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. 2025 May 27;28:102590. doi: 10.1016/j.fochx.2025.102590

Characterization of volatile organic compounds in wild thyme (Thymus serpyllum L.) from central and northern China based on comprehensive GC × GC-TOFMS and chemometrics

Shasha Qi a,1, Chengjie Hou b,1, Honglei Tian c, Ping Zhan c,⁎, Bin Qiu a,⁎
PMCID: PMC12167021  PMID: 40520703

Abstract

To optimize the compatibility with target ingredients in food processing and ensure standardized application, it is essential to characterize thyme's volatile organic compounds (VOCs) from specific species and origins. In this study, 116 VOCs were identified in thyme samples from four main Chinese production regions by two-dimensional gas chromatography/time-of-flight mass spectrometry (GC × GC-TOFMS). Among them, eight compounds- thymol (5.27 %–13.28 %), eucalyptol (9.25–12.23 %), thymoquinone (11.16–15.98 %), etc.- were identified as the most abundant constituents. Samples NX, WQ, JB, and NM were classified into different chemotypes: thymol/eucalyptol/thymoquinone/p-cymene, thymoquinone/eucalyptol/thymol, thymoquinone/eucalyptol/p-cymene, and thymoquinone/carvacrol/isoborneol, respectively. Forty odor-active compounds were determined through relative odor activity value (ROAV) analysis. Furthermore, the variable importance in projection (VIP) method was applied to identify 14 and 5 VOCs as potential markers for thyme volatility and odor activity, respectively. These findings provide a valuable reference for expanding the application of Chinese thyme as a culinary herb in the food processing industry.

Keywords: Thyme (Thymus serpyllum L.), Volatility spectrum, GC × GC-TOFMS, Multivariate statistical analysis

Graphical abstract

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Graphical abstract

Highlights

  • •

    Volatile profiles of Chinese thyme were characterized using GC × GC-TOFMS.

  • •

    Eight compounds were the most abundant of the 116 VOCs identified.

  • •

    Thymoquinone was the highest volatile compound in the WQ, JB, and NM samples.

  • •

    Forty odor-active compounds were obtained by ROAV calculations.

  • •

    Potential markers were obtained through PLS-DA.

1. Introduction

Thyme (Thymus spp.), a member of the Lamiaceae family, comprises a diverse group of species with complex taxonomic classifications. Originating primarily from the Mediterranean region, these species are now widely distributed across North Africa, North America, Europe, Greenland, Abyssinia, and Asia, predominantly in temperate zones at higher latitudes (Li et al., 2019). Wild thyme (Thymus serpyllum L.) represents one of the most significant commercial varieties globally (Jalil et al., 2024). As a medicinal plant and natural food additive, thyme has attracted much attention for a long time.

Extensive research has demonstrated various pharmacological activities of Thymus, including digestion enhancement, antihypertension, anti-inflammatory, anti-malarial, and anticancer properties (Jalil et al., 2024; Knaub et al., 2022). Furthermore, its notable antibacterial and antioxidant properties make it an essential component in food preservation and extending the shelf life of food products (Galovičová et al., 2021; Šojić et al., 2020).

Another key function of T. serpyllum lies in its culinary herb role, which improves or enhances food flavor. This traditional utilization has been documented in numerous countries (Marc et al., 2022; Peppa et al., 2023; Qi, Wang, et al., 2022; Qi, Zhan, et al., 2022) and is consistent with the modern trend of pursuing natural and healthy diets. However, the aromatic characteristics of T. serpyllum have received relatively less attention than its pharmacological properties in recent years. Volatile organic compounds (VOCs) are significant determinants of their fragrance attributes, given many volatile constituents, including terpenes, terpene alcohols, phenol derivatives, ketones, aldehydes, ethers, and esters (Sonmezdag et al., 2015). Simultaneously, it is worth noting that the composition and proportion of VOCs in thyme vary considerably owing to multiple factors such as variety, region, environment, harvesting time, harvesting location, and extraction methods (Pluhár et al., 2024; Shin & Ko, 2025; Tohidi et al., 2017). Therefore, it is crucial to elucidate the volatile profile of thyme from specific regions or sources under diverse application forms. Numerous studies have focused on the volatile components of T. serpyllum from various origins to clarify their volatile spectra and differences (Castell et al., 2023; Kim et al., 2020; Rivera-Pérez et al., 2022). Nevertheless, as far as we know, despite its wide distribution in central and northern China (Bian et al., 2007), there have been no systematic reports on its VOCs, which may impede its further application in food processing. Moreover, these related studies predominantly focus on the essential oil form of T. serpyllum, with a lack of information regarding the volatilization profiles of its culinary herb properties.

