Scent Emission Research Articles

Floral maturation and changing air temperatures influence scent volatiles biosynthesis and emission in Jasminum auriculatum Vahl

Benzenoid and terpenoid volatiles are the major compounds contributing to the unique floral scent of Jasminum auriculatum. Biosynthesis and emission of specialized scent metabolites showed maturation stage specific pattern; maximum scent emission was observed when flowers start unfurling and become fully opened under in situ condition. The activities of volatile biosynthesizing enzymes and expressions of several scent-related genes correlated well with the fragrance emission patterns. We also assessed the impact of varying air temperatures (20 °C, 25 °C, 30 °C and 35 °C) on the metabolism as well as vaporization of scent volatiles. The contents of both emitted and endogenous volatiles were higher at either 25 °C or 30 °C and showed relatively lower amounts at both border-range temperatures (20 °C and 35 °C). Further, the activities of key pathway enzymes and expressions of several scent-related genes under varying temperatures exhibited similar trends with the scent emission patterns. Analysis of non-volatile metabolites from flowers grown under different air temperatures suggests a perturbation occurring in the primary metabolism and immediate precursors of scent compounds. The knowledge base created through these studies shall be helpful in improving the yield of floral scent production from such horticulturally important plants in controlled cultivation systems.

Identification of transcription factors controlling floral morphology in wild Petunia species with contrasting pollination syndromes.

SummaryAdaptation to different pollinators is an important driver of speciation in the angiosperms. Genetic approaches such as QTL mapping have been successfully used to identify the underlying speciation genes. However, these methods are often limited by widespread suppression of recombination due to divergence between species. While the mutations that caused the interspecific differences in floral color and scent have been elucidated in a variety of plant genera, the genes that are responsible for morphological differences remain mostly unknown. Differences in floral organ length determine the pollination efficiency of hawkmoths and hummingbirds, and therefore the genes that control these differences are potential speciation genes. Identifying such genes is challenging, especially in non‐model species and when studying complex traits for which little prior genetic and biochemical knowledge is available. Here we combine transcriptomics with detailed growth analysis to identify candidate transcription factors underlying interspecific variation in the styles of Petunia flowers. Starting from a set of 2284 genes, stepwise filtering for expression in styles, differential expression between species, correlation with growth‐related traits, allele‐specific expression in interspecific hybrids, and/or high‐impact polymorphisms resulted in a set of 43 candidate speciation genes. Validation by virus‐induced gene silencing identified two MYB transcription factors, EOBI and EOBII, that were previously shown to regulate floral scent emission, a trait associated with pollination by hawkmoths.

Composition and diurnal variation of floral scent emission in Rosa rugosa Thunb. and Tulipa gesneriana L.

AbstractThis study was aimed to explore the composition and diurnal variation analyses of floral scent emission from Rosa rugosa Thunb. and Tulipa gesneriana L. The floral scent from the fresh flower were collected at different time points (9:00, 12:00, 15:00, 18:00, and 21:00) using dynamic headspace collection and were analyzed using autothermal desorber-gas chromatography/mass spectrometry (ATD-GC/MS). The results showed that a total of 62 volatile flavor compounds were detected from Rosa rugosa Thunb and a total of 70 volatile flavor compounds were detected from Tulipa gesneriana L. They were identified with eight functional categories: alcohols, fatty hydrocarbons, terpenes, aldehydes, ketones, esters, and other substances. The total release amount first decreased, and then increased with time, and arrived at the lowest at 15:00. The release amounts of different categories present distinct change patterns. Among the components, phenylethyl alcohol, citronellol, methylene chloride, hexane, and acetone showed relatively higher release amounts and were thought as the main components in floral scent of Rosa rugosa Thunb. Alpha-Farnesene, ethanol, pentadecane, beta-ocimene, longifolene, caryophyllene, and acetone showed relatively higher release amounts and were thought as the main components in floral scent of Tulipa gesneriana L. Research of roses and tulips in aromatic in the garden provides a theoretical basis and research and improvement of the aroma components of aroma.

