Next Article in Journal
Comparison of Waterlogging Tolerance of Three Peach Rootstock Seedlings Based on Physiological, Anatomical and Ultra-Structural Changes
Previous Article in Journal
Potential Use of Superabsorbent Polymer on Drought-Stressed Processing Tomato (Solanum lycopersicum L.) in a Mediterranean Climate
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Horticulturae
Volume 8
Issue 8
10.3390/horticulturae8080719
horticulturae-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
Correction published on 20 February 2023, see Horticulturae 2023, 9(2), 286.
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Dynamic Changes on Floral Aroma Composition of the Three Species from Tilia at Different Flowering Stages

by
Wenqin Bao
1 and
Yongbao Shen
1,2,3,*
1
College of Forestry, Nanjing Forestry University, Nanjing 210037, China
2
Collaborative Innovation Center of Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing 210037, China
3
Southern Tree Seed Inspection Center, National Forestry and Grassland Administration, Nanjing 210037, China
*
Author to whom correspondence should be addressed.
Horticulturae 2022, 8(8), 719; https://doi.org/10.3390/horticulturae8080719
Submission received: 13 July 2022 / Revised: 30 July 2022 / Accepted: 3 August 2022 / Published: 10 August 2022 / Corrected: 20 February 2023
(This article belongs to the Section Developmental Physiology, Biochemistry, and Molecular Biology)
Browse Figures
Versions Notes

Abstract

:
The floral aroma, sometimes known as an incorporeal gift of flowers, is one of the primary ornamental features of plants. Flowers of genus Tilia are fragrant and have great value for development and utilization. In this study, for the first time, headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC-MS) combined with chemometrics were used to analyze the dynamic variations of scent components of the three species from Tilia at different flowering stages. A total of 70 volatile components were detected, 43 aroma compounds were identified. The UpSet diagram displayed great variations in the amount of aroma components at various flowering stages for each species. Partial least squares discriminant analysis (PLS-DA) indicated the proximity of aroma composition characteristics and the significant components that can distinguish the three species from one another. Variable importance projection values (VIP) along with the Kruskal-Wallis non-parametric analysis were performed to identify 14 crucial aroma components, such as α-Farnesene, D-Limonene, Germacrene D, Linalool, etc. In the end, we discovered that, in sharp contrast to Tilia miqueliana Maxim., Tilia cordata Mill. may have a closely related phylogenetic relationship with Tilia tomentosa Moench. by evaluating the aroma similarity rates.

1. Introduction

The aroma released by plant organs is constituted of secondary metabolites which are a combination of volatile chemicals with a very low molecular weight [1]. Floral fragrance, one of a plant’s most significant ornamental characteristics, is especially crucial in influencing pollinator behavior and controlling flowering plant reproduction [2,3,4]. The various volatile substances released by plants during their growth and development result in characteristic fragrances [5] that attract specific insects for pollination [6]. Bees can use floral cues to hunt for and identify food sources [7], and the mass emission of floral material may encourage frequent activity of the primary pollinators [8]. In addition, researches demonstrated that floral fragrances can also have a synergistic effect on the visual presentation of flowers [9]. For instance, an orchid blossom can imitate the appearance and scent of a species of female bees, causing the males to attempt mating and resulting in pollination [9,10].
The floral aroma is closely associated with human life as well. Moreover, specific receptors in the human olfactory system may detect volatile fragrant substances, which can then produce pleasurable sensations [11]. Currently, more than 1700 volatile aroma substances have been identified in plants [12]. They are mainly classified as aldehydes, ketones, alcohols, esters, terpenes, alkanes, acids, ethers and aromatic compounds [13]. The range of scent compounds in various plants can reach tens or even hundreds, as evidenced by the 39 fragrant components found in Rosa odorata var. erubescens petals [14] and 89 fragrant compounds found in the entire flowers of Camellia japonica ‘Kramer’s Supreme’ [15]. Utilizing extraction from plants and artificial synthesis, floral substances have been widely used in spices, essential oils, cosmetics, food additives, air fresheners and other fields.
Trees of genus Tilia (Tiliaceae family) are fragrant, deciduous and have an abundance of flowers, making them good attractive trees for landscaping [16]. The dried inflorescences of Tilia cordata Mill. are processed into flower tea in Europe that can effectively relieve colds and coughs brought on by colds [17]. Volatile organic compounds(VOCs) in its essential oil such as α-pinene, γ-terpinene and linalool can be used for alleviating anxiety [18]. Studies on volatile organic compounds (VOCs) from Tilia species have recently concentrated on essential oils of inflorescences, bracts and leaves [19,20], but researches comparing the floral composition of different species in Tilia are scarce.
In this study, flowers of Tilia cordata Mill., Tilia tomentosa Moench. and Tilia miqueliana Maxim. in different periods were selected as experimental materials, which are adaptable and growing well in eastern and northern China. They deserve more promotion because they are superior nectar and fragrance producing plants. Therefore, headspace-solid phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC-MS) combined with chemometrics were used to explore the composition and dynamic change of floral aroma, which might also provide a scientific basis for in-depth research and utilization for the flowers of Tilia, as well as having crucial implications for plant phylogeny development.

2. Materials and Methods

2.1. Materials and Chemicals

Flowers of T. cordata, T. miqueliana and T. tomentosa were collected in Guangzhuang, Zhuzhen, Liuhe District, Nanjing (32.51° N, 118.71° E), on 27 May, 29 May and 1 June 2021 between 09:00 and 10:00. For each species, three excellent strains of similar growth and profuse flowering were selected, and inflorescences of various flowering stages were picked from the upper, middle and lower regions of the canopy in the north, south, east and west directions, packed in self-sealing polybags, placed in ice cases for temporary storage and rapidly brought back to the laboratory. According to the degree of flower opening, the flowers were divided into bud stage (a), half-open stage (b) and blooming stage (c) (Figure 1). The basic situation of the selected materials can be shown in Table 1. The C7-C40 (HJ894-2017) standards were purchased from Tan-Mo technology Co., Ltd. Changzhou, China.