When compared with gas chromatography–mass spectrometry, which is widely utilized for the detection of volatile compounds, two-dimensional gas chromatography/time-of-flight mass spectrometry (GC × GC-TOFMS) technology exhibits superior peak capacity, enhanced sensitivity, and resolution, along with faster analysis speed (Zhao et al., 2022). These inherent advantages enable GC × GC-TOFMS to more effectively identify both volatile and semi-volatile VOCs while providing a more comprehensive volatile fingerprint profile. In recent years, this technology has been successfully applied to analyze various matrices, such as fruit (Zhang et al., 2023), green tea (Wang et al., 2024), and wine (He & Jeleń, 2025), etc… Concurrently, it has also been gradually employed in the research of volatile components within diverse spices and culinary herbs (Gu et al., 2024; Yang et al., 2025).

Herein, the present study aims to systematically investigate the volatile components of thyme sourced from four main producing regions in China and comprehensively elucidate the differences in VOCs among these samples. Briefly, headspace-solid-phase microextraction (HS-SPME) combined with GC × GC-TOFMS was employed to extract and analyze VOCs; multivariate statistical analysis was used to distinguish samples and select potential specific markers. This study is expected to contribute to a deeper global understanding of Chinese thyme's volatile profiles and clarify its characteristic components, thereby enhancing its potential applications in the food processing industry.

2. Materials and methods

2.1. Plant materials and sample preparation

Thyme was produced in central and northern China and collected during the flowering period in early summer. After drying in the sun, the above-ground parts (moisture content ≤4.5 %) were separated, vacuum packaged, and then stored in a cool and ventilated place (≤ 20 °C).

The above-preserved thyme was crushed and passed through a 40-mesh sieve and was named NX, WQ, JB, and NM, respectively. The geographical and more detailed information of the thyme samples are shown in Table S1. Each thyme powder sample was subjected to testing within 24 h. The preparation of each sample was replicated in triplicate.

2.2. Chemicals and reagents

The standard of n-alkanes (C7-C40) was purchased from Sigma-Aldrich (Shanghai, China). n-Alkanes (C6-C10) was obtained from AccuStandard (New Haven, CT, USA). 1,2-Dichlorobenzene (99.7 %) was obtained from Dr. E (Shanghai, China).

2.3. Volatile compounds extraction

VOCs were extracted using the HS-SPME method according to previous work with slight modifications (Qi, Wang, et al., 2022; Qi, Zhan, et al., 2022). Each sample (0.5 g of thyme powder with 1 mL of ultrapure water) was placed into a 15 mL headspace bottle, with 3 μL of 1,2-dichlorobenzene (100 ng/μL methanol solution) as an internal standard. After sealing, the vials were placed in a thermostatic controller set at 50 °C for 15 min (equilibration) and 25 min (extraction). Then, a 1-cm 50/30 μm stable Flexdivinylbenzene/Carboxen/polydimethylsiloxane (DVB/CAR/PDMS)-coated fiber (Supelco) was promptly inserted into the GC injector. The desorption process was carried out at 250 °C for 2 min in splitless mode.

2.4. Volatile compounds analyzation

VOCs were analyzed using a gas chromatograph (Agilent 7890B, USA) equipped with a LECO Pegasus 4D-C time-of-flight mass spectrometric detector (Saint Joseph, California, USA). Chromatographic separation was achieved using a two-dimensional column system. The first-dimensional (1D) column was an Rxi-5MS column (30 m × 250 μm × 0.25 μm, Shimadzu, Japan), while the second-dimensional (2D) column was an Rxi-17Sil MS column (2 m × 250 μm × 0.25 μm, Shimadzu, Japan). The detailed analysis conditions were adapted from the method described by Zhang et al. (2024).

Compounds with a matching score of over 700 were selected by comparison with the NIST17 database. Then, the established formula was used to calculate each volatile compound's retention index (RI) (Zhao et al., 2022). Finally, the calculated RI was compared with the reference literature to identify the compound. The result represented the amount of each compound in the sample relative to the internal standard.