Assessing the Floral Volatile Constituents of Male and Female Rafflesia Kerri Meijer from Lojing Highlands, Peninsular Malaysia

Rafflesia kerri Meijer is a gigantic parasitic flowering species, endemic in Peninsular Malaysia and Thailand. The flower reported to emit a foul smell, mimicking rotten meat to attract pollinators from the Calliphoridae to visit the male and female flower. Beside the olfactory factor, the visual display with red brownish tinge colour and the enormous size believed to act as secondary attractant in luring the pollinators. However, the study of pollination biology on this species remain limited and information obtained scanty. Herein, this study was aimed to evaluate the presence of chemical compounds by qualitative phytochemical analysis and screen the floral volatile constituents (FVCs) emitted through Head Space – Solid Phase Microextraction – Gas Chromatography – Mass Spectrometry (HS-SPME-GC-MS). The phytochemical screening of both extracts showed the presence of alkaloids, triterpenoids/steroids, flavonoids and tannins, whereas the tentative floral volatile constituents (FVCs) identified were from various chemical classes such as long chain hydrocarbon, organosilicon compound, primary alcohol, aromatic acid as well as miscellaneous compound. The finding suggests that the presence of phytochemical compounds and combination of vast floral volatile constituents identified are believed to contribute in scent emission and attract the pollinators to visit the flower.

Diel scent and nectar rhythms of an African orchid in relation to bimodal activity patterns of hawkmoth pollinators.

The temporal dimensions of floral adaptation to pollinators are not yet well understood, partly because we lack accurate information on the diel rhythms of flower visitation for many pollinators. We investigated whether diel patterns of pollinator visitation to flowers of the African woodland orchid Bonatea polypodantha are synchronized with rhythms of floral anthesis, scent emission and nectar availability. Direct observations and motion-activated cameras were used to identify pollinators of B. polypodantha and to document their activity periods. The timing of pollinaria removal from flowers, emission of scent and availability of nectar was also measured. We found that B. polypodantha is pollinated exclusively by short-tongued hawkmoths. Pollinaria of the orchid are affixed between the labial palps of the moths and brush over the protruding stigmatic arms. The flowers also receive visits by long-tongued hawkmoths, but these act as nectar thieves. Tracking of pollinaria removal from flowers confirmed that pollination occurs only at night. Camera footage revealed a striking crepuscular pattern of foraging by short-tongued hawkmoths with peaks of activity during the twilight periods at dusk and at dawn. In contrast, long-tongued hawkmoths were found to visit flowers throughout the night. Flowers of B. polypodantha exhibit unimodal peaks of anthesis, scent emission (dominated by nitrogenous aromatics) and nectar availability before or around dusk. Flowers of B. polypodantha are pollinated exclusively by short-tongued hawkmoths, which show crepuscular foraging activity at dusk and dawn. Floral phenophases of the orchid are closely synchronized with the peak of pollinator activity at dusk.

Effects of Scent and Scent Emission Methods: Implications on Workers’ Alertness, Vigilance, and Memory under Fatigue Conditions

Extant studies have revealed enhancing effects of scent on performance. The role of scent, and emission method, in the context of performing repetitive tasks over prolonged duration in promoting alertness, vigilance, and memory was examined. Seventy-three participants were randomly assigned to one of three conditions (no scent, peppermint scent emitted continuously, or peppermint scent emitted intermittently) while tasked to monitor and identify suspicious cues for close to 2 hr. Pre- and post-intervention surveys, psychomotor vigilance test, and eye tracker were used as study instruments. Results showed that scent directly enhanced information recall and mitigated the deterioration of alertness, especially as subjects became fatigued. Intermittent emission showed stronger effects over continuous emission in marginally enhancing alertness and memory. Scent did not appear to promote greater vigilance. Suggestions for future studies and implications for management of employees in fatigue situations are discussed.

