Secretory Cavity Development and Its Relationship with the Accumulation of Essential Oil in Fruits of Citrus medica L. var. sarcodactylis （Noot.） Swingle

The developmental types of secretory cavities in Citrus remain controversial. The relationship between secretory cavity development and the accumulation of essential oil in fruits of Citrus species is also unknown. In order to develop better insights into these problems, histological, histochemical, and cytochemical methods were used to investigate secretory cavity development and the accumulation of essential oll at different developmental stages of fruits of Citrus medica L. var. sarcodactylis （Noot.） Swingle. The results indicate that the secretory cavity of the variety seemed to originate from an epidermal cell and a subepldermal cell. These two cells underwent successive divisions, resulting In the formation of two parts： （Ⅰ） a conical cap; and （Ⅱ） a globular gland. The formation of the lumen was schlzolysigenous. Regular changes in the size of vacuoles and the accumulation of essential oil were revealed during the process of secretory cavity development. In addition, when fruits were a light yellow or golden color, the structure of secretory cavities was well developed and the content of essential oil in a single fruit reached a maximum. It would be most appropriate to collect the fruit as a medicinal material at this time.     

杭白芷根中分泌道发生方式、分布及其挥发油积累过程研究

为探明杭白芷(Angelica dahurica var.formosana)根中分泌道发生方式、分布及其挥发油转运积累特征,该研究利用光镜及透射电子显微镜技术观察分泌道发生过程及挥发油转运特征,结合组织化学定位确定挥发油的主要积累部位。结果表明:杭白芷根中分泌道由中柱鞘细胞最先发生,次生结构中分泌道主要分布在韧皮部和皮层中;挥发油的合成不仅与分泌细胞中质体及细胞质有关,而且还与周围细胞关系密切;分泌细胞内高尔基体和内质网丰富,可能先通过形成小泡参与转运,再经由细胞壁向腔道内转移;相邻分泌细胞靠近角隅处的细胞壁分泌活动活跃,腔道内积累大量电子致密物质;成熟分泌道中分泌细胞及其腔道内积累大量油滴,因此挥发油主要积累场所为分泌细胞及其腔道。该研究明确了杭白芷根中分泌道的发生方式、分布及其挥发油积累部位,揭示了分泌道发育过程中挥发油的转运积累特征,为进一步阐明分泌组织生长发育与有效成分积累关系提供了理论依据。     

Anatomical and chemical analyses of leaf secretory cavities of Rustia formosa (Rubiaceae)

Foliar secretory cavities, commonly called leaf pellucid glands, have been reported in many families of vascular plants. In the Rubiaceae, these structures have only been found in the sister genera Rustia and Tresanthera, which are also anomalous within the family because they have poricidal anthers, and in the distantly related Heterophyllaea. General leaf anatomy, with particular attention to secretory cavities, as well as the chemical analysis of the secreted substances of Rustia formosa, is presented here for the first time. The secretory structures have been found in the lamina between the palisade and spongy parenchymas and in the cortical region of the petiole. The chemical analysis showed that the essential oil secreted is a complex mixture of at least 75 components, mostly of sesquiterpenoid composition. Illustrations of the leaf anatomy, details of the secretory structures of Rustia formosa, a gas chromatogram, and a table of the principal components of the leaf essential oil are included.     

Limonene Synthase, the Enzyme Responsible for Monoterpene Biosynthesis in Peppermint, Is Localized to Leucoplasts of Oil Gland Secretory Cells

Circumstantial evidence based on ultrastructural correlation, specific labeling, and subcellular fractionation studies indicates that at least the early steps of monoterpene biosynthesis occur in plastids. (4S)-Limonene synthase, which is responsible for the first dedicated step of monoterpene biosynthesis in mint species, appears to be translated as a preprotein bearing a long plastidial transit peptide. Immunogold labeling using polyclonal antibodies raised to the native enzyme demonstrated the specific localization of limonene synthase to the leucoplasts of peppermint (Mentha × piperita) oil gland secretory cells during the period of essential oil production. Labeling was shown to be absent from all other plastid types examined, including the basal and stalk cell plastids of the secretory phase glandular trichomes. Furthermore, in vitro translation of the preprotein and import experiments with isolated pea chloroplasts were consistent in demonstrating import of the nascent protein to the plastid stroma and proteolytic processing to the mature enzyme at this site. These experiments confirm that the leucoplastidome of the oil gland secretory cells is the exclusive location of limonene synthase, and almost certainly the preceding steps of monoterpene biosynthesis, in peppermint leaves. However, succeeding steps of monoterpene metabolism in mint appear to occur outside the leucoplasts of oil gland cells.     

Diversity of essential oil glands of clary sage (Salvia sclarea L., Lamiaceae)

The Lamiaceae is rich in aromatic plant species. Most of these species produce and store essential oils in specialised epidermal oil glands, which are responsible for their specific flavour. Two types of glands producing essential oil and possessing different morphological structure can be found in Salvia sclarea : peltate and capitate glands. The content of single oil glands from different positions on the plant (corolla, calyx and leaf) were sampled using an SPME fibre and analysed by gas chromatography in order to study variability of the essential oil composition. It was found that the composition of terpenoids is quite variable within an individual plant. Capitate oil glands mainly produce three essential oil compounds: the monoterpenes linalool and linalyl acetate, and the diterpene sclareol. Peltate oil glands, however, accumulate noticeable concentrations of sesquiterpenes and an unknown compound (m/z = 354). Furthermore, the oil composition varies within each gland type according to the plant organ. Linalool and linalyl acetate are characteristic substances of flowers, whereas the sesquiterpenes occur in higher proportions in leaves. Even within one gland type on a single leaf, the chemical variability is exceedingly high.     

Development and Essential Oil Content of Secretory Glands of Sage (Salvia officinalis)

Scanning electron microscopy of sage (Salvia officinalis L.) leaves confirmed the presence of two basic types of glandular trichomes consisting of a capitate stalked form containing a multicellular stalk and surmounted by a unicellular secretory head, and a capitate sessile form containing a unicellular stalk and unicellular, or multicellular, secretory head. In the latter type, secretory activity and filling of the subcuticular cavity may begin at virtually any stage of the division cycle affording fully developed glands containing from one to twelve cells in the secretory head. Gas liquid chromatographic analysis of the oil content of the most numerous gland species (capitate stalked, capitate sessile with one and with eight secretory cells) indicated only minor quantitative differences in essential oil composition. Thus, each gland type is capable of producing the four major monoterpene families (p-menthanes, pinanes, bornanes and thujanes) characteristic of sage.     

The content and composition of the essential oil in the course of Anthodium development in wild camomile (Matricaria chamomilla L.)

A study was made of the changes in the content of essential oil and its components (farnesene, (-)-α-bisabolol, (-)-α-bisabololoxide A, (-)-α-bisabololoxide B and spathulenol, α-bisabolonoxide A, chamazulene, cis en-in-dicycloether) and of the number of secretory canals in the receptacles during the beginning of flowering, full flowering and the termination of flowering of camomile anthodia. The total content of the essential oil was highest during the beginning of flowering and lowest during its termination. The proportion of the individual sesquiterpenes in the essential oil underwent changes mostly during the beginning and termination of flowering. The number of secretory canals in the basal and middle anthodial parts increased during the beginning of flowering and the full flowering.     

Ultrastructure and cytochemistry of the pearl gland in Piper regnellii (Piperaceae)

Pearl glands are scattered throughout the lamina of developing leaves and rarely found on adult leaves of Piper regnellii (Piperaceae). The pearl gland is a bicellular secretory trichome composed of a short broad basal cell and a spatula-like, semiglobular apical cell. Four different stages of the pearl gland were determined during its ontogenesis: origin, pre-secretory, secretory and post-secretory. During the pre-secretory stage, mitochondria, ribosomes, dictyosomes, rough endoplasmic reticulum, and plastids with electron dense inclusions were present in the cytoplasm of the apical cell. During the secretory stage, the most remarkable characteristics of the apical cell are the proliferation of dictyosomes and their vesicles, rough endoplasmic reticulum, and modified plastids. At this stage, electron-dense oil drops occur in the plastids as well as scattered within the cytoplasm, proteins and polysaccharides are seen in the plastids, vesicles, and vacuoles. Only polysaccharides are present in the periplasmic space, wall cavities, and on the surface of the apical cell. The polysaccharides are one of the main components of the mucilagenous exudate that covers the developing leaf structures. The apical cell of the senescing trichomes undergoes a progressive degeneration of its cellular components, the plastids being the first organelles to undergo lysis.     

Ultrastructural changes of the teleostean hatching gland cell during natural and electrically induced precocious secretion.

Ultrastructural changes of the hatching gland during electrically induced precocious secretion were compared with those during natural secretion in the medaka, Oryzias latipes. The gland cells are covered by a layer of epithelial cells, which adjoin one another just on the apical center of each gland cell. When the natural as well as the precocious secretion occurred, each gland cell was swollen upward and rounded, and separation of the epithelial joints occurred, giving rise to an exposure of the apical portion of the gland cells. There were marked differences between these two kinds of secretion process in the behavior of the secretory granules prior to secretion and in the mode of discharge of the secretory substances. The changes which occurred during both types of secretion and which, therefore, seemed to be essential to the secretory processes of this gland cell were the swelling up of the gland cells in the initiation of secretion and the reduction of the electron density of the zymogen granules. These secretion-associated ultrastructural changes are discussed in view of the difference in the maturation of the gland cells.     

Organization of monoterpene biosynthesis in Mentha. Immunocytochemical localizations of geranyl diphosphate synthase, limonene-6-hydroxylase, isopiperitenol dehydrogenase, and pulegone reductase

We present immunocytochemical localizations of four enzymes involved in p-menthane monoterpene biosynthesis in mint: the large and small subunits of peppermint (Mentha x piperita) geranyl diphosphate synthase, spearmint (Mentha spicata) (-)-(4S)-limonene-6-hydroxylase, peppermint (-)-trans-isopiperitenol dehydrogenase, and peppermint (+)-pulegone reductase. All were localized to the secretory cells of peltate glandular trichomes with abundant labeling corresponding to the secretory phase of gland development. Immunogold labeling of geranyl diphosphate synthase occurred within secretory cell leucoplasts, (-)-4S-limonene-6-hydroxylase labeling was associated with gland cell endoplasmic reticulum, (-)-trans-isopiperitenol dehydrogenase labeling was restricted to secretory cell mitochondria, while (+)-pulegone reductase labeling occurred only in secretory cell cytoplasm. We discuss this pathway compartmentalization in relation to possible mechanisms for the intracellular movement of monoterpene metabolites, and for monoterpene secretion into the extracellular essential oil storage cavity.     

中华补血草盐腺发育的解剖学研究

采用叶表皮印痕、扫描电镜以及石蜡切片法对中华补血草的叶片进行解剖研究,观察其成熟盐腺的结构、盐腺的发育与盐腺的密度。结果表明:(1)上表皮的盐腺密度比下表皮的略小。(2)成熟的盐腺由20个细胞构成,中央有4个分泌细胞,每一个分泌细胞外侧又伴有一个长方形的毗邻细胞;向外由2层杯状细胞包围,每一层分别有4个杯状细胞,使盐腺呈近似圆形;盐腺内部靠近叶肉细胞处有4个收集细胞;中央4个分泌细胞顶端的角质层各有一小孔,是盐分泌出的通道。(3)中华补血草盐腺是由一个单独的表皮细胞发育而成,分别经历单细胞时期、2细胞时期、4细胞时期、8细胞时期、16细胞时期和20细胞时期的不同发育阶段。     

Bracelet salt glands of the recretohalophyte Limonium bicolor: Distribution,morphology, and induction

Halophytes complete their life cycles in saline environments. The recretohalophyte Limonium bicolor has evolved a specialized salt secretory structure,the salt gland, which excretes Na⁺to avoid salt damage. Typical L. bicolor salt glands consist of 16 cells with four fluorescent foci and four secretory pores. Here, we describe a special type of salt gland at the base of the L. bicolor leaf petiole named bracelet salt glands due to their beaded-bracelet-like shape of blue auto-fluoresc...     

Molecular pathways involved in the cyclic activity of frog (Pelophylax esculentus) Harderian gland: Influence of temperature and testosterone

The Harderian gland of Pelophylax esculentus (previously: Rana esculenta) shows seasonal secretory activity changes. Specifically, the secretory activity reaches a maximum during the hottest months, i.e., July and August, drops in September and slowly increases from October onwards. Expressions of P-CaMKII, P-ERK1 and P-Akt1 correlate well with gland secretory activity; i.e., they peak immediately before the hottest part of the year (maximum secretory activity). When the gland activity declines, kinase expressions drop and remain low until February. Experiments of thermal manipulation indicate that high temperature induces the activation of CaMKII, ERK1 and Akt1, and at low temperatures, Akt1 expression decreases. Experiments of chemical castration indicate that the Harderian gland of Cyproterone acetate-treated frogs shows lower Akt1 activity as compared to controls, but the CaMKII and ERK1 activities remain unchanged. Furthermore, in a period of resumed gland activity (October–December) we observed the highest expression of PCNA, a mitotic marker. Immediately after the proliferative phase, we found the highest expression of caspase 3, an enzyme that plays a key role in apoptosis. In combination, the results suggest the following: 1) CaMKII, ERK1, and Akt1 modulate the annual secretory activity of the frog Harderian gland; 2) CaMKII and ERK1 activities are regulated by temperature, whereas both temperature and testosterone likely play a central role in Akt1 regulation; and 3) proliferation and apoptosis occur to restore and balance, respectively, an adequate cell number, which is essential to gland function.     

Phenotypic and phytochemical differences between Mentha arvensis L. and Mentha canadiensis L.

A taxonomic study of anatomical, morphological, and phytochemical characteristics of Mentha arvensis L. and Mentha canadiensis L. using hierarchical cluster analysis has been conducted and the differences between the species studied have been revealed. The ratio between the lengths of the calyx tube and the calyx lobes, the number of secretory glands on the upper and lower surfaces of the leaf, and the composition of the essential oil were shown to be the most appropriate parameters for classification.     

Regulation of oil accumulation in single glands of Eucalyptus polybractea

The accumulation of terpenoid oil was examined in the leaves of Eucalyptus polybractea at scales ranging from individual oil glands to the whole plant. Variations in oil composition and concentration of oil were measured and related to both morphological and physiological parameters. Within a plant, all glands produced oil of broadly similar composition that was not regulated by leaf age or the position of the gland within the leaf. There were, however, distinct differences between plants, suggesting that composition is controlled primarily at the whole-plant level. Oil concentration, too, was regulated primarily at the whole-plant level and was limited by gland capacity. Gland capacity was linked to leaf area and thickness, the final products of leaf expansion. Leaf and plant oil composition is determined not by a mosaic of glands specializing in producing a single or a small group of compounds, but rather by glands with remarkably similar capacities for terpenoid biosynthesis, although oil concentration, limited by gland capacity, may be linked to leaf expansion rather than biosynthetic capacity.     

Effects of light intensity on the anatomical structure,secretory structures,histochemistry and essential oil composition of Aeollanthus suaveolens Mart. ex Spreng. (Lamiaceae)

Aeollanthus suaveolens Mart. ex Spreng belongs to Lamiaceae family, and in the Amazon this species is cultivated by natives’ people, this medicinal plant is popularly known as Catinga-de-mulata, being used by the population for general pain treatment. The present study analyzed the effects of light intensity on the anatomy, secretory structures, histochemistry and composition of essential oil of leaf and stem of A. suaveolens. The anatomical structure were observed in response to two light intensities, namely 50% (half shade, HS) and 100% (full sun, FS) light. Histochemical analyses were performed to detect lipids, essential oils, alkaloids, phenolic compounds, sesquiterpene lactones, mucilage, and tannins. Secretory structures were observed under a scanning electron microscope. The results obtained in the present work indicate that the light intensity can affect the histochemistry and structures of A. suaveolens. Cross sections of the leaves and stem revealed glandular trichomes on both leaf surfaces as well as the stem surface. Essential oil was detected by histochemical analyses in all types of secretory trichomes. These anatomical and histochemical responses suggest modifications to protect the photosynthetic apparatus from excess light, in addition we note that in the chemical composition of the essential oil the class of hydrocarbons sesquiterpene prevailed.     

北柴胡分泌道的超微结构及其挥发油分泌的研究

利用透射电子显微镜技术研究了北柴胡营养器官中分泌道的发育以及挥发油的积累.并重点探讨了挥发油的分泌机制.结果显示.分泌道细胞的质体、细胞基质以及线粒体参与挥发油或其前体物质的合成,而内质网参与挥发油向分泌道腔隙的转运.在分泌道发育的后期,大量小泡与分泌细胞相邻壁的质膜融合,并将其内的物质释放入此部分细胞壁中,与此同时,此部分壁在靠近腔隙的位置结构变得松散.然后围绕腔隙的壁在相邻两分泌细胞相接的位置面向腔隙外突形成许多小泡,并将小泡释放入腔隙中.由此可见.北柴胡分泌道中挥发油主要以胞吐方式被排入分泌道的腔隙中.     

Antifungal activities of essential oils applied by dip-treatment on areca palm (Areca catechu) leaf sheath and persistence of their potency upon storage

The antifungal activities of cinnamon oil, clove oil, anise oil, and peppermint oil, and their main components (cinnamaldehyde, eugenol, trans-anethole, and menthol, respectively) against molds identified from areca palm leaf sheath (Mucor dimorphosporus, Penicillium sp., Aspergillus niger, and Rhizopus sp.) were investigated. An agar dilution method was employed to determine the minimum inhibitory concentration (MIC) of essential oils and their main components. Zone inhibition tests and the inhibitory effect of the leaf sheath dip-treated with essential oils against those molds were examined. Major components of essential oils on the leaf sheath during storage were quantified by gas chromatography analysis. The MIC values of essential oils on agar and on the leaf sheath were identical. With an MIC of 50 ??g ml⁻¹, cinnamon oil had the strongest inhibitory effect. At their MICs the oils were capable of providing protection against mold growth on the leaf sheath for at least 12 weeks during storage at 25 °C and 100% RH. Scanning electron microscope examination showed that essential oils prevented spore germination. Except for menthol in peppermint oil, the main components of the essential oils, which were fairly stable over the storage period, largely contributed to the antifungal activity.