2.5. Calculation of odor activity values

The relative odor activity value (ROAV) of VOC was obtained by the ratio of the relative concentration of each VOC to its odor threshold in water.

2.6. Statistical analysis

All experiments were performed in triplicate and results were presented on a mean ± standard deviation (SD) basis. A one-way analysis of variance (ANOVA) at a significance level of 5 % was done by the SPSS 27.0 software (SPSS Inc., Chicago, IL, USA). The sample distribution map was produced using Arcmap10.8. Principal component analysis (PCA), Partial least squares discriminant analysis (PLS-DA) models, and the important variables were predicted through MetaboAnalyst 6.0 (https://www.metaboanalyst.ca). Other pictures are generated by a CNSknowall platform (https://cnsknowall.com/#/HomePage).

3. Results and discussion

3.1. 3D plots of thyme samples by GC × GC-TOFMS

To visually and intuitively depict the distribution of VOCs in different thyme samples, three-dimensional chromatograms obtained from GC × GC-TOFMS were chosen (Fig. 1). Overall, the NX, WQ, JB, and NM samples exhibited visually similar profiles, likely due to their shared species and common origin in central and northern China. However, further observation revealed that NX and WQ demonstrated the highest degree of similarity, while the number of peaks in the NM sample was the highest. These findings suggest that despite belonging to the same species, thyme from distinct sources exhibits certain differences. Such variances could result from differences in growth environments, which may influence the biosynthesis and accumulation of VOCs in thyme plants.

Fig. 1.

Fig. 1

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3D-chromatograms of VOCs isolated from Chinese thyme samples from four regions using GC × GC-TOFMS. Column I axis represents the retention time of the compounds; column II refers to chemical polarity. NX, WQ, JB, and NM are the names of the four samples, respectively.

3.2. Analysis of the types and concentrations of VOCs in thyme samples

3.2.1. Overall analysis

Detailed information on VOCs is listed in Table S2. A total of 116 VOCs were identified and quantified in all the thyme samples, including 13 aldehydes, 14 ketones, 25 alcohols, 11 esters, 36 alkenes, 7 saturated hydrocarbons, 4 phones, 2 ethers, 2 quinones, 1 furan, and 1 acid. Overall, as shown in Fig. 2a, the number of compound types exhibited minimal variation among the four samples. However, significant differences were observed in their concentrations (Fig. 2b). Notably, in the NM sample, seven compound categories, namely ketones, alcohols, esters, alkenes, phones, quinones, and acid, displayed the highest content among the 11 categories. It could be attributed to environmental factors such as high latitude, extended illumination duration, and large diurnal temperature differences, which may enhance VOC accumulation. This observation is consistent with a study on the volatile components of thyme from different geographical distributions in Korea (Kim et al., 2020).

Fig. 2.

Fig. 2

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Composite figure of volatile profiles in thyme. a: Bubble pool plots on volatile species numbers, b: Stacked jade block diagram on content of volatile species, c: Relative proportion diagram on each volatile in its respective sample, d: Wayne diagram on the VOCs of four samples. NX, WQ, JB, and NM are the names of the four samples, respectively.

Fig. 2c illustrates the relative proportions of diverse volatile chemical classes in the four samples. Among the nine classes, alcohols represented the largest fraction, ranging from 30.57 % to 34.42 %. Phones (9.8–20.82 %), alkenes (11.83–15.35 %), and quinones (11.43–16.29 %) also constituted significantly, each exceeding 9.5 % of the total volatile. For the sake of statistical convenience, the “other compounds” category (6.93–10.77 %) was composed of three chemical classifications, namely saturated hydrocarbons, furan, and acid. Among them, a single volatile component, p-cymene, accounted for more than 95 % of this category. The relative contents of the remaining classes were as follows: ethers (5.33–9.57 %), esters (2.41–4.12 %), ketones (1.65–4.37 %), and aldehydes (1.15–1.85 %). Previous studies have shown that while the volatile profiles of Thymus essential oil vary significantly due to factors such as species, growth environment, geographical origin, and sample preparation methods, terpenes generally dominate, often accounting for over 90 % of the total composition (Hammoudi Halat et al., 2022; Sun et al., 2024). These findings are consistent with our results, further validating the consistency and reliability of our study.