2.2. Detection Methods

Randomly selected flower samples at various stages were weighed to a mass of around 0.300 g, placed in a 20 mL headspace glass vial with a screw-top, sealed and capped, then heated to 45 °C in an electric thermostatic water bath for 30 min. Subsequently, an aged 65 µm PDMS/DVB SPME fiber inserted into the vial through a PTFE spacer was placed 1 cm above the flowers samples and extracted in the headspace for 26 min. Three independent replicates were performed for each stage of each sample.
After extraction, the fiber was withdrawn and inserted into the GC inlet for desorption (3 min, 240 °C), and volatile components of the samples were analyzed using the Thermo Fisher Scientific Trace ISQ GC-MS system [21]. A DB-5MS flexible quartz capillary column (30 m × 0.25 mm × 0.25 µm) was used; the carrier gas was high purity helium He (purity > 99.999%) at a flow rate of 1 mL/min [22]; the ramp-up procedure was 60 °C for 1 min, 10 °C/min to 120 °C, 5 °C/min to 160 °C for 2 min, and 10 °C/min to 240 °C for 1 min. The mass spectrometer was set to ionization mode EI, ion ionization energy 70 eV, mass scan ranged 33-450 amu and solvent delay time was 0.5 min. The operating system was Xcalibur software (v. 2.2, Thermo Fisher Scientific, Waltham, MA, USA) [21].

2.3. Identification and Quantification of Aroma Components

Aroma components were identified by retention index (RI) and mass spectrum compared with the reference standards obtained from the NIST Chemistry WebBook (https://webbook.nist.gov/chemistry/), accessed on 15 August 2021. When reference standards were no available, identification was taken by comparing mass spectra with those in the NIST Chemistry WebBook and RI published in the literatures. Calculation of RIs according to the retention times of C7-C40 alkanes separated under the GC-MS conditions mentioned above, utilizing the formula below (Equation (1)). The relative content (%) of aroma compounds in the sample was calculated by normalization of the total ion peak area in the chromatography [23]. The average of the three measurements was taken as the relative content of each compound [24].
RI = 100 n + 100 ( t x − t n ) t n + 1 − t n
where tx is the retention time (min) for compound x to be detected; tn is the retention time (min) of n-alkanes Cn; tn+1 is the retention time (min) of n-alkanes Cn+1; n is the number of carbon atoms in an unknown compound.

2.4. Calculation of Aroma Similarity Rates

We refered to the cosine similarity method [25] to calculate the aroma similarity rates of the three species from Tilia at different flowering stages, using the relative content (%) of aroma components (Equation (2)).
S ( A , B ) = cos θ = ∑ i = 1 n a i b i ∑ i = 1 n a i 2 ∑ i = 1 n b i 2
where ai and bi are the relative content of each aroma component in samples A and B.

2.5. Statistical Analysis

Excel (Microsoft Office Standard 2019, Microsoft Corporation, Redmond, WA, USA) was used to calculate the relative content (%) of each aroma component; UpSet diagrams were performed by using the OmicShare tools, a free online platform for data analysis (https://www.omicshare.com/tools), accessed on 5 June 2022; SPSS Statistics 26.0 software (IBM, Armonk, NY, USA) was used for the Kruskal–Wallis non-parametric test; SIMCA 14.1.0.2047 software (Umetrics, Umeå, Sweden) was used for the PLS-DA model, drawing score scatter plot and loading scatter plot, calculation of VIP value and permutation test; Matlab R2021a software (Math Works Corporation, Natick, MA, USA) was used for calculating the aroma similarity rates; aroma characteristics were collected from ‘The Good Scents’ company network database (www.thegoodscentscompany.com), accessed on 5 January 2022. OriginPro 2021 software (OriginLab Corporation at Northampton, MA, USA) was applied to draw heat map.

3. Results

3.1. Dynamic Changes of Floral Aroma at Different Stages

A total of 70 volatile compounds were detected in the three species of Tilia at different flowering stages, 43 aroma compounds were identified, 27 components remained uncertainly labelled as NI (Table 2). At different stages, there were 28, 28 and 26 components from T. cordata; 17, 20 and 17 from T. tomentosa; 22, 24 and 26 from T. miqueliana, respectively. Figure 2 demonstrated that the three species had a wide range of floral aroma compound quantities, and the relative content of terpenoids were dominating in the identified aroma components.
The primary aroma components released from T. cordata flowers are β-Ocimene, Germacrene D and Linalool. With the gradual blossoming, the relative content of β-Ocimene increased from 29.47% to 34.44% and lastly to 36.90% at different flowering stages; Linalool was 14.74%, 18.73% and 21.30%. Both β-Ocimene and Linalool showed an increasing trend. The relative content of Germacrene D, on the other hand, kept on diminishing from 22.30% to 18.67% and 16.83%.
β-Ocimene, D-Limonene, and Cyclohexene, 1-methyl-4-(1-methylethylidene)- were predominant fragrant substances of T. tomentosa. However, β-Ocimene and Cyclohexene,1-methyl-4-(1-methylethylidene)- did not vary significantly at the three flowering stages. The relative content of D-Limonene increased marginally, containing 19.88% and 19.04% at the bud and half-open stages, respectively, and going up to 22.55% during the blooming period.
The dominating aroma substance of T. miqueliana was α-Farnesene, with an inconspicuous increase in relative content at the three flowering stages, containing 58.63%, 59.35% and 60.37%, respectively.
The amount of fragrance compounds present in the three species at various flowering phases varied greatly, as shown by Table 2 and Figure 2.
At different flowering phases, T. cordata had 25 aroma substances in common at the three flowering stages; the bud stage shared 2 exclusive substance with the half-open stages, and only 1 particular component in the half-open stage and the blooming stage. T. tomentosa contained 14 aroma substances in common across the three flowering stages, the half-open stage shared 2 unique components with the bud stage and the blooming stage, respectively. T. miqueliana possessed 20 fragrant compounds that appeared in all three flowering stages, the bud stage shared 1 particular component with the half-open stage and the blooming stage, respectively. Some aroma components were only available during special periods, for example, there was only 1 exclusive component in the bud stage of T. cordata; For T. tomentosa, the bud stage and the blooming stage had 1 exclusive component, respectively, and 2 exclusive components appeared in the half-open stage; For T. miqueliana, 3 exclusive components in the half-open stage, 5 exclusive components in the blooming stage. Moreover, Tridecane was detected at the half-open stage from T. miqueliana, which has been proven to be one of the ingredients of plant waxes that covers not only leaves but also other plant organs like flowers and fruits [26]. Thus, it is possible to assume that wax layers are present on the floral surface of T. miqueliana.
Additionally, T. cordata shared 2 aroma components with T. tomentosa: β-Ocimene and Germacrene D, while only sharing Lilac aldehyde (isomer II) with T. miqueliana. 4 aroma compounds including D-Limonene, α-farnesene, Phenylethyl Alcohol, and Acetic acid, 2-phenylethyl ester were discovered in both T. miqueliana and T. tomentosa. However, only 2 components in commom among the 3 species. Considering β-Ocimene, Germacrene D, D-Limonene and α-farnesene diversely all exhibited high relative percentages diversely among the three species, it may be inferred that they are the typical scent chemicals in flowers of Tilia.

