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    Topic review

    Dendrobium Essential Oil

    Subjects: Plant Sciences
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    Definition

    A detailed chemical composition of Dendrobium essential oil has been only reported for a few main species. This article is the first to evaluate the essential oil composition, obtained by steam distillation, of five Indian Dendrobium species: Dendrobium chrysotoxum Lindl., Dendrobium harveyanum Rchb.f., and Dendrobium wardianum R.Warner (section Dendrobium), Dendrobium amabile (Lour.) O’Brien, and Dendrobium chrysanthum Wall. ex Lindl. (section Densiflora). We investigate fresh flower essential oil obtained by steam distillation, by GC/FID and GC/MS. Several compounds are identified, with a peculiar distribution in the species: Saturated hydrocarbons (range 2.19–80.20%), organic acids (range 0.45–46.80%), esters (range 1.03–49.33%), and alcohols (range 0.12–22.81%). Organic acids are detected in higher concentrations in D. chrysantum, D. wardianum, and D. harveyanum (46.80%, 26.89%, and 7.84%, respectively). This class is represented by palmitic acid (13.52%, 5.76, and 7.52%) linoleic acid (D. wardianum 17.54%), and (Z)-11-hexadecenoic acid (D. chrysantum 29.22%). Esters are detected especially in species from section Dendrobium, with ethyl linolenate, methyl linoleate, ethyl oleate, and ethyl palmitate as the most abundant compounds. Alcohols are present in higher concentrations in D. chrysantum (2.4-di-tert-butylphenol, 22.81%), D. chrysotoxum (1-octanol, and 2-phenylethanol, 2.80% and 2.36%), and D. wardianum (2-phenylethanol, 4.65%). Coumarin (95.59%) is the dominant compound in D. amabile (section Densiflora) and detected in lower concentrations (range 0.19–0.54%) in other samples. These volatile compounds may represent a particular feature of these plant species, playing a critical role in interacting with pollinators.

    1. Introduction

    The Orchidaceae family, with its huge number of species that evolved different pollination systems, is known for the variety and complexity of its floral scents, which according to Kaiser (1993), could potentially cover all the spectrum of fragrances occurring in nature [1]. Floral scent, which derives from the composition of volatile organic compounds emitted by the flowers’ tissues (floral VOCs), is fundamental for the defense against pathogens/herbivores and pollinator responses [2]. This trait, together with other characteristics of flowers, such as the color, the presence of nectar, and other peculiarities of the reproductive portions, contributes indeed to defining pollination syndromes [3]. The genus Dendrobium Sw., 1799 (Epidendroideae; Dendrobiinae), which accounts for about 1100 species distributed in Pacific Islands, Asia, and Australia, is one of the largest of the family [4]. As potted and cut flowers, Dendrobium species and hybrids are of great economic interest, being at the top ten among the most commercially traded orchid taxa [5]; several species are also grown and sold for medicinal purposes [6][7]. A large number of taxa, the great morphological diversity, and the wide distribution range have contributed to taxonomic ambiguities that are currently under debate [4][8][9]. In the phylogenetic revision of the genus, Takamiya et al. (2014) considered the presence of papillae on the flower’s lip in entities belonging to different clades. They demonstrated