3.2.2. Important volatiles analysis

As shown in Fig. 2d, among the 116 identified VOCs, 102 were shared across all four samples. Sample NM contained three unique compounds: (E)-2-Nonenal, o-Acetyltoluene, and p-Mentha-1(7),8(10)-dien-9-ol. Sample NX had one unique compound, octanoic acid methyl ester, while the other two samples lacked unique compounds. Additionally, the number of substances shared by two or three samples was relatively small. Whether these compounds can serve as distinguishing markers for different samples remains uncertain and warrants further analysis.

The 102 shared compounds represent the primary volatile profile characteristics of thyme from central and northern China. In sample NX, 10 compounds with a relative content exceeding 2000 μg/kg. Thymol had the highest content at 13.28 % of the total VOCs, followed by eucalyptol (11.95 %), thymoquinone (11.16 %), p-cymene (10.26 %), isoborneol (5.45 %), 1-octen-3-ol (4.87 %), carvacrol (3.89 %), carvacrol methyl ether (3.87 %), γ-terpinolene (3.64 %) and thymol methyl ether (3.34 %) in sequence. To emphasize the content of the main VOCs in the sample, based on the classification conventions for thyme essential oil, NM can be assigned to the thymol/eucalyptol/thymoquinone/p-cymene chemotype. Thymol, a prevalent compound in Thymus species, has long been a key factor in evaluating the commercial value of thyme. Recently, its excellent antibacterial, antioxidant, and antifungal properties, coupled with high safety, have led to extensive applications in the food industry (Sivaram et al., 2022). Furthermore, thymol enhances the flavor of culinary thyme during cooking, showing good synergistic effects in meat dishes (Qi, Wang, et al., 2022; Qi, Zhan, et al., 2022; Wu et al., 2023).

Samples WQ and NM shared high similarity in their main volatile compositions (content >2000 μg/kg). In sample WQ, thymoquinone (15.98 %) had the highest content, followed by eucalyptol (12.23 %), thymol (9.64 %), p-cymene (8.92 %), isoborneol (6.55 %), carvacrol (4.37 %), 1-octen-3-ol (4.22 %), carvacrol methyl ether (4.14 %) and caryophyllene (3.06 %). Similarly, WQ thyme can be classified as a thymoquinone/eucalyptol/thymol chemotype. The high thymoquinone content aligns with the findings by Alipour et al. (Alipour et al., 2025). Notably, such a high thymoquinone concentration is relatively rare in thyme, potentially because most studies focus on its essential oil rather than the plant itself. Thymoquinone exhibits excellent pharmacological effects, such as anticancer and anti-inflammatory properties (Tabassum et al., 2021). Its application in the food industry is mainly due to its antioxidant (Yildiz et al., 2020) and antimicrobial properties (Wang et al., 2021).

In sample JB, thymoquinone again had the highest content at 13.51 % of the total VOCs, while thymol (5.27 %) was significantly lower compared to the previous two samples. Other high-content compounds in JB included eucalyptol (11.79 %), p-cymene (9.99 %), 1-octen-3-ol (6.93 %), carvacrol methyl ether (6.49 %), isoborneol (5.25 %), carvacrol (4.19 %), caryophyllene (3.15 %) and thymol methyl ether (3.07 %). Based on these findings, JB thyme can be categorized as a thymoquinone/eucalyptol/p-cymene chemotype. Interestingly, eucalyptol ranked as the second-highest compound in JB, NX and WQ. Research by Manukyan (Manukyan, 2019) revealed that lower air temperature (15 °C) favored higher eucalyptol production. This could explain our results, as the annual average temperatures at the collection sites were mostly not higher than 15 °C, with relatively dry environmental conditions. Although some studies (Horvathova et al., 2014) indicated that eucalyptol has a lower antioxidant capacity compared to phenolic terpenes, it has demonstrated satisfactory antibacterial effects in various aspects (El-Kased & El-Kersh, 2022).