3.2. Chemometric Analysis of Floral Aroma Components

Partial least squares discriminant analysis (PLS-DA) is a multivariate statistical analysis method for discriminant analysis [27] that enhances group differences by using information about class members provided by the auxiliary matrix in the form of codes [28]. In order to ascertain whether there were variations in the quantities of floral scent components in the three species of Tilia, a PLS-DA model was employed in this study to interpret the data collected by GC-MS. In all, 9 groups of samples with positive contribution rates were used as independent variables in the mathematical model. After fitting, the PLS-DA model contained 2 principal components (PC1 = 0.598 and PC2 = 0.345), and the fitting parameters R2X=0.942, R2Y=0.999, and Q2=0.997, with a cumulative explanatory degree of 94.3%, can comprehensively reflect the original information of floral fragrance for the three species of Tilia, which has good predictability.
The scent features of each species at various flowering phases were similar in the score scatter plot (Figure 3a) with good aggregation, and the data points of the three species were distributed within a 95% confidence interval. Samples of T. miqueliana were separated in the t1 direction, T. tomentosa and T. cordata were separated in the t2 direction. In the loading scatter plot (Figure 3b), the far distance of component points from the centre, the greater the weight value for the sample variability and give rise to the greater effect on sample dispersion. The contributions of aroma components to PC1 and PC2 raised as the distance from the coordinate axis’ origin increased and vice versa. Great contributions from β-Ocimene (P7) and α-Farnesene (P39) offered significant contributions to PC1: 0.463 and −0.620, respectively; while D-Limonene (P5), Germacrene D (P37) and Linalool (P14) offered significant contributions to PC2: 0.443, −0.363 and −0.364, respectively.
Overfitting is a well-known issue with the PLS-DA model, thus a permutation test was used to validate the model’s accuracy (Figure 4). The three species included in the model were subjected to a permutation test, with 200 replacements. The results were shown as follows: R2 = (0.0, 0.23) and Q2 = (0.0, −0.307) of T. cordata; R2 = (0.0, 0.277) and Q2 = (0.0, −0.299) of T. miqueliana; R2 = (0.0, 0.244) and Q2 = (0.0, −0.283) of T. tomentosa. In general, the intercept values of prediction points of the fragrant compositions for the three species were all lower than the original model (Q2 < R2), and the y-intercept of Q2 is less than 0, indicating that the PLS-DA model was not over-fitting and the analysis results were reliable.

3.3. Determination of Crucial Components

The contribution of different variables to the overall classification can be explained by the variable importance for the projection (VIP) [29]. The more notable the difference in variables across groups, the higher the VIP value is. When the VIP value is higher than 1.00, the corresponding variable can be defined as the key variable of the discriminant model. To make the analysis more accurate, a Kruskal–Wallis nonparametric test was used to further examine the compounds with VIP greater than 1.00, along with the P-values less than 0.05 as the selection criteria. 14 crucial components were eventually identified (Table 3), which were consistent with the results of the PLS-DA loading scatter plot, indicating statistical variations of the three species. Moreover, the olfactory properties of the 14 crucial components are varied, primarily manifesting as citrus, woody, floral.
A heat map was utilized for the analysis to visualize the distribution of the 14 crucial components at the different flowering stages of the three species, as shown in Figure 5. The three species could be totally separated from one another by the 14 crucial components, which had varying distributions and high abundances.

3.4. Aroma Similarity Rates

The statistics between T. cordata and T. tomentosa in terms of aroma similarity rates were higher than 0.570 in the three flowering stages, which means that they might have a very close phylogenetic relationship (Table 4). In contrast, the aroma similarity rates between T. miqueliana and T. cordata or T. tomentosa in the three flowering stages were all lower than 0.1, revealed that they were basically dissimilar. Nevertheless, the aroma similarity rates of each species at various flowering stages were all greater than 0.970, suggesting that the overall olfactory characteristics fluctuated a little. It may be deduced that the primary floral scent constituents of the three species were synthesized during the bud stage, with only a minor amount of decomposition and transformation occurring during the half-open stage and the blooming stage.

