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  • 1 Technology Centre, China Tobacco Guangxi Industrial Co. Ltd., Nanning, 530001, P.R. China
  • 2 University of Science and Technology of China, Hefei, 230052, P.R. China
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A credible method for determination of the aglycon moieties of glycosidically bound aroma compounds in Flos Chrysanthemi by comprehensive two-dimensional gas chromatography–time-of-flight mass spectrometry (GC × GC–TOFMS) has been proposed. The aglycon moieties of glycosidically bound aroma compounds were isolated using methyl-tert-butyl ether (MTBE) extraction following enzymatic hydrolysis. The GC × GC–TOFMS analysis was performed to comprehensively identify different forms of the released aroma components in Flos Chrysanthemi. The result shows that the limit of detection of the released aglycon moieties ranged from 0.3 to 3.1 ng/mL, the recovery of the released 1-octanol was better than 98.3%, and the intra-day and inter-day precisions of this method were 0.2 to 8.9% and 1.3 to 9.1%, respectively. The proposed method was applied to the analysis of four types of Flos Chrysanthemi (Chuju, Boju, Hangju, and Gongju). A total of 60 aglycon moieties of interest were identified in the four types of Flos Chrysanthemi. These aglycones mainly consisted of aliphatic, aromatic, monoterpene, C13-norisoprenoids, and miscellaneous compounds.

Abstract

A credible method for determination of the aglycon moieties of glycosidically bound aroma compounds in Flos Chrysanthemi by comprehensive two-dimensional gas chromatography–time-of-flight mass spectrometry (GC × GC–TOFMS) has been proposed. The aglycon moieties of glycosidically bound aroma compounds were isolated using methyl-tert-butyl ether (MTBE) extraction following enzymatic hydrolysis. The GC × GC–TOFMS analysis was performed to comprehensively identify different forms of the released aroma components in Flos Chrysanthemi. The result shows that the limit of detection of the released aglycon moieties ranged from 0.3 to 3.1 ng/mL, the recovery of the released 1-octanol was better than 98.3%, and the intra-day and inter-day precisions of this method were 0.2 to 8.9% and 1.3 to 9.1%, respectively. The proposed method was applied to the analysis of four types of Flos Chrysanthemi (Chuju, Boju, Hangju, and Gongju). A total of 60 aglycon moieties of interest were identified in the four types of Flos Chrysanthemi. These aglycones mainly consisted of aliphatic, aromatic, monoterpene, C13-norisoprenoids, and miscellaneous compounds.

Introduction

Flos Chrysanthemi, dried flower of Compositae Chrysanthemum morifolium Ramat, which belongs to the Compositae family, is well-known for its physiological functions, such as antipyresis, antitumor, antifungal, antiviral, anti-oxidation, and immunomodulatory activities [14]. The major quality attributes of Flos Chrysanthemi are appearance, aroma, and color [5]. However, the quality of Flos Chrysanthemi and its market price are commonly judged by the aroma. The volatile chemical compounds of Flos Chrysanthemi are mainly composed of monoterpenes, sesquiterpenes, aldehydes, acids, esters, and alcohols [6]. Additionally, these compounds can also combine with sugars and be stored in the form of odorless glycosides in tissues serving as flavor precursors. Glycosides are able to release free aroma compounds by enzymatic or chemical cleavage during plant maturation and can be considered as a source of latent and potential aroma [7]. Thus, the development of reliable analytical method to identify and quantify glycosidically bound aroma compounds in Flos Chrysanthemi has assumed enormous significance.

Numerous methods have been developed to analyze glycosidically bound aroma compounds. The glycosides can be directly determined by liquid chromatography–tandem mass spectrometry (LC–MS/MS) [8] or detected by one-dimensional gas chromatography–mass spectrometry after derivatization which can directly provide structural information of glycoside [9]. However, because the reference glycosides are not commercially available, indirect methods have more often been applied for the determination of glycosidically bound aroma compounds in plant [10]. Indirect methods included acidic and enzymatic hydrolysis; acid hydrolysis is relatively simple but always with harsh condition which could change the structure of released aglycon [10, 11]. Because of the specificity of enzymatic hydrolysis and gentle condition, the aglycones released by enzymatic hydrolysis could provide more original structural information [9, 12]. Moreover, traditional one-dimensional gas chromatography–mass spectrometry, which has poor peak capacity and sensitivity, can identify about 20 volatile components in Flos Chrysanthemi [1]. Recently, research has confirmed that GC × GC–TOFMS analysis accurately identified more aroma components than one-dimensional gas chromatography–mass spectrometry [9, 10, 1416]. However, there are very few studies investigating the quantitative analysis of the aglycon moieties of glycosidically bound aroma compounds in Flos Chrysanthemi by enzymatic hydrolysis and GC × GC–TOFMS.

The main objective of this study is the development of a credible methodology for extraction and analysis of the aglycon moieties of glycosidically bound aroma compounds in Flos Chrysanthemi. The aglycon moieties of glycosidically bound aroma compounds were isolated using methyl-tert-butyl ether (MTBE) extraction following enzymatic hydrolysis. The GC × GC–TOFMS analysis was performed to comprehensively identify different forms of the released aroma components in Flos Chrysanthemi. The proposed method was applied to the analysis of four types of Flos Chrysanthemi.

Experimental

Materials

All reference aroma compounds were supplied by Bioko century Biotechnology Co., Ltd. (Beijing, China). Octyl-β-d-glucopyranoside, phenyl-β-d-glucopyranoside (internal standard), and β-glucosidase from almonds were purchased from TCI (Tokyo, Japan). The purity of all chemical standards was ≥97%. Methanol (MEOH), dichloromethane (DCM), methyl-tert-butyl ether (MTBE), cyclohexane (CYHEX), petroleum ether (PETH), and chloroform (CHL) were of analytical grade and supplied by Tedia Company, Inc. (Faifield, USA). n-Paraffins C6–C40 (purity, >95.5%) were from Sigma-Aldrich (St. Louis, MO, USA). High-purity water was prepared by a Milli-Q water purification system (Millipore, Bedford, MA, USA). The internal standard solution of phenyl-β-d-glucopyranoside (25 mg/mL) was prepared by methanol.

Chuju was supplied from Research Department of Chuju in Chuzhou city (China). Boju, Gongju, and Hangju were purchased from a local pharmacy in Bozhou, Hangshan, and Tongxiang city (China), respectively. About 1 kg of each cultivar was dried at 40 °C for 5 h and then grounded to a 40 mesh powder. Chuju was used to optimize the method.

Sample Preparation

One gram powder of Chuju was introduced into a 50 mL capped flask, and 10 μL of internal standard was added. The extraction was performed under sonication with 20 mL of methanol for 30 min. The extraction was repeated twice more. The three extracts were combined, filtered, and dried under nitrogen stream. Then the solid residue was dissolved in 5 mL of 0.2 mol/mL sodium citrate buffer (pH = 5.5). The solution was then extracted three times with 20 mL of MBTE to remove any traces of free aglycones. The conjugated aroma components were insoluble in MBTE and, thus, remained in the aqueous phase. To generate the free form of the conjugated aroma components, 5 units of β-glucosidase from almonds were added. The mixture was sealed and incubated at 40 °C for 48 h. The conjugated aroma components were hydrolyzed, and the released aglycones were extracted with 3 mL of MBTE; 2 μL of the upper organic phase was used for GC × GC–TOFMS analysis. A blank sample was run without addition of β-glucosidase from almonds, which was replaced by 0.2 mol/mL sodium citrate buffer (pH = 5.5). All assays were performed in triplicate.

Equipment and Conditions

A GC × GC–TOFMS system consisted of an Agilent 7890 gas chromatography unit (Agilent Technologies, Palo Alto, CA, USA) equipped with a cold jet loop modulator (KT 2001; Zoex, Lincoln, NE, USA), and a Pegasus 4D TOFMS (LECO, St. Joseph, MI, USA) was used. An HP-5MS column (30 m × 0.25 mm × 0.25 μm, Agilent Technologies, Palo Alto, CA, USA) was used as the first dimension column and a DB-17 column (1.5 m × 0.1 mm × 0.1 μm, Agilent Technologies, Palo Alto, CA, USA) as the second dimension column. Helium (99.999% purity), provided by Guangxi Guoxin Gas Co., Ltd. (Nanning, China), was used as the carrier gas at a rate of 1.0 mL/min. The primary oven temperature was set at 60 °C for 1 min, ramped at 5 °C/min to 270 °C, and held for 1 min. The secondary oven temperature was programmed to be 10 °C higher than the primary oven. The injector temperature was 250 °C with a split ratio, 1:50. The total modulation time was 4 s. The transfer line temperature was 260 °C, and the ion source temperature was 230 °C. The electron energy was 70 eV. Data acquisition was performed with an acquisition rate of 100/spectra in the mass range of a mass-to-charge ratio of m/z = 33–450. Tentative identification of compounds was based on a comparison with NIST 08 mass spectral library after deconvolution performed using Chroma TOF software (version 4.42; LECO).

Results and Discussion

Optimization of Sample Preparation Conditions

Enzymatic hydrolysis is a complex process with many parameters that may be influenced by many factors such as the type of enzyme, the hydrolysis pH, the process time, temperature, etc. [1618]. According to a previous study [9], 0.2 mol/mL sodium citrate buffer (pH = 5.5), 40 °C, and 48 h were ultimately selected as the hydrolysis pH, temperature, and time for this study, respectively. To further optimize the conditions for hydrolysis of the aglycones from Flos Chrysanthemi, the type and volume of extraction solvent were investigated. Figure 1a shows that MTBE was the most efficient solvent for extracting the aglycones from Flos Chrysanthemi, and MTBE was ultimately selected as the extraction solvent for this study. This is probably due to the fact that the polarity of MTBE is similar to that of most of the aglycones [14]. No obvious differences in the sum peak area were observed using 3 mL, 4 mL, and 5 mL of MTBE, as shown in Figure 1b. In addition, considering the cost, 3 mL of MTBE is sufficient for the aglycones extraction process.

Figure 1.
Figure 1.

a: Effect of the type of extraction solvent on the extraction yield; b: total peak area of all analytes using different volumes of MTBE

Citation: Acta Chromatographica Acta Chromatographica 30, 3; 10.1556/1326.2017.00278

GC × GC Analysis

In the GC × GC analysis, the selection of column system was important. Considering that the majority of enzyme-released aglycones from Flos Chrysanthemi are medium or weak polar components, a non-polar column was used as the first column and a mid-polar column as the second column. In this case, the first column separation is based on volatility, whereas, in the second column, analytes are separated mainly according to their polarity. In this way, orthogonality has been achieved and an apparent group-type separation of some major enzyme-released aglycones in the sample was obtained, as shown in Figure 2. The components identified as aliphatic, aromatic, and monoterpene compounds were located in the region marked (a)–(c), respectively. Similarly, some components identified as C13-norisoprenoids were located in region (d). The peaks were distributed over the entire chromatogram, and no wrap-around phenomenon was observed, as shown in Figure 2. All these results indicate that the column system used in this study was appropriate.

Figure 2.
Figure 2.

The GC × GC–TOFMS total ion current chromatogram of Chuju extract

Citation: Acta Chromatographica Acta Chromatographica 30, 3; 10.1556/1326.2017.00278

Identification and Quantification

A contour plot (total ion current chromatogram, TIC) of the aglycones released by β-d-glucosidase hydrolysis from glycosidic fractions in Chuju is shown in Figure 2. Autoprocessing resulted in a peak table containing over 257 peaks (indicated as bubbles) with S/N of ≥100. According to the literature [19, 20], a similarity and reverse number above 800 with the assistance of linear temperature programed retention indices (LTPRIs) were employed as criteria for tentative identification of compounds. Finally, 41 aglycones of interest, including 12 aliphatic compounds, 13 aromatic compounds, 9 monoterpene, 5 C13-norisoprenoids, and 2 miscellaneous compounds were tentatively identified from glycosidic fractions in Chuju using GC × GC–TOFMS (listed in Table 1). Compared to previous study [9, 10], the GC × GC–TOFMS analysis of this study accurately identified more adequate components.

Table 1.

Analytical performance of the method for analysis of the aglycones by GC × GC–TOFMS

No.ComponentsR.T. (s)CASLTPRISimilarityReverseCalibration curveCorrelation coefficientLOD (ng/mL)Intra-day precision (%, n = 5)Inter-day precision (%, n = 5)
SampleReferencea
Monoterpene
16-Camphenol899b, 2.090c3570-04-5111911108098541.62.3
2Verbenol927, 1.980473-67-6113411538368391.62.6
3Pinocarveol*d967, 2.060547-61-511511145910910y = 0.138x - 0.8590.99951.53.34.3
4Lavandulol*999, 1.94058461-27-111711170915915y = 0.237x - 1.2320.99920.33.45.1
5Isoborneol*1011, 2.140124-76-511761160882904y = 0.146x - 0.7320.99922.10.21.5
64-Terpinenol1027, 2.010562-74-3118811808968961.43.8
7Myrtenol*1059, 2.10019894-97-412001198804818y = 0.866x - 0.8240.99910.43.54.1
88-Hydroxylinalool1311, 2.37064142-78-5128413368919092.22.9
9Z-a-trans-bergamotol1777, 2.19088034-74-6169717088278456.36.8
C13-Norisoprenoids
103-Oxo-a-ionol1739, 2.56034318-21-3166716468938931.32.6
113-Hydroxy-7,8-dihydro-a-ionol1795, 2.500172705-13-4170917008568587.09.1
12Blumenol C1823, 2.49036151-02-7172417138468470.72.3
13Solanone1663, 2.29054868-48-316087807856.57.1
145,6-Epoxy-3-hydroxy-ß-ionone1787, 2.48038274-01-0170316908668662.32.8
Aromatic compounds
15Benzaldehyde*655, 2.210100-52-7966967928928y = 0.290x - 1.5700.99930.71.62.1
16Benzyl alcohol*775, 2.310100-51-610421035903904y = 0.268x - 0.7990.99981.21.01.6
17Benzene acetaldehyde*791, 2.320122-78-110511044949949y = 0.200x - 1.6640.99991.18.39.0
183-Methyl-phenol839, 2.240108-39-4107910709159188.08.5
19Phenylethyl alcohol*915, 2.36060-12-811231118950954y = 0.270x - 1.2240.99980.34.06.1
202,3-Dihydro-benzofuran1083, 2.410496-16-2120912198328513.03.8
21Benzenepropanol1111, 2.170122-97-4122012289479532.02.8
22Cuminaldehyde*1127, 2.370122-03-212261237923936y = 0.279x - 1.2870.99971.11.51.9
23Hydroquinone*1171, 2.760123-31-912401241890905y = 0.554x - 0.8960.99911.22.83.9
244-Methoxy-benzenemethanol1199, 2.590105-13-5124812739109104.15.1
252-Methoxy-4-vinylphenol*1247, 2.4007786-61-012621309947947y = 0.237x - 1.2690.99921.23.74.2
262-Methoxy-3-(2-propenyl)-phenol1315, 2.3301941-12-4128413629479471.12.9
274-Hydroxy-benzeneethanol1419, 2.820501-94-0143714318768922.23.1
Aliphatic compounds
282-Methyl-1-butanol*359, 1.5101565-80-6737934934y = 0.336x - 0.2680.99931.11.82.9
293-Methyl-2-buten-1-ol*395, 1.630556-82-1777779896896y = 0.142x - 0.2400.99890.33.14.6
302-Methyl-1-pentanol*459, 1.650105-30-6830824926926y = 0.114x - 0.2400.99993.10.51.9
313-Hexen-1-ol*491, 1.740544-12-7857858944944y = 0.118x - 0.6480.99971.30.41.3
321-Hexanol*503, 1.710111-27-3866867917917y = 0.280x - 2.3870.99920.82.42.8
332-Heptanol*547, 1.700543-49-7899887957957y = 0.142x - 0.8120.99910.91.92.9
341-Octen-3-ol*671, 1.7803391-86-4979978952952y = 0.129x - 0.5400.99901.20.51.3
35Sulcatol691, 1.8104630-06-29949219295.37.1
363-Octanol699, 1.760589-98-09949959149217.68.2
371-Octanol*827, 1.840111-87-510721070923923y = 0.343x - 5.6270.99911.95.37.6
386-Methyl-5-hepten-2-one*687, 1.900110-93-0989987956956y = 0.560x - 6.8900.99922.14.75.7
393-Methyl-2-cyclohexen-1-one*823, 2.3601193-18-610671064945945y = 0.214x - 0.2400.99952.31.51.8
Miscellaneous compounds
40Furfural*459, 2.00098-01-1836835943943y = 0.150x - 1.0820.99941.58.99.1
414-Hydroxydihydro-2(3H)-furanone983, 3.2905469-16-9116111539379373.06.9

