

1-hexanol

52

1-octanol

53

1-octen-3-ol

54

(Z)-2-octen-1-ol

55

2-ethyl-1-hexanol

56

2,4-dimethyl-cyclohexanol

57

4,5-dimethyl-2-hepten-3-ol

58

R-(-)-1,2-propanediol

19.45

933

1576

59

[S-(R*,R*)]-2,3-butanediol

19.12

901

1519

60

α-terpineol

22.45

912

1679

61

4-terpineol

19.91

892

62

3-phenyl-2-propen-1-ol

31.47

881

63

benzyl alcohol

64

anisic alcohol

butanoic acid

66

pentanoic acid

67

hexanoic acid

68

heptanoic acid

16.08

837

1338

904

1433

1.54

812/831

18.46

0.05

MS

14.77

1.53

812/869

31.74

0.42

MS

29.51

1.67

868/884

17.77

0.23

MS

22.74

1.59

926/929

75.74

0.81

MS

1.8

845/888

39.03

0.19

MS

1.61

862/896

43.13

0.2

MS

1353

1445

29.15

36.14

1473

1473

26.42

1697

TE D

838

1596

745

1602

2245

EP

17.09

AC C

65

13.58

9.3

2239

RI PT

3-methyl-2-buten-1-ol

SC

50

M AN U

Alcohols

26.54

1.94

817/829

25.74

0.06

MS

30.22

2.12

800/800

12.15

0.16

MS

7.16

1.65

916/940

59.44

1.16

MS

9.54

1.81

917/937

63.44

2.06

MS

38.07

2.08

905/936

69.44

0.36

MS S

37.24

1.99

898/901

51.83

0.42

MS

44.85

3.47

870/884

71.37

0.83

MS

25.71

3

869/891

50.68

0.26

MS

43.06

3.71

855/868

36.46

0.11

MS

9.78

1.51

867/869

58.71

0.26

MS

15.6

1.72

879/891

33.67

0.15

MS

Acids

20.33

839

1611

1630

23.37

812

1710

26.51

813

1817

1833

23.09

1.91

866/884

68.92

1.43

MS

29.60

713

1925

1943

29.63

1.82

880/883

54.75

0.16

MS

37

70

32.14

845

2030

36.76

1.87

(S)-2-hydroxypropanoic acid

17.15

2.39

71

3-methyl-butanoic acid

12.99

72

2-methyl-butanoic acid

13.7

73

dehydroacetic acid

47.82

59.15

0.64

MS

822/887

36.81

1.13

MS

1.52

815/846

75.86

0.19

MS

1.5

874/891

47.06

0.15

MS

3.9

835/862

25.03

0.06

MS

Esters 74

butyrolactone

75

dihydro-3-hydroxy-4,4-dimethyl

20.23

856

1608

-2(3H)-furanone

827/857

RI PT

octanoic Acid

SC

69

M AN U

ACCEPTED MANUSCRIPT

15.13

4.84

945/968

80.68

0.54

MS

24.40

3.82

908/919

32.29

0.37

MS

26.9

1.64

927/931

82.87

0.56

MS

44.13

2.65

922/929

44.31

0.77

MS

38.19

2.46

828/843

19.76

0.06

MS S

60.18

1.98

870/887

60.41

0.3

MS

48.65

2.79

929/930

22.77

2.67

MS

47.46

3.80

796/814

70.65

0.05

MS AM

eucalyptol

9.80

930

77

1-methoxy-4-(1-propenyl)-

25.94

920

21.53

benzene estragole

79

butylated hydroxytoluene

80

2-methoxy-3-(2-propenyl)-phen ol

81

580

AC C

78

34.15

946

1197

1209

TE D

76

EP

Ethers

1797

1650

Phenols

2129

2,6-dimethoxy-phenol

38

ACCEPTED MANUSCRIPT

874/926

37.03

0.08

MS

83

undecane

32.96

1.12

846/882

15.06

0.23

MS

84

dodecane

40.09

1.16

927/933

24.18

0.76

MS

85

2,6,11-trimethyl-dodecane

46.99

1.18

838/884

5.55

0.31

MS

86

1,3-dimethyl-benzene

1.55

911/929

35.78

1.29

MS

87

1,3-diethyl-benzene

1.79

840/869

31.15

0.12

MS

88

1,3-diethenyl-benzene

89

1,4-diethenyl-benzene

90

ethylbenzene

91

1-ethenyl-4-ethyl-benzene

92

1-ethenyl-3-ethyl-benzene,

93

styrene

94

D-limonene

95

azulene

7.19

7.91

869

1097

884

1125

1099

1134

15.13 28.44

7.56

9.49

764

920

1112

1122

1185

1192

RI PT

1.08

SC

decane

TE D

82

EP

a

25.47

M AN U

Hydrocarbons

32.36

2.26

959/962

35.78

1.29

MS

33.2

2.31

876/919

24.53

0.14

MS

14.53

1.56

911/948

65.5

0.12

MS

31.06

2.02

942/951

37.88

0.52

MS

30.46

2.01

928/941

22.38

1.48

MS

16.2

1.86

951/954

64.48

1.25

MS

27.02

1.48

900/901

21.62

0.39

MS

37.12

2.96

917/932

36.18

0.39

MS

RI: retention index, bRI*: retention index from http://www.odour.org.uk/, cIdentification method: MS, compared with NIST08 Mass Spectra Database; AM, agrees

AC C

with the accurate mass determination; S, agree with mass spectrum of authentic standards.

39

ACCEPTED MANUSCRIPT


A fast and effective extraction method (ASE-SAFE) is set up.




An accurate identification method (GC×GC/HR-TOFMS) is set up.




Some very polar and unstable odour-active and trace constitutes can be qualified.




6

Compounds

of

2-enals

or

2,4-dineals

are

only

identified

by

The problem of overlapped peaks in TIC of GC-MS is resolved by

EP

TE D

M AN U

SC

GC×GC/HR-TOFMS.

AC C




RI PT

GC×GC/HR-TOFMS.