4. Discussion

The fundamental parenchyma of the floral organ or specialized glandular epidermal cells are usually the places plant fragrance is produced [30]. The formation and accumulation of flower scent are influenced by a variety of elements, but mostly depend on genetic effect. However, the release of VOCs from floral organs does not hinge on the volatilization of its components, but on the selective release of these components into the membrane structure of the synthesis zone by specific carriers [31,32]. The primary floral aroma components in T. cordata and T. tomentosa flowers in this study vary markedly from the VOCs of their inflorescence essential oils in previous research [17,18]. The synthesis and emission of VOCs may be affected by different genotypes and stand conditions together. Studies have also found that the main components of floral volatiles vary greatly even among species of proximal origin [12], and that the primary components of flower volatiles are irregular even within the same genus [33].
Moreover, volatile substances frequently undergo dynamic changes throughout flowers’ blossom. Variables were found in the quantity and relative content of fragrance compounds in the three species of Tilia at various flowering periods, which coincided with the analysis of volatile components at different flowering stages in other fragrant plants, such as Hosta [34], Polianthes tuberosa [35], Prunus mume [36], and Osmanthus fragrans [37]. According to the volatile components, which are controlled by the actions of related biosynthases within the flowers, reactions like dehydrogenation, deoxygenation, catalysis and degradation are carried out from opening to decay in order to accumulate or generate new compounds to take part in body metabolism [38]. This ultimately results in variations in categories and relative contents of the volatile components released over time.
The emissions of major aroma components in the three species of Tilia were basically in the blooming stage as seen by the results. Previous research on floral aroma production indicates that some plants, such as Acacia Cyclops was in the bud stage [39]; however, Osmanthus fragrans ‘Boye Jingui’ was in their early bloom stage [40]. As a result, the floral aroma of different plants has varying compositions and release patterns, which could have an impact on the pollinators’ attraction to certain plants [41]. Environmental influences and the internal biological clock both have an impact on the pattern of scent emission [42]. In the diurnal variation of the full blooming stage of Dendrobium Chrysotoxum, the release volume of α-pinene and β-Ocimene reached a peak at 11:00 and 14:00, respectively [43]. Under various light intensity treatments, there were noticeable variations in the content and concentration of fragrance from Oncidium Sharry Baby ‘Sweet Fragrance’ [44]. Additionally, high temperature inhibited the blooming and aroma emission of Jasminum sambac Ait, while high humidity and a plentiful water supply can encourage fragrance production [45].
Several components of the 14 key floral scent components determined in this research can not only participate in plant growth and development, environmental response and other physiological processes, but also have biological activities. For instance, α-Farnesene plays an important role in plant defense [46] and can be used as a precursor for modern biofuel farnesane production [47]. D-Limonene can be utilized as a synergist for hygienic pesticides [48] and as a raw ingredient for artificial neroli and lemon oils, both of which have phlegmatic and anticancer properties [49]. Linalool contains antibacterial, anti-inflammatory and anti-infective properties and is frequently utilized in the synthesis of essential oils [50]. Escherichia coli and Bacillus subtilis are significantly inhibited by the antimicrobial activity of Germacrene D [51]. They provide a variety of fragrant qualities that are beneficial to human mood and health. It is obvious that the three species of Tilia are highly valuable in terms of commerce, environment, and medicine. Consequently, based on the results of this research, we recommend more investigation into the metabolic pathways, regulatory genes, associated enzymatic activities of floral components and subcellular localization of Tilia.

5. Conclusions

In this study, HS-SPME/GC-MS combined with chemometrics were used to analyze the aroma components for the three species of Tilia at different flowering stages. The majority of the scent compounds were detectable at each stage, however, several fragrance components were only present at certain intervals. Phylogeny and coevolutionary history have an impact on the variation in volatile components found in flowers. The three species belonging to the same genus show different degrees of similarity to each other regarding aroma. It’s also intriguing that, in sharp contrast to T. miqueliana, T. cordata and T. tomentosa have a tighter relationship, implying that the three species have diverse proximity of ecological origins.

Author Contributions

Conceptualization, W.B.; methodology, W.B.; software, W.B.; validation, W.B.; formal analysis, W.B.; investigation, W.B.; data curation, W.B.; writing-original draft preparation, W.B.; writing-review and editing, W.B.; visualization, W.B.; resources, Y.S.; supervision, Y.S.; project administration, Y.S.; funding acquisition, Y.S. All authors have read and agreed to the published version of the manuscript.