In the NM sample, thymoquinone also exhibited the highest concentration. Nevertheless, other VOCs with content exceeding 2000 μg/kg showed more significant differences compared to NX, WQ, and JB. NM contained 11 such high-content compounds, with their respective proportions as follows: thymoquinone (11.92 %), carvacrol (11.71 %), isoborneol (9.48 %), eucalyptol (9.25 %), thymol (8.83 %), p-cymene (6.61 %), carvacrol methyl ether (3.44 %), 1-octen-3-ol (2.69 %), p-mentha-1(7),8(10)-dien-9-ol (2.45 %), 2-bornanone (2.20 %) and linalool (2.10 %). Correspondingly, NM thyme can be classified as a thymoquinone/carvacrol/ isoborneol chemotype. Carvacrol, an isomer of thymol, shares highly similar bioactivities and aroma properties with thymol. Consequently, these two volatiles are frequently applied jointly in antibacterial, antifungal, and preservation applications, among others. This approach achieves a certain degree of synergy (Milos & Makota, 2012; Nieto, 2020; Sun et al., 2024) and reduces the toxicity associated with their individual use (Jalil et al., 2024).

Apart from highlighting the high levels and differences of these substances in thyme, it is equally intriguing to explore the biosynthesis pathway of important VOCs in thyme. This exploration will enhance our understanding of why these important compounds vary. As shown in Fig. 3, geranyl diphosphate can be cyclized to γ-terpinene, which is subsequently aromatized through a series of reactions to yield p-cymene. p-Cymene can then be hydroxylated to produce carvacrol or thymol, which can be further oxidized to thymoquinone (Botnick et al., 2012). This pathway has also been verified in chemical synthesis studies (Shin & Ko, 2025).

Fig. 3.

Fig. 3

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Possible biosynthetic pathways for several monoterpenes.

3.3. Main aroma active compounds analysis

Thyme has been utilized as a culinary herb since ancient times, a practice closely linked to its distinctive flavor profile. The unique flavors of thyme are primarily attributed to the aroma-active compounds among its numerous VOCs. Therefore, focusing on these substances is essential for evaluating the flavor quality of thyme. Generally, VOCs with ROAV greater than one are often considered to possess aroma activity and contribute significantly to the overall aroma. In this study, a total of 40 VOCs were identified as aroma-active compounds (ROAV >1), as detailed in Table 1. Among these, 1-octen-3-one, trans-β-ionone, 1-octen-3-ol, eucalyptol, linalool, isoborneol, bornyl acetate, β-myrcene, p-cymene, and eugenol exhibited particularly high ROAV values (exceeding 200). These compounds have also been recognized as effective odorants in other thyme varieties through gas chromatography-olfactometry analysis (Goodner et al., 2006; Sonmezdag et al., 2015). Previous studies have identified thymol and carvacrol as the primary odorants of thyme, as they exhibit the highest sensory intensity regardless of the extraction method used (E Dellacassa & Minteguiaga, 2023). However, in our study, their ROAV values were relatively low, ranging from 1 to 6. This discrepancy is not contradictory; the compound content was determined using a relatively quantitative method, and the calculated ROAV provides an approximation for assessing aroma contribution.

Table 1.

ROAVs and odor description of 40 odor-active compounds in thyme.