Sources of floral scent variation in the food-deceptive orchid Orchis mascula

Intraspecific variation of floral signals, and particularly floral scent, is widespread in orchid species. Pollinator-mediated selection is often hypothesized to govern such polymorphism. For example, circadian rhythms of floral emissions have been associated to the daily activity patterns of pollinators in several plant species. However, most of the studies on floral scent variation usually consider only one factor of variation (diel variation, pollination, flower age …). In this study, we investigated simultaneously seven sources of variation susceptible to induce changes in floral scent emissions in the food-deceptive orchid Orchis mascula: circadian rhythm, flower age, pollination, inflorescence morphology, herbivory, leaf spots and habitat. We found that all the factors studied, except plant morphology (spots on leaves, inflorescence morphology), were associated with significant changes in the volatile profile of O. mascula inflorescences. Electroantennography experiments with Bombus terrestris, the main pollinator of O. mascula, revealed that bumblebees were able to detect almost all the compounds involved in floral scent variation. Whether pollinator behaviour is influenced by changes in floral odours induced by these different factors, and whether it can affect plant reproductive success will have to be tested in further studies.

Detecting small and cryptic animals by combining thermography and a wildlife detection dog

Small and cryptic species are challenging to detect and study in their natural habitat. Many of these species are of conservation concern, and conservation efforts may be hampered by the lack of basic information on their ecological needs. Brown hare (Lepus europaeus) leverets - one example of such a small, cryptic and endangered animal - are notoriously difficult to detect, and therefore data on wild leverets are virtually non-existent. Novel technologies and methods such as thermal imaging and the use of wildlife detection dogs represent suitable means for the detection of such species by overcoming the problem of camouflage, using heat or scent emission respectively. Our study on brown hare leverets provides information on how to apply these new techniques successfully for the detection of small and cryptic species, thus enabling the collection of data that was previously inaccessible (e.g. behavioural observation, radio tagging). We found that the choice of method should be made according to vegetative structure. While the handheld thermal imaging camera is best used in areas with no or low vegetative cover, the thermal drone can be used up to medium vegetative cover, whereas the detection dog method is best applied where vegetation is very dense and not suitable to be searched using thermography. Being able to search all sort of different vegetation types, our combined approach enables the collection of a balanced and unbiased dataset regarding habitat type and hence selection of study specimen. We hope that the use of these new techniques will encourage research on many cryptic species that formerly have been neglected because they could not be detected using conventional methodologies.

Unravelling the structure and function of the petal appendages in the tribe Schwenckieae (Solanaceae).

The tribe Schwenckieae (Solanaceae) is characterised by the presence of appendages on the corolla, a diagnostic trait for the group. These appendages constitute a median distal projection of the three-lobed petal and occur in the genera Melananthus and Schwenckia but are absent in Heteranthia. We investigated the micromorphology and anatomical structure of the appendages and lateral petal lobes of Schwenckia americana (two varieties), S.angustifolia, S.curviflora and S.novaveneciana, and Melananthus fasciculatus. We also performed histochemical tests to determine if the appendages are involved in the production of volatiles, acting as a fragrance secretory structure (osmophore). The appendages have a uniseriate epidermis, whose cells store phenolics and lipids. The parenchyma is starch-rich just prior to anthesis in all species studied. The sensory test and anatomical analyses identified scent-secreting tissues, not only in the appendages, but also in the lateral petal lobes, whose cells are papillose with a sculptured surface. The α-naphthol p-phenylenediamine (NADI) reaction detected volatile (essential oils) compounds in S.americana var.americana and S.americana var.angustifolia. We demonstrated the secretory tissues and the production of lipids in the corolla appendages of Schwenckia and Melananthus, which indicate their osmogenic function and probable scent emission to attract pollinators.

I've been robbed! - Can changes in floral traits discourage bee pollination?