Funding

This work was supported by Innovation and popularization of forestry technology in Jiangsu Province, “Long-term scientific research base for the in vitro conservation of rare native tree germplasm resources in Jiangsu Province: LYKJ[2021]03”.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Not applicable.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Pichersky, E.; Gang, D.R. Genetics and Biochemistry of Secondary Metabolites in Plants: An Evolutionary Perspective. Trends Plant Sci. 2000, 5, 439–445. [Google Scholar] [CrossRef]
  2. Jurgens, A.; Witt, T.; Gottsberger, G. Flower Aroma Composition in Dianthus and Saponaria Species (Caryophyllaceae) and Its Relevance for Pollination Biology and Taxonomy. Biochem. Syst. Ecol. 2003, 31, 345–357. [Google Scholar] [CrossRef]
  3. Klahre, U.; Gurba, A.; Hermann, K.; Saxenhofer, M.; Bossolini, E.; Guerin, P.M.; Kuhlemeier, C. Pollinator Choice in Petunia Depends on Two Major Genetic Loci for Floral Aroma Production. Curr. Biol. 2011, 21, 730–739. [Google Scholar] [CrossRef] [PubMed]
  4. Raguso, R.A.; Levin, R.A.; Foose, S.E.; Holmberg, M.W.; McDade, L.A. Fragrance Chemistry, Nocturnal Rhythms and Pollination “Syndromes” in Nicotiana. Phytochemistry 2003, 63, 265–284. [Google Scholar] [CrossRef]
  5. Jin, L.; Yang, M.L.; Wang, J.Z. Research Progress of Floral Metabolism in Plants. Mod. Gard. 2019, 5, 8–10. [Google Scholar] [CrossRef]
  6. Piechulla, B.; Pott, M.B. Plant Scents-Mediators of Inter- and Intraorganismic Communication. Planta 2003, 217, 687–689. [Google Scholar] [CrossRef]
  7. Burger, H.; Dotterl, S.; Ayasse, M. Host-Plant Finding and Recognition by Visual and Olfactory Floral Cues in an Oligolectic Bee. Funct. Ecol. 2010, 24, 1234–1240. [Google Scholar] [CrossRef]
  8. Hoballah, M.E.; Stuurman, J.; Turlings, T.C.J.; Guerin, P.M.; Connetable, S.; Kuhlemeier, C. The Composition and Timing of Flower Odour Emission by Wild Petunia axillaris Coincide with the Antennal Perception and Nocturnal Activity of the Pollinator Manduca sexta. Planta 2005, 222, 141–150. [Google Scholar] [CrossRef]
  9. Pichersky, E.; Gershenzon, J. The Formation and Function of Plant Volatiles: Perfumes for Pollinator Attraction and Defense. Curr. Opin. Plant Biol. 2002, 5, 237–243. [Google Scholar] [CrossRef]
  10. Schiestl, F.P. On the Success of a Swindle: Pollination by Deception in Orchids. Naturwissenschaften 2005, 92, 255–264. [Google Scholar] [CrossRef]
  11. Miao, M.M.; Chen, J.C.; Zhang, Z.D. Physiology, Genetics and Manipulation of Flower Fragrance. Mol. Plant Breed. 2007, 5, 67–74. [Google Scholar] [CrossRef]
  12. Knudsen, J.T.; Eriksson, R.; Gershenzon, J.; Stahl, B. Diversity and Distribution of Floral Aroma. Bot. Rev. 2006, 72, 1–120. [Google Scholar] [CrossRef]
  13. Qiu, Y.X.; Chen, J.Y.; Tian, M.J.; Xie, Z.H.; Chen, X.M.; Zhan, J.S. Study on the Volatile Components in Flowers of 12 Camellia Species. For. Res. 2015, 28, 358–364. [Google Scholar] [CrossRef]
  14. Wang, H.; Zhu, J.J.; Feng, S.C. The Aroma Components Analysis of Rosa odorata var. Erubescens and Its Application in Urban Gardens. Chin. Agric. Sci. Bull. 2021, 37, 71–75. [Google Scholar] [CrossRef]
  15. Fan, Z.Q.; Li, J.Y.; Li, X.L.; Yin, H.F. Analysis on the Aroma Components of Different Floral Organs of Aromatic Camellia‘Kramer’s Supreme’Based on HS-SPME/GC-MS. Bull. Bot. Res. 2014, 34, 136–142. [Google Scholar] [CrossRef]
  16. Yi, H.Z.; Chuanyuan, D. Study on Anatomical Characters and Garden Application of Some Species in Tilia L. Master’s Thesis, Fujian Agriculture and Forestry University, Fuzhou, China, April 2011. [Google Scholar]
  17. Fitsiou, I.; Tzakou, O.; Hancianu, M.; Poiata, A. Volatile Constituents and Antimicrobial Activity of Tilia tomentosa Moench and Tilia Cordata Miller Oils. J. Essent. Oil Res. 2007, 19, 183–185. [Google Scholar] [CrossRef]
  18. Zori, M.; Kosti, S.; Kebert, M.; Bozin, B.; Orlovi, S. Volatile organic compounds of Tilia cordata Mill. from Serbia, in terms of ecosystem services. Topola 2020, 206, 21–28. [Google Scholar] [CrossRef]
  19. Toker, G.; Baser, K.H.C.; Kürkçüoglu, M. and Özek, T. The composition of essential oils from Tilia L. species growing in Turkey. J. Essent. Oil Res. 1999, 11, 369–374. [Google Scholar] [CrossRef]
  20. Kowalski, R.; Baj, T.; Kalwa, K.; Kowalska, G.; Sujka, M. Essential oil composition of Tilia cordata flowers. J. Essent. Oil Bear. Plants 2017, 20, 1137–1142. [Google Scholar] [CrossRef]
  21. Ji, X. Comparative Investigation of Volatile Components and Bioactive Compounds in Beers by Multivariate Analysis. Flavour Fragr. J. 2021, 36, 374–383. [Google Scholar] [CrossRef]
  22. Zhang, Z.Q.; Zang, M.W.; Zhang, K.H.; Wang, S.W.; Li, D.; Li, X.M. Effects of Phospholipids and Reheating Treatment on Volatile Compounds in Phospholipid-Xylose-Cysteine Reaction Systems. Food Res. Int. 2021, 139, 109918. [Google Scholar] [CrossRef] [PubMed]
  23. Oliveira, L.C.; Nunes, C.E.P.; Brito, V.L.G.; Caetano, A.P.S. Floral Oil Production in a Family Dominated by Pollen Flowers: The Case of Macairea radula (Melastomataceae). Flora 2022, 288, 152008. [Google Scholar] [CrossRef]