Codea Compounds Odor threshold (μg/kg)b Odor descriptionc ROAVd
NX WQ JB NM
N2 3-methyl-Butanal 1.1 Cocoa, almond 48.96 34.36 49.2 31.38
N3 2-methyl-Butanal 1 Malt 34 37.14 51.72 31.29
N4 Hexanal 5 Green, fresh, grass 3.62 5.04 4.5 5.01
N5 (E)-2-Hexenal 428.6 Green, leaf < 1 1.44 1.65 1.1
N6 (E,E)-2,4-Hexadienal 1.8 Green 5.48 12.22 14.69 10.42
N8 Benzeneacetaldehyde 6.3 Hawthorne, honey, sweet 25.03 25.22 17.02 13.98
N9 Nonanal 1.1 Citrus, green, citronella grass 48.42 33.03 41.6 30.15
N11 (E)-2-Nonenal 0.19 Cucumber, fat -⁎ – – 277.53
N12 Decanal 3 Soap, orange peel, tallow 8.41 9.91 8.45 2.53
N14 1-Octen-3-one 0.003 Intense earthy, metallic, mushroom-like 326,356.67 40,056.67 – –
N15 3-Octanone 21.4 Musty, mushroom, ketonic 24.08 22.89 43.33 58.29
N19 2-Bornanone 1360 Camphoreous, medicinal, mentholic < 1 < 1 < 1 1.6
N22 p-Acetyltoluene 21 Sweet, coumarin, cherry-like 2.04 1.6 2.65 2.79
N24 Carvone 27 Caraway 3.64 2.4 5.45 7.41
N26 Geranyl acetone 60 Magnolia, green 1.55 1.88 2.09 2.31
N27 trans-β-Ionone 0.007 Seaweed, violet, flower, raspberry 3585.71 5240 4577.14 4878.57
N28 1-Octen-3-ol 1.5 Fishy, earthy, mushroom 2260.88 1866.09 3358.45 1779.12
N29 3-Octanol 250 Mushroom, fat < 1 < 1 < 1 1.39
N30 Eucalyptol 4.6 Mint, sweet 1810.7 1764.25 1862.41 1993.34
N35 1-Nonen-3-ol 10 Oily, creamy, green, earthy, mushroom 12.08 11.55 16.97 5.93
N36 Linalool 6 Floral, sweet, woody, green 209.42 212.32 300.83 346.17
N39 Isoborneol 16 Must, camphor 237.54 271.76 238.41 587.39
N41 Terpinen-4-ol 1200 Turpentine, nutmeg, must < 1 < 1 < 1 1.4
N54 (Z)-3-Hexen-1-ol acetate 31 Fresh, green, sweet, fruity 1.69 1.01 1.4 < 1
N55 Benzoic acid methyl ester 73 chemical phenolic, cherry 3.06 1.8 2.1 1.44
N58 Methyl salicylate 40 Peppermint 5.31 3.19 3.48 4.28
N60 Bornyl acetate 0.056 Woody, pine, herbal, cedar, spice 6264.11 6897.5 3736.43 14,316.25
N66 α-Pinene 14 Fresh, earthy, sweet 16.43 13.32 14.64 20.21
N72 β-Myrcene 1.2 Balsamic, must, spice 305.53 216.18 437.25 625.78
N73 α-Phellandrene 40 Turpentine, mint, spice 5.63 5.1 8.03 8.23
N74 α- Terpinolen 200 Sweet, fresh 3.69 3.19 4.35 4.16
N76 trans-β-Ocimene 34 NI# 1.61 1.24 2.19 5.08
N77 γ-Terpinolene 1000 Gasoline, turpentine 2.53 1.85 1.47 1.92
N78 β-Pinene 140 Resinous, woody, pine < 1 < 1 < 1 6.3
N83 Caryophyllene 64 Sweet, woody 25.85 31.7 35.8 12.79
N95 Ionene 2 NI 51.66 77.22 72.34 123.77
N100 p-Cymene 5.01 Solvent, gasoline, citrus 1427.04 1180.95 1449.46 1307.36
N108 Thymol 1700 Thyme like 5.44 3.76 2.25 5.15
N109 Carvacrol 2290 Thyme like 1.18 1.27 1.33 5.07
N110 Eugenol 0.71 Clove, woody 519.54 351.14 308.65 348.17
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⁎

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a

Code same as Table S2.

b

Odor thresholds in water, data from the Compilation of Odor Threshold Levels in Air, Water and Other Media (second enlarged and revised edition), Oliemans Punter & Partners BV, The Netherlands.

d

Odor activity value.

3.4. Multivariate statistical analysis

To further clarify the disparities in volatiles among thyme samples from different origins, a multivariate statistical analysis was conducted on the identified compounds. Unsupervised PCA effectively revealed the natural grouping patterns among the samples. Additionally, the variable importance projection (VIP) score analysis applied to the PLS-DA model was employed to identify significant VOCs. According to Fig. 4a, the first two principal components cumulatively explained 89.4 % (PC1 = 66.5 %, PC2 = 22.9 %) of the total variance, indicating that these two dimensions effectively captured the overall volatile characteristics of the samples. Samples NX, WQ, and JB were relatively clustered, with a certain degree of overlap, suggesting that the volatile profiles of thyme from these three habitats are similar. In contrast, NM samples were clustered separately and distantly from the others, signifying that they possess a unique volatile profile.

Fig. 4.

Fig. 4

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Multivariate statistical analysis of volatile profiles. a: The scores plot of PCA based on GC × GC-TOFMS, b: Important volatile compounds (VIP > 1.0) identified by PLS-DA based on GC × GC-TOFMS, c: The scores plot of PCA based on compounds with ROAV >1, d: Important volatile compounds (VIP > 1.0) identified by PLS-DA based on compounds with ROAV >1. NX, WQ, JB, and NM are the names of the four samples, respectively. The code of every compound is the same as in Table S2.