Some floral visitors collect nectar by piercing flower external whorls, acting as nectar robbers. They leave robbery vestiges, which can cause changes in floral characteristics, including physical and chemical signals that may influence flower recognition by pollinators. If pollinating bees associate these changes with absence or reduction in nectar volume, they can avoid these flowers, negatively affecting pollination. We aimed to investigate the effect of robbery on primary and secondary attractants. Additionally, we experimentally investigated if the visual signs present in robbed flowers affect the bee pollination of this plant species by discouraging pollinator visits. This study was performed in a very common pollinator-plant-cheaters system comprised by a bee-pollinated Bignoniaceae species and a nectar-robber bee that lands on the corolla tube and makes slits at its base during the nectar robbery. We experimentally isolated the effect of nectar consumption by this nectar-robber and investigated if the slits caused by the nectar-robbers affected the floral scent emission. In addition, we experimentally evaluated the effect of visual signs (slits) associated to the nectar robbery and the effect of nectar depletion on the pollination of Jacaranda caroba (Bignoniaceae). The robbers visited around 75% of the flowers throughout the day and removed significant amounts of nectar from them. However, the damages the robbers cause did not affect floral scent emission and we did not verify significant differences on pollen deposition neither when comparing flowers with slits and control nor when comparing flowers with and without nectar. We showed that even though nectar-robbers visually honestly signal the robbery and deplete high amounts of nectar, they did not affect pollinator visitation. These results showed that presumably antagonistic interactions might in fact not be so.

Pollination elicits an accelerated reduction in nocturnal scent emission by flowers of the dioecious herb Silene latifolia

More than 85% of plants rely on biotic pollen vectors attracted by olfactory or visual signals. While these signals are beneficial for plant reproduction, they are also costly in terms of resource investment, and may involve additional costs related to herbivore attraction, transmission of diseases, and increased transfer of self-pollen among flowers of an individual plant. Given the potential costs of signalling to pollinators, we hypothesized that plants should cease signalling to pollinators as soon as the costs of being attractive exceed the benefits. Here, we used the herb Silene latifolia Poir., which is pollinated by the nocturnal seed-predator Hadena bicruris, to investigate the possible reduction of scent emission after pollination. We compared the progressive decrease of scent and attractive compound emission between pollinated and unpollinated flowers on an hourly basis, overnight. We found that total scent emission decreased significantly faster over time for pollinated than for non-pollinated flowers. Our findings suggest that the costs of maintaining the attractiveness of pollinated flowers of Silene latifolia exceed the benefits, which could be linked with the risks of parasitism.

The Snapdragon LATE ELONGATED HYPOCOTYL Plays A Dual Role in Activating Floral Growth and Scent Emission

The plant circadian clock controls a large number of internal processes, including growth and metabolism. Scent emission displays a circadian pattern in many species such as the snapdragon. Here we show that knocking down LATE ELONGATED HYPOCOTYL in Antirrhinum majus affects growth and scent emission. In order to gain an understanding of the growth kinetics, we took a phenomic approach using in-house artificial vision systems, obtaining time-lapse videos. Wild type flowers showed a higher growth speed than knockdown plants. The maximal growth rate was decreased by 22% in plants with lower LHY expression. Floral volatiles were differentially affected as RNAi plants showed advanced emission of compounds synthesized from cinnamic acid and delayed emission of metabolites of benzoic acid. The monoterpenes myrcene and ocimene were delayed, whereas the sesquiterpene farnesene was advanced. Overall, transgenic lines showed an altered volatile emission pattern and displayed a modified scent profile. Our results show that AmLHY plays an important role in the quantitative and qualitative control of floral growth and scent emission.

MYB1 transcription factor regulation through floral scent in Cymbidium cultivar ‘Sael Bit’