  24. Xu, W.; Qixiang, Z. Study of Emission Patterns of Characteristic Volatiles Identified in Lagerstroemia caudata Flowers and Key Generelated to MEP Pathway. Ph.D. Dissertation, Beijing Forestry University, Beijing, China, June 2019. [Google Scholar] [CrossRef]
  25. Wang, H.F.; Li, M.J.; Liu, Z.H.; Wang, Z.H.; Shi, Z.P. The Aroma Change of Fu Brick Tea during Its Blooming. J. Tea Sci. 1991, 81–86. [Google Scholar] [CrossRef]
  26. Allebone, J.; Hamilton, R.J.; Knights, B.A.; Middleditch, B.S.; Power, D.M. Cuticular Leaf Waxes Part II. Chenopodium album L. and Lolium perenne L. Chem. Phys. Lipids 1970, 4, 37–46. [Google Scholar] [CrossRef]
  27. Hirri, A.; Bassbasi, M.; Platikanov, S.; Tauler, R.; Oussama, A. FTIR Spectroscopy and PLS-DA Classification and Prediction of Four Commercial Grade Virgin Olive Oils from Morocco. Food Anal. Methods 2016, 9, 974–981. [Google Scholar] [CrossRef]
  28. Yang, Z.; Ren, H.Q.; Jiang, Z.H. Study on Discriminant Analysis of Wood Bio-Decay by PLS-DA Method. Spectrosc. Spectr. Anal. 2008, 4, 793–796. [Google Scholar] [CrossRef]
  29. Liu, B.Q.; Chen, X.Q.; Wu, X.G.; Zhang, W.; Wang, Z. Study of Pu’er Raw Materials Grade Classification by PCA and PLS-DA. J. Tea Sci. 2015, 35, 179–184. [Google Scholar] [CrossRef]
  30. Effmert, U.; Große, J.; Röse, U.S.R.; Ehrig, F.; Kägi, R.; Piechulla, B. Volatile composition, emission pattern, and localization of floral scent emission in Mirabilis jalapa (Nyctaginaceae). Am. J. Bot. 2005, 92, 2–12. [Google Scholar] [CrossRef]
  31. Xu, J.; Li, Y.Y.; Zheng, C.S.; Wang, C.; Yoo, Y.K. Studies of Aroma Compounds in Chrysanthemum in Different Florescence and Inflorescence Parts and Aroma Releasing. Acta Bot. Boreali-Occident. Sin. 2012, 32, 722–730. [Google Scholar] [CrossRef]
  32. Goodwin, S.M.; Kolosova, N.; Kish, C.M.; Wood, K.V.; Jenks, M.A. Cuticle Characteristics and Volatile Emissions of Petals in Antirrhinum majus. Physiol. Plant. 2010, 117, 435–443. [Google Scholar] [CrossRef]
  33. Li, Y.Y.; Xu, H.Y.; Ma, H.; Li, Z.H. Temporal Variation of Flora Aroma Emission of a Hybrid Variety of Luculia. J. Shanxi Agric. Univ. Nat. Sci. Ed. 2020, 40, 8–18. [Google Scholar] [CrossRef]
  34. Liu, Q.; Sun, G.F.; Zhang, J.Z.; Li, X.D. Study on Floral Aroma of the Genus Hosta. Sci. Agric. Sin. 2015, 48, 4323–4334. [Google Scholar] [CrossRef]
  35. Lin, R.Y.; Zhong, H.X.; Huang, M.L.; Luo, Y.H.; Lin, B. Study on the Temporal and Spatial Dynamics of Polianthes tuberosa. J. Nucl. Agric. Sci. 2017, 31, 2434–2442. [Google Scholar] [CrossRef]
  36. Zhao, Y.Q.; Pan, H.T.; Zhang, Q.X.; Sun, M.; Pan, C.B. Dynamics of Fragrant Compounds from Prunus mume Flowers. J. Beijing For. Univ. 2010, 18, 310–315. [Google Scholar] [CrossRef]
  37. Shi, T.T.; Yang, X.L.; Wang, L.G. Dynamic Characteristics of Floral Components and Anatomical Observation of Petals in Three Cultivars of Osmanthus fragrans. J. Nanjing For. Univ. Sci. Ed. 2020, 44, 12–20. [Google Scholar] [CrossRef]
  38. Dudareva, N.; Murfitt, L.M.; Mann, C.J.; Gorenstein, N.; Kolosova, N.; Kish, C.M.; Bonham, C.; Wood, K. Developmental Regulation of Methyl Benzoate Biosynthesis and Emission in Snapdragon Flowers. Plant Cell 2000, 12, 949–961. [Google Scholar] [CrossRef]
  39. Kotze, M.J.; Jürgens, A.; Johnson, S.D.; Hoffmann, J.H. Volatiles Associated with Different Flower Stages and Leaves of Acacia cyclops and Their Potential Role as Host Attractants for Dasineura dielsi (Diptera: Cecidomyiidae). S. Afr. J. Bot. 2010, 76, 701–709. [Google Scholar] [CrossRef]
  40. Shi, T.T.; Yang, X.L.; Wang, L.G. Study on the Aroma Component Emission Pattern of Osmanthus fragrans‘Boye Jingui’. J. Nanjing For. Univ. Sci. Ed. 2018, 42, 97–104. [Google Scholar] [CrossRef]
  41. Liu, C.; Zeng, X.L.; Zheng, R.R.; Luo, J.; Wang, C.Y. Cloning and Expression of the Alcohol Acyltransferase Gene from Osmanthus fragrans Flowers. J. Huazhong Agric. Univ. Nat. Sci. Ed. 2016, 35, 36–42. [Google Scholar] [CrossRef]
  42. Kong, Y.; Sun, M.; Pan, H.T.; Zhang, Q.X. Advances in Metabolism and Regulation of Floral Aroma. J. Beijing For. Univ. 2012, 34, 146–154. [Google Scholar] [CrossRef]
  43. Huang, X.L.; Zheng, B.Q.; Wang, Y. Study of Aroma Compounds in Flowers of Dendrobium chrysotoxum in Different Florescence Stages and Diurnal Variation of Full Blooming Stage. For. Res. 2018, 31, 142–149. [Google Scholar] [CrossRef]
  44. Zhang, Y.; Tian, M.; Wang, C.X. Components and Sensory Evaluation of Flower Aroma of Oncidium Sharry Baby‘Sweet Fragrance’under Different Light Conditions. J. Plant Resour. Environ. 2018, 27, 107–109. [Google Scholar] [CrossRef]
  45. Gao, L.P.; Wang, L.M.; Zhang, Y.Q.; Xia, T.; Wan, X.C. Studies on the Aroma Releasing Enzymes of Jasmine Flowers. J. Tea Sci. 2001, 2, 140–143. [Google Scholar] [CrossRef]
  46. Wang, Q.; Zhang, Y.H. Transcriptional Regulation of α-Farnesene Synthesis by Md MYC2 and Md ERF3 in Apple Fruit. Master’s Thesis, Shandong Agricultural University, Taian, China, May 2019. [Google Scholar]
  47. Peralta-Yahya, P.P.; Ouellet, M.; Chan, R.; Mukhopadhyay, A.; Keasling, J.D.; Lee, T.S. Identification and Microbial Production of a Terpene-Based Advanced Biofuel. Nat. Commun. 2011, 2, 483. [Google Scholar] [CrossRef] [PubMed]