Subsequently, as evident from Fig. 4b, a total of 14 volatile compounds with VIP >1 were analyzed in the thyme samples, including carvacrol (N109), thymol (N108), carvacrol methyl ether (N113), 1-octen-3-ol (N28), isoborneol (N39), tridecane (N103), linalool (N36), eucalyptol (N30), 3-octanone (N15), p-mentha-1(7),8(10)-dien-9-ol (N47), bornanone (N19), γ-terpinolene (N77), thymoquinone (N114) and trans-pinocarvyl acetate (N61). PC1(70.3 %)-PC2 (25 %) explained 95.3 % of the total variance of aromatic compounds with an ROAV >1(Fig. 4c). The clustering patterns of the four samples were highly consistent with those observed previously, thereby validating the accuracy of each other's grouping. Differences in aroma-active compounds can more effectively reflect the sensory aroma characteristics of thyme. This may serve as the basis for selecting different types of thyme according to the compatible objects in food processing. According to Fig. 4d, compounds with VIP >1, such as linalool (N36), isoborneol (N39), 1-octen-3-ol (N28), thymol (N108), and carvacrol (N109) were identified as the key potential markers for discerning aroma differences among thyme samples from four distinct regions.

4. Conclusions

In this study, GC × GC-TOFMS was employed to comprehensively characterize the volatile profiles of thyme sourced from four major geographical regions in central and northern China and precisely determine the differences among them. The results revealed that alcohols, alkenes, and phenols were predominant in thyme, with most of these compounds belonging to terpenes. Notably, all samples were found to be rich in eight key compounds: thymol, eucalyptol, thymoquinone, p-cymene, isoborneol, 1-octen-3-ol, carvacrol, and Carvacrol methyl ether. Based on the classification approach of thyme essential oil, thyme samples were classified into distinct chemotypes. Subsequently, odor-active compounds were identified through ROAV calculations. Multivariate statistical analysis methods were then utilized to discriminate between samples and select key volatile markers. PCA results revealed that NX, WQ, and JB thyme exhibited similar volatile and aroma profiles, while NM thyme showed significant differences. The establishment of the PLS-DA model identified 14 with a variable importance in projection (VIP) > 1 as markers of thyme volatility and 5 VOCs (VIP > 1) as markers of odor activity. These findings clarify the chemical basis underlying the culinary properties of thyme (Thymus serpyllum L.), providing a solid foundation for evaluating its flavor quality in major Chinese production areas. Furthermore, this research offers valuable insights into expanding the global diversity of thyme volatiles and enhancing its applications in food processing.

CRediT authorship contribution statement

Shasha Qi: Writing – original draft, Methodology, Investigation, Formal analysis, Data curation, Conceptualization. Chengjie Hou: Writing – review & editing, Validation, Methodology, Formal analysis, Data curation. Honglei Tian: Resources, Project administration, Funding acquisition. Ping Zhan: Writing – review & editing, Validation, Supervision, Resources, Project administration, Funding acquisition, Conceptualization. Bin Qiu: Writing – review & editing, Supervision, Resources, Project administration, Funding acquisition.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

This research was conducted with support from National Natural Science Foundation of China (32072343 and 32272459), Special Support Plan of Shaanxi Province (TZ0432), Science and Technology Innovation Team of Shaanxi Province (2022TD-14), Qin Chuangyuan “Scientist and Engineer” Team Construction (2024QCY-KXJ-078), and Agricultural Science and Technology Innovation Project of Shandong Academy of Agricultural Sciences (CXGC2025F08).

Footnotes

Appendix A

Supplementary data to this article can be found online at https://doi.org/10.1016/j.fochx.2025.102590.

Contributor Information

Ping Zhan, Email: zhanping0993@126.com.

Bin Qiu, Email: qiubin2009@live.cn.

Appendix A. Supplementary data

Supplementary material 1

mmc1.docx (17KB, docx)

Supplementary material 2

mmc2.docx (51KB, docx)

Data availability

Data will be made available on request.

References

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Supplementary material 1

mmc1.docx (17KB, docx)

Supplementary material 2

mmc2.docx (51KB, docx)

Data Availability Statement

Data will be made available on request.

Articles from Food Chemistry: X are provided here courtesy of Elsevier

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