that this character evolved as an adaptation to bee pollination by Dendrobium species [4]. As stated in previous studies, bee-pollinated orchid flowers exhibit papillose carpets, identified as osmophores, structures of accumulation of substances responsible for floral fragrances [10][11]. Takamiya et al. (2014) recorded odor-producing cells in all species of Section Densiflora and the majority of the Section Dendrobium, thus hypothesizing that this character has probably been acquired after the divergence between the Asian and the Australasian Superclades [4]. Despite the great number of studies aimed at optimizing in vitro propagation protocols (i.e., Marting and Madassery, 2006; Teixera da Silva et al., 2015; Calevo et al. 2020; and references therein) [12][13][14], and at characterizing anatomical and chemical traits (Carlsward et al., 1997; Xu et al., 2013; Devadas et al., 2016 and references therein) [15][16][17], the genus Dendrobium has been little investigated from the point of view of the reproductive biology, and even less is known about floral volatilome [18]. To the best of our knowledge, only a few authors had carried out characterizations of floral volatiles from Dendrobium species. Flath and Ohinata (1982) investigated the VOCs of D. superbum Rchb. f. (syn. D. anosmum Lindl.), which is pollinated by the melon fly (Dacus cucurbitae), finding a significant amount of 4-phenylbutan-2-one, whose structure is closely related to another known fly attractant [19]. Brodmann et al. (2009) worked on D. sinense Tang and F.T.Wang and reported that this species emits (Z)-11-eicosen-1-ol (a molecule present in the alarm pheromone of honeybees) to attract hornets for pollination [20]. Silva et al. (2015) recognized terpenes as the most abundant class of compounds in the floral volatiles of D. nobile Lindl. [21]. Julsrigival et al. (2013) found a prevalence of 2-pentadecanone in D. parishii Rchb.f. [22]. Robustelli della Cuna et al. (2017), instead, compared the essential oil of different portions of D. moschatum (Buch.-Ham.) Sw., including the inflorescence: They observed differences among the volatile compositions, and then hypothesized that compounds like ketones or long-chain methyl and ethyl esters play a role as pollinator attractants [23]. The few reports dedicated to reproductive biology have stated that there are various ways for which Dendrobium species attract pollinators: There are cases of shelter mimicry [24][25][26][27][28], nectar rewarding [18], chemical and visual attraction [29], rest and mating place offering, or generalized food deception strategies like a simulation of other co-flowering species occurring in the same habitat [30]. In this work, we aimed to characterize and compare the floral volatiles of five Dendrobiums belonging to sections Dendrobium and Densiflora of the Asian Superclade [4][9]. In particular, we characterized the volatile fractions of the inflorescences of D. chrysanthum Wall. ex Lindl. (Figure 1A), D. harveyanum Rchb. f. (Figure 1B) and D. wardianum R.Warner (Figure 1C) from section Dendrobium, Core subclade of Clade A, and D. chrysotoxum Lindl. (Figure 1D) and D. amabile (Lour.) O’Brien (Figure 1E) from Clade A and C, respectively, of section Densiflora (according to Takamiya et al. 2014) [4].
    Figure 1. Dendrobium chrysanthum (A), D. harveyanum (B), D. wardianum (C), D. amabile (D), and D. chrysotoxum (E), greenhouse-grown plants cultivated in Turin (Italy).

    2. Current Researches and Results

    The yields of D. amabileD. chrysanthumD. chrysotoxumD. harveyanum, and D. wardianum essential oils obtained by steam distillation from fresh flowers were evaluated as 0.09%, 0.34%, 0.33%, 0.39%, and 0.33% (weight/dry weight basis), respectively. Table 1 shows the results of qualitative and quantitative oil analyses on the Elite-5MS column. The compounds are listed in order of their elution and are reported as percentages of the total essential oil. Differences in the qualitative and quantitative compositions of the obtained essential oils have been observed. As shown in the Venn’s diagram (Figure 2), only palmitic acid was shared by all five taxa. On the other hand, 30 compounds were uniquely identified in D. chrysotoxum, and nine, eight, four, and three in D. wardianumD. harveyanumD. chrysanthum, and D. amabile, respectively. Furthermore, 21 compounds were found shared by D. chrysotoxum and D. wardianum. Below, the qualitative and quantitative description of essential oils for each taxon. The Pie chart (Figure 3) shows that the essential oils were different depending on the different species: It can be observed that the main constituents were compounds belonging to saturated hydrocarbons, acids, esters, coumarin, and alcohol classes.
    Figure 2. Venn’s diagram shows both the number of compounds shared and unshared/peculiar among the five Dendrobium species. Percentages are referred to the total number of compounds found, not to the relative abundance.
    Figure 3. Pie chart of distribution of the classes.
    Table 1. Essential oils composition of inflorescences from the five Dendrobium species.
    Compound a RI b RI c Section Dendrobium Section Densiflora  
    D. chrysotoxum D. harvejanum D. wardianum D. amabile D. chrysanthum Identification d
    % % % % %  
    Octane 800 800 - 0.15 - - - RI, NIST
    Hexanal 802 801 0.73 0.06 0.02 - - RI, NIST
    2-hexanol 804 808 - 0.12 - - - RI, NIST
    Diacetone alcohol 841 841 - - - - 0.68 RI, NIST
    α-pinene 939 931 0.21 - - - - MS, NIST
    Benzaldehyde 960 958 0.14 - - - - RI, NIST
    β-pinene 979 973 0.03 - - - - MS, NIST
    Caproic acid 1005 1003 0.06 - - - - RI, NIST
    α-terpinene 1017 1015 0.10 - - - - RI, NIST
    o-Cymene 1026 1023 0.09 - - - - RI, NIST
    Limonene 1029 1027 0.17 - - - - RI, NIST
    Benzyl alchol 1032 1035 0.21 - 0.52 - - RI, NIST
    β-Isophorone 1042 1041 0.51 -   - - RI, NIST
    Phenylacetaldehyde 1042 1043 0.84 - 0.06 - - RI, NIST
    2-octenal 1056 1058 - 0.13 - - 0.06 RI, NIST
    γ-Terpinene 1060 1059 0.76 - 0.04 - - RI, NIST
    Unidentified - 1065 - - 2.89 - - -
    cis-sabinene hydrate 1070 1067 0.27 - - - - MS, NIST
    dihydromyrcenol 1073 1073 - 0.04 - - 0.06 RI, NIST
    1-octanol 1070 1074 2.80 - 0.41 - - MS, NIST
    trans-sabinene hydrate 1098 1098 0.20 - - - - RI, NIST
    Linalool 1097 1101 0.34 0.08 - - - MS, NIST
    Nonanal 1102 1105 - 0.16 - - - RI, NIST
    2-phenylethanol 1107 1115 2.36 - 4.65 - - MS, NIST
    Methyl octanoate 1127 1127 0.04 - - - - RI, NIST
    cis-verbenol 1141 1142 0.92 - - - - RI, NIST
    trans-verbenol 1145 1148 4.60 - - - - RI, NIST
    Camphor 1150 1157 - 0.12 - - - MS, NIST
    Nonenal 1162 1161 0.41 - 0.17 - - RI, NIST
    α-phellandren-8-ol 1170 1169 2.15 - - - - RI, NIST
    Terpinen-4-ol 1177 1179 1.53 - - - - RI, NIST
    Diethyl succinate 1182 1184 0.33 - - - - RI, NIST
    p-cymen-8-ol 1183 1186 0.29 - - - - RI, NIST
    α-terpineol 1189 1192 0.18 - - - 0.28 RI, NIST
    Ethyl octanoate 1196 1199 0.20 - - - - RI, NIST
    Decanal 1202 1206 - - 0.04 - - RI, NIST
    Verbenone 1205 1210 0.20 - - - - MS, NIST
    2,4-nonandienal 1212 1214 - - 0.03 - - RI, NIST
    4-vinylphenol 1224 1221 - - 0.52 0.08 - RI, NIST