  48. Hao, H.L.; Dai, H. Application and challenges of plant essential oils in health pest control. Chin. J. Hyg. Insectic. Equip. 2019, 25, 284–288. [Google Scholar] [CrossRef]
  49. Elegbede, J.A.; Elson, C.E.; Qureshi, A.; Tanner, M.A.; Gould, M.N. Inhibition of DMBA-Induced Mammary Cancer by the Monoterpene d-Limonene. Carcinogenesis 1984, 5, 661–664. [Google Scholar] [CrossRef]
  50. Huo, M.; Cui, X.; Xue, J.; Chi, G.; Gao, R.; Deng, X.; Guan, S.; Wei, J.; Soromou, L.W.; Feng, H.; et al. Anti-Inflammatory Effects of Linalool in RAW 264.7 Macrophages and Lipopolysaccharide-Induced Lung Injury Model. J. Surg. Res. 2013, 180, E47–E54. [Google Scholar] [CrossRef]
  51. Zhang, J.S.; Li, B.; Chen, J.K.; Zhou, T.S. Chemical Constituents and Antimicrobial Activity of Volatile Oil from Solidago canadensis L. J. Fudan Univ. Nat. Sci. 2006, 3, 412–416. [Google Scholar] [CrossRef]
Horticulturae 08 00719 g001 550
Figure 1. Flowers of the three species from Tilia at different flowering stages: bud stage (a), half-open stage (b), and blooming stage (c).
Figure 1. Flowers of the three species from Tilia at different flowering stages: bud stage (a), half-open stage (b), and blooming stage (c).
Horticulturae 08 00719 g001
Horticulturae 08 00719 g002 550
Figure 2. UpSet diagrams based on the aroma components at different flowering stages of the three species. (a) T. cordata; (b) T. tomentosa; (c) T. miqueliana. Note: “set size” represents the dataset of aroma components in each species of Tilia at different flowering periods; the horizontal bar chart on the left represents the statistical value of floral fragrance components in each period; the single point of the intermediate matrix represents the existence of unique aroma components in a certain period, and 2 or 3 points are connected by lines to represent the unique components with intersection of multiple periods; the numbers at the top of the vertical bars represent the values of the unique aroma components at the intersection points at different periods.
Figure 2. UpSet diagrams based on the aroma components at different flowering stages of the three species. (a) T. cordata; (b) T. tomentosa; (c) T. miqueliana. Note: “set size” represents the dataset of aroma components in each species of Tilia at different flowering periods; the horizontal bar chart on the left represents the statistical value of floral fragrance components in each period; the single point of the intermediate matrix represents the existence of unique aroma components in a certain period, and 2 or 3 points are connected by lines to represent the unique components with intersection of multiple periods; the numbers at the top of the vertical bars represent the values of the unique aroma components at the intersection points at different periods.
Horticulturae 08 00719 g002
Horticulturae 08 00719 g003 550
Figure 3. PLS-DA model for aroma components at different stages of the three species. (a) Score scatter plot; (b) Loading scatter plot.
Figure 3. PLS-DA model for aroma components at different stages of the three species. (a) Score scatter plot; (b) Loading scatter plot.
Horticulturae 08 00719 g003
Horticulturae 08 00719 g004 550
Figure 4. Permutation test of PLS-DA model. (a) T. cordata; (b) T. miqueliana; (c) T. tomentosa.
Figure 4. Permutation test of PLS-DA model. (a) T. cordata; (b) T. miqueliana; (c) T. tomentosa.
Horticulturae 08 00719 g004
Horticulturae 08 00719 g005 550
Figure 5. Heat map of content distributions about 14 crucial components in the three species from Tilia.
Figure 5. Heat map of content distributions about 14 crucial components in the three species from Tilia.
Horticulturae 08 00719 g005
Table
Table 1. Basic situation of plant materials selected from the three species.
Table 1. Basic situation of plant materials selected from the three species.
SpeciesAbbreviationBlooming Stages
Flower Diameter (mm)		Collection Date
T. cordataTC10–1227 May 2021
T. miquelianaTM11–1329 May 2021
T. tomentosaTT12–141 June 2021
Table
Table 2. Aroma components and relative contents in samples of the three species from Tilia.
Table 2. Aroma components and relative contents in samples of the three species from Tilia.
No.Compounds NameCAS
Number		RIRelative Content (%)
TC-aTC-bTC-cTT-aTT-bTT-cTM-aTM-bTM-c
1NI-1-----3.30-----
2NI-2---------0.06-
3NI-3----------0.65
4NI-4-731------0.170.200.19
5NI-5-749------0.43-0.19
6(1R)-2, 6, 6-Trimethylbicyclo [3.1.1]hept-2-ene7785-70-8948---2.412.854.04---
7β-Myrcene123-35-3996----2.904.40---
8NI-6-1004------7.335.213.62
9NI-7-10110.700.900.99------
10NI-8-1013----0.83----
11α-Phellandrene99-83-21014-----0.32---
12D-Limonene5989-27-51038---19.8819.0422.550.210.31-
13trans-β-Ocimene3779-61-110400.881.030.96------
14β-Ocimene13877-91-3105229.4734.4436.9032.6232.4432.68---
15Benzene, 1-methoxy-4-methyl-104-93-81056--------0.19
16γ-Terpinene99-85-41067---4.804.975.53---
17Bicyclo [3.1.0]hexan-2-ol, 2-methyl-5-(1-methylethyl)-, (1α,2β,5α)-15537-55-01079----0.330.36 ---