LTPRI reference values according to NIST database (http://webbook.nist.gov/chemistry/).

The first dimension retention time.

The second dimension retention time.

Compounds with an asterisk confirmed with standards. Compounds without an asterisk were tentatively identified and semi-quantified.

Five-point calibration curves were calculated by plotting the peak area ratio (y) of the analyte and internal standard against the concentrations (x, μg/mL). All compounds marked with an asterisk (*) were quantified by the linear equation, as shown in Table 1. The limit of detection (LOD) defined as a signal-to-noise ratio of 3:1 ranged from 0.3 to 3.1 ng/mL. Correlation coefficient (r) of the calibration curves were greater than 0.9989. The quantitative data of other compounds with no chemical standards were obtained by the semi-quantitative method using phenyl-β-d-glucopyranoside as reference, without considering calibration factors, i.e., setting the slope and intercept value of the linear equation as 1 and 0, respectively. The most abundant characteristic ion was used as quantitative ion.

Accuracy and Repeatability

The validation of the method was carried out with accuracy and repeatability, and the accuracy of the method was estimated by the recovery of known amounts of octyl-β-d-glucopyranoside. The recovery of the released 1-octanol was better than 98.3%. The intra-day and inter-day precisions of this method were evaluated using multiple preparations of the same sample. Five replicate samples were prepared and analyzed in a single day and on five different days. As shown in Table 1, the intra-day and inter-day precisions of this method were 0.2 to 8.9% and 1.3 to 9.1%, respectively.

Application

The practical applicability of this technique was evaluated by analyzing the four cultivars of Flos Chrysanthemi. Quantitative data of the four cultivars of Flos Chrysanthemi were shown in Table 2. As can be observed in Table 2, 60 aglycones, including 17 aliphatic compounds, 19 aromatic compounds, 12 terpenoids, 6 C13-norisoprenoids, and 6 miscellaneous compounds, have been detected. Also, 21 aglycones (e.g., benzyl alcohol, phenylethyl alcohol, blumenol C, 3-oxo-α-ionone, 3-hydroxy-7,8-dehydro-β-ionol, 1-octanol) are present in all four cultivars of Flos Chrysanthemi. In addition, the total concentration of the aglycones released by β-d-glucosidase hydrolysis from glycosidic fractions in four Flos Chrysanthemi range from 90 to 304 mg/kg, Hangju proved to have the highest level of aglycones, and Boju has the lowest level. Meanwhile, a Fisher's least significant difference (LSD) test was used to rank the means and to identify the means that were different. The result illustrated that the glycosidic fractions of the four Flos Chrysanthemi were significantly different.

Table 2.

The concentrations (mg/kg) of aglycones released by enzymatic hydrolysis from glycosidic fractions in four types of Flos Chrysanthemi

No.ComponentsChujuBojuHangjuGongju
Monoterpenes
16-Camphenol5.04 ± 0.08a3.64 ± 0.02 bb
2Pinocarveol*c1.25 ± 0.02 a1.80 ± 0.01 b3.16 ± 0.05 c4.67 ± 0.10 d
3Verbenol1.31 ± 0.04 a1.01 ± 0.01 b
4Lavandulol*1.18 ± 0.04 a1.06 ± 0.05 b
5Verbenol2.07 ± 0.06 a1.07 ± 0.04 b
6Isoborneol*5.81 ± 0.01 a3.03 ± 0.02 b0.84 ± 0.02 c0.72 ± 0.03 d
74-Terpinenol5.03 ± 0.07 a0.98 ± 0.03 b
8Myrtenol1.42 ± 0.05 a1.09 ± 0.11 b1.15 ± 0.05 b
9Carveol1.08 ± 0.04
108-Hydroxylinalool2.72 ± 0.06 a1.64 ± 0.02 b1.8 ± 0.2 c2.4 ± 0.2 d
11Z-α-bergamotol1.44 ± 0.09 a1.38 ± 0.02 b
C13 Norisoprenoids
123-Hydroxy-7,8-dihydro-α-ionol2.3 ± 0.2 a1.39 ± 0.03 ab23 ± 6 c8.8 ± 0.1 d
133-Oxo-α-ionol39.2 ± 0.5 a16.14 ± 0.04 b67.22 ± 0.05 c59.6 ± 0.9 d
14Blumenol C14.7 ± 0.1 a13.40 ± 0.06 b22.0 ± 0.7 c15.3 ± 0.2 d
153,4-Dehydro-β-ionone1.43 ± 0.04
16Solanone0.93 ± 0.06 a2.03 ± 0.06 b2.9 ± 0.1 c
175,6-Epoxy-3-hydroxy-β-ionone0.44 ± 0.01 a1.68 ± 0.05 b0.72 ± 0.03 c
Benzene derivatives
18Benzaldehyde*5.09 ± 0.08 a2.4 ± 0.1 b5.5 ± 0.1 c12.4 ± 0.1 d
19Benzyl alcohol*32.4 ± 0.3 a25.4 ± 0.1 b40.5 ± 0.1 c45 ± 2 d
20Benzeneacetaldehyde*1.2 ± 0.1 a1.08 ± 0.02 ab0.66 ± 0.05 c1.33 ± 0.03 ad
212-Methylphenol0.72 ± 0.04 a0.53 ± 0.06 b
223-Methyl-phenol0.50 ± 0.04 a0.61 ± 0.01 b0.36 ± 0.03 c
232-Methoxyphenol0.56 ± 0.03
24Phenylethyl alcohol*9.9 ± 0.4 a11.4 ± 0.9 ab63.4 ± 0.3 c70 ± 2 d
252,3-Dihydro-benzofuran0.66 ± 0.02 a0.14 ± 0.02 b0.65 ± 0.02 ac0.52 ± 0.01 d
26Benzenepropanol0.51 ± 0.01 a0.65 ± 0.01 b1.71± 0.1 c1.06 ± 0.01 d
27Cuminaldehyde*0.65 ± 0.02
28Hydroquinone*1.44 ± 0.04 a0.41 ± 0.04 b1.17 ± 0.06 c0.69 ± 0.04 d
294-Methoxy-benzenemethanol3.2 ± 0.1 a1.75 ± 0.02 b7.97 ± 0.03 c
303-Phenyl-2-propen-1-ol0.56 ± 0.03
312-Methoxy-4-vinylphenol*1.07 ± 0.04 a0.67 ± 0.02 b2.64 ± 0.06 c
322-Methoxy-3-(2-propenyl)-phenol2.73 ± 0.03 a1.44 ± 0.07 b
332-(4-Methoxyphenyl)ethanol5.92 ± 0.01
344-Hydroxy-benzeneethanol0.46 ± 0.01 a1.53 ± 0.02 b0.61 ± 0.05 c
354-Hydroxy-acetylphenol0.82 ± 0.02 a0.83 ± 0.02 a
364-(4-Hydroxyphenyl)-2-butanone0.58 ± 0.05
37Homovanillyl alcohol0.66 ± 0.04
Aliphatic compounds
382-Methyl-1-butanol*4.89 ± 0.09 a0.14 ± 0.01 b7.05 ± 0.05 c5.13 ± 0.05 d
391-Hepten-4-ol0.85 ± 0.04
401-Pentanol2.39 ± 0.06
413-Methyl-2-buten-1-ol*3.2 ± 0.1 a3.16 ± 0.01 a1.26 ± 0.02 b
422-Methyl-1-pentanol*1.82 ± 0.01 a0.89 ± 0.02 b1.14 ± 0.02 c
433-Hexen-1-ol*5.43 ± 0.02 a3.4 ± 0.2 b6.92 ± 0.03 c16.34 ± 0.03 d
441-Hexanol*4.2 ± 0.1 a1.4 ± 0.1 b2.11 ± 0.08 c4.07 ± 0.04 a
452-Heptanol*0.52 ± 0.01 a0.72 ± 0.01 b0.62 ± 0.01 c
464-Hepten-1-ol1.04 ± 0.02
471-Heptanol1.07 ± 0.06
481-Octen-3-ol*4.07 ± 0.02 a0.60 ± 0.06 b12.3 ± 0.1 c5.5 ± 0.3 d
49Sulcatol0.94 ± 0.05 a1.4 ± 0.2 b
503-Octanol1.18 ± 0.09 a0.86 ± 0.03 ab1.26 ± 0.04 ac
511-Octanol*0.75 ± 0.04 a0.86 ± 0.04 a1.75 ± 0.08 b0.98 ± 0.08 c
52Hexanal0.67 ± 0.01 a0.26 ± 0.01 b1.05 ± 0.02 c
536-Methyl-5-hepten-2-one*0.85 ± 0.04 a1.24 ± 0.02 b
543-Methyl-2-cyclohexen-1-one*1.33 ± 0.02 a1.26 ± 0.05 b
Miscellaneous
552-Furylmethanol0.93 ± 0.02
56Furfural*0.45 ± 0.04 a2.5 ± 0.3 b0.75 ± 0.07cd0.70 ± 0.02 ad
573-Hydroxy-2,3-dihydromaltol0.25 ± 0.01
582-Hydroxycineole0.43 ± 0.05 a0.26 ± 0.03 b
594-Hydroxydihydro-2(3H)-furanone0.66 ± 0.02 a0.72 ± 0.03 b0.83 ± 0.01 c0.90 ± 0.03 d
607-Hydroxycoumarin2.7 ± 0.3
Total17391304279

Concentrations of the same compound with different letters show significant differences (based on Fisher's LSD test, p < 0.05).

Not detected.

Compounds with an asterisk confirmed with standards. Compounds without an asterisk were tentatively identified and semi-quantified.

Conclusion

A reliable method for determination of the aglycon moieties of glycosidically bound aroma compounds in Flos Chrysanthemi has been proposed. The aglycon moieties of glycosidically bound aroma compounds were isolated using MTBE extraction following enzymatic hydrolysis. The GC × GC–TOFMS analysis was performed to comprehensively identify different forms of the released glycosidically bound aroma components in Flos Chrysanthemi. The proposed method was applied to the analysis of four types of Flos Chrysanthemi. A total of 60 aglycone moieties of interest were identified in four types of Flos Chrysanthemi (Chuju, Boju, Hangju, and Gongju). These aglycone moieties mainly consisted of aliphatic, aromatic, monoterpene, C13-norisoprenoids, and miscellaneous compounds. Additionally, 21 aglycones (e.g., benzyl alcohol, phenylethyl alcohol, blumenol C, 3-oxo-α-ionone, 3-hydroxy-7,8-dehydro-β-ionol, 1-octanol) are present in all four types of Flos Chrysanthemi.

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    Humpf, H. U.; Schreier, P. J. Agric. Food Chem. 1991, 39, 1830.

  • 11.

    Martínez-Gil, A. M.; Angenieux, M.; Pardo-García, A. I.; Alonso, G. L.; Ojeda, H.; Salinas, M. R. Food Chem. 2013, 138, 956.

  • 12.

    Zazoğlu, S.; Anilanmert, B.; Aydin, M.; Cengiz, S. Acta Chromatogr. 2017, 29, 253.

  • 13.

    García-Carpintero, E. G.; Sánchez-Palomo, E.; Gallego, M. A. G.; González-Viñas, M. A. Food Chem. 2012, 131, 90.

  • 14.

    Yu, D.; Zhu, L.; Liu, S.; Yu, H.; Dai, Y. J. Chromatogr. A 2013, 1280, 122.

  • 15.

    Sghaier, L.; Cordella, C. B. Y.; Rutledge, D. N.; Watiez, M.; Breton, S.; Kopczuk, A.; Sassiat, P.; Thiebaut, D. J. Vial Chromatographia 2015, 78, 805.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 16.

    Perrault, K. A.; Nizio, K. D.; Forbes, S. L. Chromatographia 2015, 78, 1057.

  • 17.

    Sarry, J. E.; Günata, Z. Food Chem. 2004, 87, 509.

  • 18.

    Ortizserrano, P.; Gil, J. V. J. Agric. Food Chem. 2007, 55, 9170.

  • 19.

    Shellie, R. A.; Marriott, P. J.; Huie, C. W. J. Sep. Sci. 2003, 26, 1185.

  • 20.

    Mühlen, C. V.; Zini, C. A.; Caramão, E. B.; Marriott, P. J. J. Chromatogr. A 2008, 1200, 34.

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    Wu, Q.; Deng, C.; Shen, S.; Song, G.; Hu, Y.; Fu, D.; Chen, J.; Zhang, X. Chromatographia 2004, 59, 763.

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    Kai, C.; Xiang, Z.; Pan, W.; Zhao, H.; Zhu, R.; Bo, L.; Geng, Z. J. Chromatogr. A 2013, 1311, 149.

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    Humpf, H. U.; Schreier, P. J. Agric. Food Chem. 1991, 39, 1830.

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    Martínez-Gil, A. M.; Angenieux, M.; Pardo-García, A. I.; Alonso, G. L.; Ojeda, H.; Salinas, M. R. Food Chem. 2013, 138, 956.

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    Zazoğlu, S.; Anilanmert, B.; Aydin, M.; Cengiz, S. Acta Chromatogr. 2017, 29, 253.

  • 13.

    García-Carpintero, E. G.; Sánchez-Palomo, E.; Gallego, M. A. G.; González-Viñas, M. A. Food Chem. 2012, 131, 90.

  • 14.

    Yu, D.; Zhu, L.; Liu, S.; Yu, H.; Dai, Y. J. Chromatogr. A 2013, 1280, 122.

  • 15.

    Sghaier, L.; Cordella, C. B. Y.; Rutledge, D. N.; Watiez, M.; Breton, S.; Kopczuk, A.; Sassiat, P.; Thiebaut, D. J. Vial Chromatographia 2015, 78, 805.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 16.

    Perrault, K. A.; Nizio, K. D.; Forbes, S. L. Chromatographia 2015, 78, 1057.

  • 17.

    Sarry, J. E.; Günata, Z. Food Chem. 2004, 87, 509.

  • 18.

    Ortizserrano, P.; Gil, J. V. J. Agric. Food Chem. 2007, 55, 9170.

  • 19.

    Shellie, R. A.; Marriott, P. J.; Huie, C. W. J. Sep. Sci. 2003, 26, 1185.

  • 20.

    Mühlen, C. V.; Zini, C. A.; Caramão, E. B.; Marriott, P. J. J. Chromatogr. A 2008, 1200, 34.

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