Cymbidium belongs to Orchidaceae, one of the most abundant angiosperm families. Cymbidium cultivars have a high aesthetic value and commercially valued characteristics such as a pleasant scent and a variety of flower colors. Cymbidium cultivar ‘Sael Bit’, which was developed by National Institute of Horticultural & Herbal Science (NIHHS), is highly fragrant with small green and light yellow flowers. An analysis of ‘Sael Bit’ floral scent emissions by an electronic nose showed that the petal and initial flowering stage emitted a strong scent. In an headspace-solid phase micro extraction-gas chromatography-mass spectrometry (HS-SPME-GC-MS) ‘Sael Bit’ study, we identified alkenes and benzenoids, such as tridecane, benzene, and Z-2-octenal, as major volatile organic compounds. V-myb myeloblastosis viral oncogene homolog (MYB) transcription factors (TFs) play a positive role in floral scent regulation. However, little is known about the MYB transcriptional regulation of floral scent biosynthesis. Due to the importance of the plant’s biology, we need to study the floral scent MYB TFs in this plant. In the present work, we isolated and cloned the full-length cDNA of Cymbidium cultivar ‘Sael Bit’ MYB1 from fully open flower. CsMYB1 encodes an 840-bp open reading frame and 280 amino acids by functional characterization. In q-RT-PCR expression analysis, CsMYB1 TF showed the highest expression in petals and columns compared to sepals and labella. Moreover, this MYB gene expressed various flower developmental stages and was highly expressed at the fully open flower stage. The phenylpropanoid/benzenoid and ester compounds responsible genes expressed through CsMYB1 TF regulation in q-RT-PCR analysis. According to these results, we suggest that CsMYB1 might be involved in the regulation of phenylpropanoid/benzenoid genes in floral scent profile. In future studies, we will focus on develop regulation models of floral scent in Cymbidium cultivar ‘Sael Bit’.

Flowering Stage and Daytime Affect Scent Emission of Malus ioensis "Prairie Rose".

Flowering crabapple is an important ornamental flower. It is vital to understand the floral scent properties and the associated release dynamics for carrying out fragrant flower breeding or floral regulation of crabapple. Static headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry was used to detect the volatile compounds in Malus ioensis “Prairie Rose” flowers at different flowering stages and at different day-night time. The results showed that methylheptenone, phenylethanol, geranylacetone, 2-(4-methoxyphenyl)ethanol, α-cedrene were the major compounds in M. ioensis “Prairie Rose”, but the compounds released during different stages and different day-night time were significantly different (P < 0.0001). A total of 25 volatile compounds were identified from the four flowering stages. The floral scents in the initial and flowering stages were the most similar (dissimilarity 0.21). The main compounds in these two stages were geranylacetone and methylheptenone, and the contents of geranylacetone and phenylethanol were positively correlated with the flowering stages. From the bud stage to the end of flowering, the total amount of volatile compounds released showed an initial increase followed by a decrease and the amounts of compounds released during the initial flowering stage were the highest. The aliphatic and benzenoids content was significant higher in the daytime than at night. A total of 15 compounds were detected in the five time periods. Methylheptenone and phenylethanol were particularly released in the 10:00–12:00 and 15:00–17:00 time periods. There were only three common compounds among the five time periods and the types of flower volatiles released during the daytime were obviously higher than those released at night. From the nocturnal to diurnal, the amount of flower volatiles released first increased, then decreased, and the release reached a peak between 10 am and 12 noon, which was consistent with the pollination biological characteristics of Malus flowers. Our findings are important for understanding the mechanism of insect visits to crabapple and the regulation of crabapple flower scent.

Isolation of 2-phenylethanol biosynthesis related gene and developmental patterns of emission of scent compounds in Persian musk rose (Rosa moschata Herrm.)

Abstract The molecular properties of 2-phenylethanol were investigated in the single petals of Persian Musk rose at four floral developmental stages. The chemical compositions of the floral scent emissions were analyzed by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). In this plant, the relative percentage of 2-phenylethanol increased as flowers developed. In the early stage of flower development, the patterns of emission of 2-phenylethanol were very low and then progressively increased reaching its highest value at full bloom stage. The partial CDS of phenyl acetaldehyde reductase (PAR) were isolated in the petals of Persian musk rose. Moreover, the expression pattern of RmPAR showed a significant difference during flower developmental stages and floral organ parts of Rosa moschata. The expression level of RmPAR constantly decreased from the budding stage (stage 1) to the full bloom stage (stage 3); It is reached at the highest level in the budding stage and rapidly decreases in the full bloom stage, and then increases again in the senescence stage (stage 4). In addition, the highest expression level of RmPAR was in the stamen and slightly higher in the petal, and the expression amount in the pistil was about half that in the stamen. Nevertheless, in the calyxes, RmPAR was lowly expressed.

Intense Floral Scent Emission in Polianthes tuberosa L. (Tuberose) Variants Sprouted from γ-Irradiated Tubers

Tuberose (Polianthes tuberosa L.) is a tuberous, perennial, night-blooming ornamental plant which is commercially cultivated in different parts of India. In the present study, the tubers of tuberose were subjected to γ-radiation in order to identify variants with altered scent profile. The growth of plants sprouted from both control and irradiated tubers at different doses was monitored and the lethal dose was calculated based on the survival rate of plants. The growth rate significantly decreased in plants sprouted from tubers exposed to a higher dose. Growth pattern of plants sprouted from irradiated tubers and control also correlated with the chlorophyll content. The emitted scent volatiles from flowers developed from both control and irradiated plants were analysed. Floral volatile emission rate was increased in the lowest dose (10 Gy) as compared to control plants. The phenomenon of ‘hormesis’ was observed in the plants since tubers exposed to the lower dose (10 Gy) showed enhanced growth rate, higher volatile content in comparison with control plants. Plants sprouted from tubers exposed to a higher dose of radiation showed lethal effects. In order to study genetic variation among the control and plants sprouted from irradiated tubers, inter simple sequence repeat marker analysis was also carried out and it was observed that out of 74 loci, 67 were polymorphic (90.54%). The genetic similarity coefficient values were also calculated among control and variant lines thus validating morpho-physiological variations.

The Petunia CHANEL Gene is a ZEITLUPE Ortholog Coordinating Growth and Scent Profiles.

The floral perianth, comprising sepals and petals, conceals the sexual organs and attracts pollinators. The coordination of growth and scent emission is not fully understood. We have analyzed the effect of knocking down CHANEL (PhCHL), the ZEITLUPE ortholog in petunia (PhCHL) by hairpin RNAs. Plants with low PhCHL mRNA had overall decreased size. Growth evaluation using time lapse image analysis showed that early leaf movement was not affected by RNAi:PhCHL, but flower angle movement was modified, moving earlier during the day in knockdown plants than in wild types. Despite differences in stem length, growth rate was not significantly affected by loss of PhCHL. In contrast, petal growth displayed lower growth rate in RNAi:PhCHL. Decreased levels of PhCHL caused strongly modified scent profiles, including changes in composition and timing of emission resulting in volatile profiles highly divergent from the wild type. Our results show a role of PhCHL in controlling growth and development of vegetative and reproductive organs in petunia. The different effects of PhCHL on organ development indicate an organ-specific interpretation of the down regulation of PhCHL. Through the control of both timing and quantitative volatile emissions, PhCHL appears to be a major coordinator of scent profiles.

Butterfly pollination of Bonatea cassidea (Orchidaceae): Solving a puzzle from the Darwin era

Although moths and butterflies are closely related, they tend to respond to different floral cues for foraging and thus select for different suites of floral traits. Shifts between moth and butterfly pollination in the same plant clade provide an opportunity to assess the traits that are required to impart a shift between these pollination systems. The orchid genus Bonatea consists mostly of night-scented moth-pollinated taxa, but here we document a butterfly pollination system in Bonatea cassidea, a century after Mansel Weale's initial observations of butterflies carrying pollinaria of this taxon in South Africa were communicated to the Linnean Society by Charles Darwin. We recorded visits by a number of species of butterflies from the family Hesperiidae and Pieridae, but contrary to Weale, we found that the majority of pollinaria were firmly affixed between the palpi and not on the sternum. Removal of pollinaria from flowers during the day, but not at night, suggested that moths do not contribute to pollination, despite the white flower colouration. Floral traits found in B. cassidea but not its moth-pollinated congeners include diurnal anthesis, scent emission which is weak in the day and virtually absent at night, and the production of small amounts of relatively dilute sucrose-rich nectar in short spurs. These features are consistent with those expected for butterfly-pollinated plants in general. This study represents the first detailed report of butterfly-pollination in Bonatea and one of very few cases of orchid pollination by pierid butterflies.

Dynamic trajectories of volatile and non‐volatile specialised metabolites in ‘overnight’ fragrant flowers of Murraya paniculata

Ephemeral flowers, especially nocturnal ones, usually emit characteristic scent profiles within their post-anthesis lifespans of a few hours. Whether these flowers exhibit temporal variability in the composition and profile of volatile and non-volatile specialised metabolites has received little attention. Flowers of Murraya paniculata bloom in the evenings during the summer and monsoon, and their sweet, intense fragrance enhances the plant's value as an ornamental. We aimed to investigate profiles of both volatile and non-volatile endogenous specialised metabolites (ESM) in nocturnal ephemeral flowers of M.paniculata to examine whether any biochemically diverse groups of ESM follow distinct patterns of accumulation while maintaining synchrony with defensive physiological functions. Targeted ESM contents of M.paniculata flowers were profiled at ten time points at 2-h intervals, starting from late bud stage (afternoon) up to the start of petal senescence (mid-morning). Emitted volatiles were monitored continuously within the whole 20-h period using headspace sampling. The ESM contents were mapped by time point to obtain a highly dynamic and biochemically diverse profile. Relative temporal patterns of ESM accumulation indicated that the active fragrance-emitting period might be divided into 'early bloom', 'mid-bloom' and 'late bloom' phases. Early and late bloom phases were characterised by high free radical generation, with immediate enhancement of antioxidant enzymes and phenolic compounds. The mid-bloom phase was relatively stable and dedicated to maximum fragrance emission, with provision for strong terpenoid-mediated defence against herbivores. The late bloom phase merged into senescence with the start of daylight; however, even the senescent petals continued to emit fragrance to attract diurnal pollinators. Our study suggests that dynamic relations between the different ESM groups regulate the short-term requirements of floral advertisement and phytochemical defence in this ephemeral flower. This study also provided fundamental information on the temporal occurrence of emitted volatiles and internal pools of specialised metabolites in M.paniculata flowers, which could serve as an important model for pollination biology of Rutaceae, which includes many important fruit crops.

Anatomical features related with pollination of Neottia ovata (L.) Bluff & Fingerh. (Orchidaceae)

The survey of anatomical features of Neottia ovata confirmed that the lip base and the groove function as a nectary. The secretory tissue was present in buds about 1,3 mm width. The sweet scent comes from the aromatic constituents of the nectar and also is released by the lip lobes and margins of all tepals. Additionally, the stomata present on the keels on the abaxial surface of sepals and large substomatal cavities may be connected with fragrance emission. The secretory function was not observed on the stem trichomes. The groove is formed by a single-layered epidermis, few layers of subepidermal cells and underlying parenchyma. The abundant starch grains present in buds, absent in sepals and lip epidermis in larger buds and almost absent in open flowers (visible in plastids in TEM) suggest gradual reduction during floral development and nectar secretion. The nectar was gathered under the cuticle, which was visible in the groove and lip base. The main mode of nectar release is by diffusion through micro-channels. The presence of irregular plasmalemma, dictyosomes, and profiles of ER could also indicate the granulocrine mode of secretion. The vesicles originated from ER or dictyosomes are fused with irregular plasmalemma. The irregular plasmalemma with invaginations (observed in groove) and cell wall protuberances (observed at the lip base) along the outer periclinal wall may function as highly specialized cells - transfer cells. Numerous idioblasts, visible in all tepals and in the staminodium, attract pollinators. In the anther, the endothecial cell thickenings were composed of numerous complete rings or helical segments in a closely spaced parallel arrangement (type I), providing the mechanical forces for anther dehiscence and opening and release of pollen grains.