    3-phenyl-1-propanol 1232 1231 - - 0.08 - - RI, NIST
    Phenylacetic acid ethyl ester 1247 1247 0.15 - 0.72 - - RI, NIST
    Nerol 1254 1256 0.06 - - - - RI, NIST
    2,4-decadienal (E,E) 1291 1295 0.40 0.39 0.39 0.16 - RI, NIST
    2-methoxy-4-vinyl-phenol 1315 1315 - - 0.24 - - RI, NIST
    2,4-decadienal (E,Z) 1319 1317 0.63 0.88 0.48 0.72   RI, NIST
    2-nonenoic acid-γ-lactone 1345 1344 0.39 - 0.49 - - RI, NIST
    Capric acid 1359 1359 - 0.32 -     RI, NIST
    Eugenol 1367 1366 - - - 0.10 - RI, NIST
    1-tetradecene 1390 1393 - 0.07 -   0.57 MS, RI
    3,4-dihydrocoumarin 1398 1399 - - - 0.10 - RI, NIST
    Coumarin 1434 1436 0.71 0.19 0.54 95.49 - RI, NIST
    9-epi-(E)-caryophyllene 1466 1458 - - 1.32 - - MS, NIST
    Ethyl-cinnammate 1467 1468 - - 0.55 - - RI, NIST
    2,4-di-tert-butylphenol 1494 1489   -   0.12 22.81 MS, NIST
    β-selinene 1494 1489 0.25 - 1.30 - - MS, NIST
    9-oxo-ethyl-nonanoate 1507 1510 1.28 - - - - MS, NIST
    Lauric acid 1566 1568 0.23 - - - - RI, NIST
    Ethyl laurate 1593 1596 0.15 - - - - RI, NIST
    Unidentified - 1658 - 5.16 - - - -
    Pentadecan-2-one 1667 1667 - - 0.26 - - RI, NIST
    Heptadecane 1700 1700 0.31 - 0.54 - - RI, NIST
    Unidentified - 1767 0.39 - 3.04 - - -
    Myristic acid 1780 1776   - 3.59 - - MS, NIST
    1-octadecene 1790 1796 0.32 - 0.41 - - MS, RI
    Methyl pentadecanoate 1820 1828 0.04 - - - - MS, NIST
    Unidentified - 1879 5.74 - - - - -
    Ethyl pentadecanoate 1890 1896 0.36 - 0.19 - - MS, NIST
    Heptadecan-2-one 1902 1903 0.11 -   - - RI, NIST
    Methyl palmitate 1927 1928 0.34 - 0.44 - - RI, NIST
    cis-9-hexadecenoic acid 1942 1943 - - - - 4.06 RI, NIST
    Z-11-Hexadecenoic acid 1953 1953 - - - - 29.22 RI, NIST
    Palmitic acid 1958 1960 0.05 7.52 5.76 0.61 13.52 RI, NIST
    Neocembrene 1960 1966 0.52 - 3.07 - - MS, NIST
    Ethyl palmitate 1992 1997 3.05 - 0.99 - - MS, NIST
    Octadecan-1-ol 2074 2071 0.17 - 0.60 - - MS, NIST
    Eicosane 2000 2000 - 40.42 - - 0.55 RI, NIST
    Unidentified - 2037 - 2.06 - -   -
    Methyl linoleate 2051 2068 7.48 2.50 13.17 - 1.03 MS, NIST
    10-Heneicosene 2060 2073 - - - 0.43 - MS, RI
    Heneicosane 2100 2100 1.01 2.92 1.66 0.25 - RI, NIST
    Linoleic acid 2144 2147 0.12 - 17.54 - - RI, NIST
    Ethyl linolenate 2169 2171 26.98 - 32.24 - - RI, NIST
    Ethyl oleate 2179 2181 5.39 - 0.72 - - RI, NIST
    Ethyl octadecanoate 2193 2198 0.80 - 0.31 - - RI, NIST
    Docosane 2200 2204 1.66 26.82 - 1.94 17.53 RI, NIST
    9-Triacosene 2279 2275 0.31 - - - - MS, RI
    Tricosane 2300 2307 9.33 - - - - RI, NIST
    Tetracosane 2400 2401 0.40 0.90 - - 2.07 RI, NIST
    9-Pentacosene 2474 2475 0.07   - -   MS, RI
    Pentacosane 2500 2501 0.95 6.53 - - 6.40 RI, NIST
    Hexacosane 2600 2600 - 2.46 - - - RI, NIST
    9-Eptacosene 2676 2676 - - - - 1.15 MS, RI
    Heptacosane 2700 2701 0.18 - - - - RI, NIST
    Aldehydes     3.15 1.62 1.20 0.88 0.06  
    Alcohols     7.97 0.12 7.02 0.30 22.81  
    Acids     0.45 7.84 26.89 0.61 46.80  
    Coumarin     0.71 0.19 0.54 95.59 -  
    Esters     46.59 2.50 49.33 - 1.03  
    Ketones     0.62 0.12 0.26 - 0.68  
    Saturated hydrocarbons     22.84 80.20 2.20 2.19 26.55  
    Unsaturated hydrocarbons     0.69 0.07 0.41 0.43 1.72  
    Terpenes     2.04 - 5.73 - -  
    Oxygenated terpenes     8.31 0.11 - - 0.34  
    Miscellanea     0.48 - 0.49 - -  
    Unidentified     6.13 7.22 5.92 - -  
    a) Compounds are listed in order of elution from an Elite-5 column. b) Retention Indices according to Adams [31], unless stated otherwise. c) Retention index (mean) determined on an Elite-5 column using a homologous series of n-hydrocarbons, d) Method of identification: MS, mass spectrum; NIST, comparison with library [32]; RI, retention indices in agreement with literature values.

    3. Discussion

    Little is known about the pollinators of the studied species, but as argued by Dobson (2006) and Witjes et al. (2011), it is possible to reconstruct the pollinator community behind a certain species by analyzing the volatile composition of flowers [33][34]. While research is still needed to identify pollinators, our analyses constitute a first contribution for the study of compounds possibly involved in plant-animal interactions. However, other functions of floral volatiles, that may play a crucial role in herbivory avoidance and as defensive molecules against pathogens, cannot be excluded [35][36]. Differences in the floral scents of related taxa could play a role in reproductive isolation by influencing pollinator’s behavior and choices [37][38][39][40]. Indeed, in some cases, a simple change in the amount of one floral VOC has been linked with strong reproductive isolation, as seen in Silene dioica (L.) Clairv. and S. latifolia Poir. [41]. However, this ethological type of isolation seems to be more or less pivotal depending on the specialization of both the plants and pollinators considered, highlighting the need to carry out additional detailed behavioral experiments to understand plant-pollinator interactions [3].
    In this work, the relative composition in floral VOCs of the five Dendrobium species was qualitatively studied. The highest number of species-specific compounds were recorded for entities from section Dendrobium. Palmitic acid was the only compound shared by all the five taxa examined. This molecule is frequently found in the volatilome of several plant species (Orchidaceae included) [23][35][42], and also in other organisms; we observed that it was relatively abundant in D. chrysanthum (13%), followed by D. harveyanum (7.52%) and D. wardianum (5.76%), while in the remaining two species it was less represented.
    The scent recognized for both D. chrysotoxum and D. wardianum could be due to the high presence of esters in floral VOCs that we detected during our analyses. Esters are produced by the reaction of alcohols with organic acids; they typically have fruity smells and are indeed among the molecules responsible for the odors of many fruits [43]. High content of volatile esters has been linked with the strong flavor of the “snow chrysanthemum” cultivar of Coreopsis by Kim et al. (2020) [44]. In D. moschatum, a putative role as semiochemicals involved in pollinator attraction has been hypothesized for methyl and ethyl esters by Robustelli della Cuna et al. (2017) [23]. According to da Silva et al. (1999) and Cseke et al. (2007), terpenes are more abundant in flower VOCs of species pollinated by food-seeking bees [45][46]. As shown in Table 1D. wardianum had the highest level (5.73%) of terpenes in the essential oil, followed by D. chrysotoxum (2.04%), but this class of compounds was not the predominant one in these two species. Conversely, oxygenated terpenes have been detected only in D. chrysotoxum (8.31%), while they were present in lower percentages in D. harveyanum and D. chrysanthum. Therefore, due to their ester and terpenoid contents, and considering similar results obtained by Flath and Ohinata (1982) for D. superbum, we cannot exclude that D. chrysotoxum and D. wardianum could rely on the action of frugivorous flies or bees, or other animals for their pollination [19].
    It is noteworthy that the VOCs spectrum of D. amabile, a scented orchid, was almost entirely dominated by coumarin, a compound having a sweet smell that resembles vanilla. On the contrary, this compound was present only in very small percentages in all the other Dendrobiums considered. As previously stated by Robustelli della Cuna et al. 2017, coumarin was abundant, although less represented in respect to D. amabile, also in VOCs from inflorescences and leaves of D. moschatum [23]. In this species, authors hypothesized a phytoalexin-like defensive role for coumarin. In the future, a possible role of coumarin in plant-pollinator interactions should be investigated. Interestingly, D. chrysanthum showed a distinctive floral volatile composition compared to the other species. Indeed, this entity displayed the highest amounts of acids (accounting for 46.8% of the total essential oil), together with a good representation of alcohols (22.8%) if compared to the other species considered. Among acids, the most representative one (29.2%) was Z-11-Hexadecenoic acid, a known sex pheromone in moths [47]. Considering the relatively high content of this compound, we can again hypothesize its possible role as pollinator (putatively, moth) attractant. Concerning alcohols, 2,4-di-tert-butylphenol was relatively abundant in D. chrysanthum. This molecule was also present in traces in D. amabile. Zhang et al. (2017) and Huang et al. (2018) recorded the occurrence of this alcohol in flowers of D. moniliforme (L.) Sw. and rhizomes of Gastrodia elata Blume, respectively [48][49]. This compound, known for its toxicity, exerts several bioactivities and has insecticidal, nematicidal, antibacterial, and antifungal properties (Zhao et al., 2020 and references therein) [50]. Therefore, a defensive role for 2,4-di-tert-butylphenol in D. chrysanthum cannot be excluded. Finally, it is interesting to notice that among the Dendrobium and Densiflora sections, three self-incompatible species, D. amabile and D. harveyanum, and D. chrysanthum, respectively, showed a reduced spectrum of volatiles [51][52]. It is tempting to hypothesize that this has a role in pollination biology; indeed, discouraging pollinators from pollinating more flowers of the same plant and inducing pollinators to visit different individuals, would result in a higher fruit set.

    This entry is adapted from 10.3390/plants10081718

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