Structural Investigation of the Glandular Trichomes of Endemic Salvia Smyrnea L

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South African Journal of Botany 76 (2010) 572–578 www.elsevier.com/locate/sajb

Structural investigation of the glandular trichomes of endemic Salvia smyrnea L. ⁎ P. Baran, K. Aktaş , C. Özdemir

Celal Bayar University, Faculty of Art and Science, Department of Biology, 45030-Muradiye, Manisa, Turkey Received 17 May 2009; received in revised form 4 November 2009; accepted 23 April 2010

Abstract

The morphology, anatomy and distribution of glandular trichomes on the aerial organs of Salvia smyrnea L. endemic to Turkey have been investigated with light microscopy (LM) and scanning electron microscopy (SEM). This species is evaluated in endangered (EN) category. Two morphologically distinct types of glandular trichomes were determined. Various types of capitate glandular trichomes consist of a 1–4 celled base, a1–8 stalk celled or no stalk and a uni- or bicellular head. In capitate trichomes the neck cell that has an important role especially for xeromorfic plants, acting to prevent the backflow of secreted substance through the apoplast, has been distinctively observed in the investigated species. The capitate trichomes were present abundantly on all aerial organs of S. smyrnea. Peltate glandular trichomes had a large secretory head comprising 1–2 central and 4,6, 8–10 peripheral cells. This study pointed out that S. smyrnea was of rich glandular trichomes in point of diversity and quantity. © 2010 SAAB. Published by Elsevier B.V. All rights reserved.

Keywords: Capitate trichome; Glandular trichome; Lamiaceae; Peltate trichome; Salvia smyrnea

1. Introduction which can be distinguished by head size and stalk length. As a rule, in a capitate trichome, the length of the stalk should be The essential oil produced by glandular trichomes is one of more than half the height of the head, whereas peltate trichomes the features characteristic for Lamiaceae family (Ascensão et are short with a uni- or bicellular stalk and a large secretory head al., 1995). Glandular trichomes that develop from epidermal with 4 to 18 cells arranged in one or two concentric circles. cells are generally considered as the site of biosynthesis or Capitate trichomes are very variable in stalk length, head shape accumulation of essential oils (Nishizawa et al., 1992; Turner et and secretion process and can be subdivided into various types al., 2000). Volatile oil produced by glandular trichomes is (Werker et al., 1985a). important for pesticide, pharmaceutical, flavouring, fragrance In the Lamiaceae family, the morphology, distribution and and cosmetic industries (Serrato-Valenti et al., 1997; Zeybek and frequency of glandular trichomes are used as discriminative Zeybek, 2002). These glandular trichomes are widely distributed characters at subfamiliar level (Ascensão et al., 1995). over the aerial reproductive and vegetative organs of the plants of According to Bisio et al. (1999) most of the aromatic genera Lamiaceae. They are the primary secretory organs of these plants are in the subfamily Nepetoideae which includes also the genus and their structures can vary widely among species (Serrato- Salvia L. Furthermore Metcalfe and Chalk (1972) emphasized Valenti et al., 1997). Ascensão and Pais (1998) determined the that Salvia is the genus that has the largest number of glandular two main types of glandular trichome, peltate and capitate, occur trichomes of the family. Volatile oil that provides a special fragrance is characteristic of many Salvia species (Stace, 1991). Salvia species are an important group of useful plants which have not lost their importance since ancient times. The genus ⁎ Corresponding author. Fax: +90 236 2412158. Salvia derived from “Salveo” which means “to save, to recover” E-mail address: [email protected] (K. Aktaş). in Latin (Hamlyn, 1969). Salvia, commonly known as sage, has

Fig. 1. Photographs of peltate glandular trichomes: cu. cuticle, gs. glandular space, pc. periphery cells, cc. central cells. multiple uses such as food additive, spice and herbal tea Taxonomical description of the plant followed Hedge (Demirci et al., 2005). Besides their medicinal value, Salvia sp. (1982). The paraffin method was applied for preparing cross- are also grown in parks and gardens as ornamental plants sections of the mature plant organs as petiole, leaf, pedicel, (Nakipoğlu, 1993). The antibacterial, antituberculous and calyx, corolla using rotary microtome (Microm HM 325). The antiphlogistic activities of the constituents of Salvia species thickness of the cross-sections prepared with microtome was are well established (Ulubelen et al., 2001). 15 µm (Algan, 1981). The cross-sections taken by hand were Salvia, the largest genus of the family represents an enormous stained in Sartur reagent (Baytop, 1981). Trinoculer light and cosmopolitan assemblage of nearly 1000 species displaying a microscope (Olympus BX50F-3) with camera (Olympus PM-C remarkable range of variation. The genus comprises 500 spp. in 35) was used for taking photographs of the cross-sections. For Central and South America, 250 spp. in Central Asia/Mediterra- scanning electron microscope investigation small petiole, leaves, nean, and 90 spp. in Eastern Asia (Walker et al., 2004). The first pedicel, calyx, and corolla were coated with a thin layer gold revision of Salvia species in Turkey was made by Hedge (1982), in POLARON SC7620 Sputtering Device. Observations were whorecognized86species(Davis et al., 1988). Since then, six new carried out on a JEOL JSM 6060 scanning electron microscope. species have been described (Huber-Morath, 1982; Vural and Results were presented in photographs, micrographs and tables. Adıgüzel, 1996; Dönmez, 2001; Hamzaoğlu et al., 2005; Ilcim et al., 2009; Celep and Doğan, 2010) and three new records have been reported from Turkey (Behçet and Avlamaz, 2009; Celep et al., Table 1 2009; Kahraman et al., 2009); the total has now reached 95. Forthy Cell numbers of parts of peltate trichomes found on organs of of Salvia eight of these Salvia species in Turkey are endemic. Salvia smyrnea smyrnea L. is an endemic species to Turkey and is recognized by the Peltate trichomes International Union for Conservation of Nature (IUCN) as endangered category (Ekim et al., 2000). Centre Periphery Centre Periphery Any study on the glandular trichomes of S. smyrnea that is Stem 1 4 Corolla 1 4 the subject of this paper has not been found in literature. Of the 2 6 16 18 great number of Salvia species which have not been examined Leaf 1 4 2 6 with respect to the structural properties of their glandular 16 trichomes, we aimed to investigate the morphology, cell number 1 8 Pedicel 1 8 and distribution of glandular trichomes of S. smyrnea. 28 14 19 16 2. Materials and methods Calyx 1 4 1 6 Petiole 1 6 The plant samples were collected in 2003 from the following 18 18 location: İzmir: Kemalpaşa, and Nif mountain, around the fire 110 26 tower, 1500 m, 07.2003 (Baran 015). 28 18 574 P. Baran et al. / South African Journal of Botany 76 (2010) 572–578

Table 2 in which the secreted material accumulated, developed by the Cell numbers of parts of capitate trichomes found on organs of Salvia elevation of the cuticle together with the outermost layer of the smyrnea L. secretory cell walls. Two different ways of the release of Capitate trichomes secreted product were seen. They were present on the stem, Type I Type II Type III leaf, pedicel, corolla, calyx and petiole, especially being Base Stalk Head Base Stalk Head Base Stalk Head abundant on leaves and calyx both in diversity and density – Stem 1–30,2–41–21–31–2, 4–51 1–43–4,7 1 (Fig. 1; Fig. 4M U; Table 1). Petiole 1–40–51 1–31–811–31–51 Leaf 1–31–61–21–21–2, 4–51 1–31–51 3.2. Capitate trichomes Pedicel 1–22–51–21–31–311–32–41 – – – – – – – – Calyx 1 20,231213141213251 Capitate trichomes greatly varied in structure, size and Corolla 1–2 0,2,4 1 1–32–411–30,3–41 secretion mode, which were consisted of a base formed by 1–4 epidermal cells, 1–8 stalk celled, usually with a short neck cell, The classification of glandular was made according to Werker or no stalk and a uni- or bicellular head (Table 2). The neck cell et al. (1985a) and Özdemir and Şenel (2001). with thickened walls was adjacent to the head and usually short in length and appeared as filled with the secreted product. We 3. Results observed three types of capitate trichomes. Type I had uni- or bicellular round head with droplets on the cuticle, with very Glandular trichomes were of two main types, peltate and small subcuticular space or without subcuticular space (Fig. 2; capitate, being different in size, structure and mode of secretion Fig. 4B–D); type II had a pear-like or round head and larger (Fig. 4A–L). subcuticular space filled with secreted material and subsequent broken cuticle which was obviously elevated forming large 3.1. Peltate trichomes subcuticular space surrounding the cell until the base of it (Fig. 3A–D; Fig. 4E–G); type III had a cup-like head cell with Peltate glandular trichomes had a base including epidermal a broken cuticle (Fig. 3E–J; Fig. 4H–L). Type I, II and III cells and a very short stalk cell and a large secretory head, capitate trichomes were all observed on stem, leaf, petiole, comprising 1,2 central cell and 4,6, 8–10 peripheral secretory pedicel, calyx and corolla, especially being abundant on res- cells arranged in two concentric circles, which is a typical pectively pedicel, calyx, leaves and petiole both in diversity and feature in the Lamiaceae. At the time of secretion a large space, density.

Fig. 2. Photographs of type I capitate glandular trichomes: hc. head cell, nc. neck cell, sc. stalk cell, cu. cuticle, s. secretory. P. Baran et al. / South African Journal of Botany 76 (2010) 572–578 575

Fig. 3. Photographs of capitate glandular trichomes (A–D: type II, E–J: type III): hc. head cell, nc. neck cell, sc. stalk cell, cu. cuticle, bc. base cell, ss. subcuticular space.

4. Discussion in Salvia aurea L, which occurred in S. smyrnea as type I and III. In Nepeta racemosa L. (Bourett et al., 1994)andS. aurea Recently some researchers observed peltate and various (Serrato-Valenti et al., 1997), small capitate glands with two head capitate trichomes on some Salvia species (Baran et al., 2008; cells were found. In our study, the capitate trichomes with a Moon et al., 2009; Özkan, 2008). As in plants of most bicellular head had a 1–3 stalk celled or no stalk cell, while Lamiaceae species, the aerial parts of S. smyrnea, especially the capitate trichomes with a bicellular head in S. sclarea (Özdemir leaves and flowers were densely covered glandular trichomes, and Şenel, 1999) had no stalk cell and those in S. forskahlei including two main types, peltate and capitate trichomes, which (Özdemir and Şenel, 2001)hada1–2 stalk celled. Özdemir and differed in structure and secretion process. Şenel (1999, 2001) were the first authors to report the base cell A similar arrangement in periphery cells of peltate trichomes number of capitate glandular trichomes in the genus Salvia.We in our study is also determined in Origanum L. species also determined the base cell number of capitate glandular (Bosabalidis and Tsekos, 1984), Satureja thymbra L. (Bosaba- trichomes of S. smyrnea in the present study. The capitate lidis, 1990), Salvia forskahlei L. and S. sclarea L. (Özdemir trichomes of S. smyrnea had 1–4 celled base, while they and Şenel, 1999, 2001) and other species of the Lamiaceae possessed 1–2celledbaseinS. sclarea and S. forskahlei examined by Werker et al. (1985a,b), but in other species of the (Özdemir and Şenel, 1999, 2001). Other researchers deter- same family, such as Leonotis leonurus (L.) R. Br. (Ascensão mined the base cell number as 1–2, 1–5, 1–3inSalvia huberi et al., 1995)andOcimum basilicum L. (Werker, 1993), a small Hedge, S. viridis L., S. recognita Fisch. & Mey. respectively number of head cells are arranged in a single circle. (Baran et al., 2008; Özdemir and Altan, 2005; Özkan and Soy, The types of capitate glandular trichomes observed in S. 2007). smyrnea corresponded respectively to the three types of capi- The morphology of glandular hairs is often related to the tate trichomes described by Werker et al. (1985a) in their study kind of secreted product. In some Lamiaceae secretion from of some Lamiaceae species, including the Salvia. Serrato-Valenti capitate trichomes contains mostly polysaccharides and only et al. (1997) observed two types of capitate glandular trichomes small quantities of essential oil. On the other hand, the peltate 576 P. Baran et al. / South African Journal of Botany 76 (2010) 572–578 trichomes are considered the site of production and storage of In the peltate trichomes of S. smyrnea another, less common essential oil (Bini Maleci et al., 1983; Werker et al., 1985a). release mechanism has been observed as well, namely droplets Furthermore, Werker et al. (1985b) reported that the different of secretory material were seen on the external surface of the morphological structures of some capitate trichomes might head, which appeared to pass through the cuticular layer. correspond to the production of the different materials. On the In type I capitate trichomes of S. smyrnea, droplets of other hand, by a functional point of view, Werker (1993) secreted material were seen on the head cell (Fig. 2), which was classified the glandular trichomes into two types, according to released by cuticular exudation. A similar phenomenon different modes and timings of secretion: the short-term was observed in the secretory trichomes of Inula viscosa L. glandular trichomes, which start and end secretion rapidly and (Werker and Fahn, 1981), Majorana striaca L. (Werker et al., the long-term glandular trichomes, in which secretory material 1985a) as well as S. aurea (Serrato-Valenti et al., 1997) and S. is accumulated in a large subcuticular space. In the peltate blepharophylla Brandegee ex Epling (Bisio et al., 1999). In trichomes of S. smyrnea, the secretory product remained trapped type II capitate trichomes of S. smyrnea, accumulation of the between the cuticular sheath and the head cells, following a secreted product into an extended subcuticular space by cuti- pattern already described for other aromatic Lamiaceae (Bosaba- cular elevation and subsequently the rupture of cuticle was lidis and Tsekos, 1982; Bourett et al., 1994; Bruni and Modenesi, observed, as reported in Salvia officinalis L. (Corsi and Bottega, 1983; Werker et al., 1985a,b). As proposed by Ascensão et al. 1999), S. blepharophylla (Bisio et al., 1999), S. dominica L., S. (1995), external factors such as high temperatures, low air sclarea (Werker et al., 1985b), S. officinalis, S. fruticosa Miller. humidity or animal aggression may cause the cuticular rupture. and some other species of Lamiaceae (Werker et al., 1985a). In

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