18Benzeneacetaldehyde122-78-11088------5.48 6.415.40
19NI-9-1090------0.270.150.15
20trans-Linalool oxide (furanoid)34995-77-210981.55 1.040.87 ------
21Cyclohexene, 1-methyl-4-(1-methylethylidene)-586-62-91099---12.2212.3511.39---
22Benzene, (2-methoxyethyl)-3558-60-91102------0.250.390.43
23Linalool78-70-6110914.7418.7321.30------
24NI-10-1113----1.79----
252, 4, 6-Octatriene, 2, 6-dimethyl-673-84-711391.561.21 1.19 ------
26NI-11-11492.223.453.406.794.215.0015.1712.039.12
27Lilac aldehyde (isomer II)-1166-0.590.41 -----0.10
28Phenylethyl Alcohol60-12-81180---7.437.17 5.700.681.59 1.77
29(3R,6S)-2,2,6-Trimethyl-6-vinyltetrahydro-2H-pyran-3-ol39028-58-511960.510.340.29 ------
303-Cyclohexen-1-ol, 4-methyl-1-(1-methylethyl)-, (R)-20126-76-51204---0.350.320.37 ---
31Benzoic acid, ethyl ester93-89-01208---0.62 0.94 0.70---
32Benzene, (2-methoxyethenyl)-4747-15-31216------0.13 0.19 0.22
33NI-12-12190.73--------
34Estragole140-67-01225---3.963.87 2.32 ---
35Lilac alcohol (isomer III)-12280.640.40 -------
36Lilac alcohol (isomer IV)-12410.88 0.46 -------
37Lilac alcohol D33081-37-71259--------0.10
38Acetic acid, 2-phenylethyl ester103-45-71295---0.440.50 -0.160.29 0.61
39Tridecane629-50-51300-------0.19 -
40NI-13-1301--------0.09
41NI-14-13190.831.211.121.951.181.793.612.612.11
42NI-15-13590.840.530.46------
43α-Cubebene17699-14-813700.43 0.340.34------
44Aromadendrene109119-91-714091.03 0.820.85------
45Phenol, 2-methoxy-4-(1-propenyl)-97-54-11434---1.142.20 1.00 ---
46β-Copaene18252-44-314364.393.02 2.54 ------
47Methyleugenol93-15-21443------3.71 7.0010.26
48NI-16-14462.571.741.47------
49(E)-β-Famesene18794-84-81459------0.09 0.110.16
50NI-17-14611.180.860.81------
51NI-18-1468------0.120.110.13
52NI-19-14701.711.161.11------
53cis-Muurola-4(15),5-diene157477-72-014790.630.560.50 ------
54NI-20-1489------0.250.180.18
55γ-Muurolene30021-74-014911.52 1.25 1.11 ------
561, 3, 6, 10-Dodecatetraene, 3, 7, 11-trimethyl-, (Z,E)-26560-14-51498------1.231.28 1.68
57Germacrene D23986-74-5150122.3018.6716.830.480.660.90---
58NI-21-15110.890.800.84------
59α-Farnesene502-61-41514---0.750.930.9558.6359.3560.37
60NI-22-15161.260.960.80------
61NI-23-15191.000.620.48------
62Naphthalene, 1,2,3,4,4a,5,6,8a-octahydro-7-methyl-4-methylene-1-(1-methylethyl)-, (1α,4aβ,8aα)-39029-41-915331.811.591.45------
63NI-24-1535------0.340.350.36
64Naphthalene, 1,2,3,5,6,8a-hexahydro-4,7-dimethyl-1-(1-methyl ethyl)-, (1S-cis)-483-76-115393.07 2.692.44 ------
65NI-25-1550-------0.06-
66Naphthalene, 1,2,4a,5,6,8a-hexahydro-4,7-dimethyl-1-(1-methylethyl)-, [1S-(1α,4aβ,8aα)]-24406-05-115560.660.59 0.54 ------
67NI-26-1603------0.590.620.69
68Benzene, 1,2,3-trimethoxy-5-(2-propenyl)-487-11-61609------0.60 0.76 0.66
69NI-27-1623------0.550.550.57
70Benzyl Benzoate120-51-41744---0.86 0.52 ----
Total 100.00100.00100.00100.00100.00100.00100.00100.00100.00
Note: RT—Retention time; RI—Retention index; NI—Not identified.
Table
Table 3. Characteristics of the 14 crucial components in the PLS-DA model.
Table 3. Characteristics of the 14 crucial components in the PLS-DA model.
No.Components NameVIPp-ValueAroma Characteristics
1α-Farnesene3.700.02Citrus, herbal, lavender
2D-Limonene2.760.03Citrus, orange
3Germacrene D2.500.02Woody, spice
4Linalool2.470.02Citrus, woody, floral
5Cyclohexene, 1-methyl-4-(1-methylethylidene)-2.110.02Citrus, woody
6Phenylethyl Alcohol1.550.02floral, rose
7γ-Terpinene1.380.02Oily, woody, lemon
8NI-111.330.03-
9Methyleugenol 1.210.02Spicy, cinnamon, clove
10Benzeneacetaldehyde1.160.02Honey, floral, cocoa
11Estragole1.100.02Spice, herbal, anise
12NI-61.090.02-
13(1R)-2,6,6-Trimethylbicyclo[3.1.1]hept-2-ene1.060.02Terpene, aromatic, minty
14β-Copaene1.040.02-
VIP: variable importance in projection.
Table
Table 4. Aroma similarity rates between the three species of Tilia at different flowering stages.
Table 4. Aroma similarity rates between the three species of Tilia at different flowering stages.
TC-aTC-bTC-cTT-aTT-bTT-cTM-aTM-bTM-c
TC-a1.000 0.986 0.972 0.578 0.585 0.572 0.015 0.012 0.009
TC-b 1.000 0.997 0.626 0.631 0.616 0.021 0.016 0.012
TC-c 1.000 0.635 0.641 0.625 0.020 0.015 0.012
TT-a 1.000 0.991 0.986 0.063 0.058 0.048
TT-b 1.000 0.993 0.052 0.050 0.043
TT-c 1.000 0.056 0.052 0.044
TM-a 1.000 0.996 0.987
TM-b 1.000 0.997
TM-c1.000
Note: The closer clustering relationship is, the aroma similarity rate draws nearer to 1.000.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

Bao, W.; Shen, Y. Dynamic Changes on Floral Aroma Composition of the Three Species from Tilia at Different Flowering Stages. Horticulturae 2022, 8, 719. https://doi.org/10.3390/horticulturae8080719

AMA Style

Bao W, Shen Y. Dynamic Changes on Floral Aroma Composition of the Three Species from Tilia at Different Flowering Stages. Horticulturae. 2022; 8(8):719. https://doi.org/10.3390/horticulturae8080719

Chicago/Turabian Style

Bao, Wenqin, and Yongbao Shen. 2022. "Dynamic Changes on Floral Aroma Composition of the Three Species from Tilia at Different Flowering Stages" Horticulturae 8, no. 8: 719. https://doi.org/10.3390/horticulturae8080719

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop