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Analysis of the leaf surface in Globularia meridionalis (Podp.) O. Schwarz and Globularia punctata Lapeyr. using optical and environmental scanning electron microscopy (ESEM)

The genus Globularia belongs to the family of Plantaginaceae (former Globulariaceae) and includes 22 to 23 species distributed mainly in the Mediterranean area. Some members of the genus are known for their biological properties and are used for a variety of therapeutic actions such as hypoglycaemic, laxative, cholagogue and other [1]. The characteristic anatomical feature of the family are small glandular hairs that have a short stalk cell and 2 (rarely 4) convexly arched cells separated from one another by vertical walls [2]. The aim of this study was to record and compare the structure of the leaf surface of two species growing in Croatia, G. meridionalis (Podp.) O. Schwarz and G. punctata Lapeyr. [3], with emphasis on the structure, size and layout of the glands. To our knowledge, precise data and images for the abovementioned species do not exist. The analysis of the surface was made by using an optical microscope and an environmental scanning electron microscope (ESEM). Sudan III was used as a staining agent for lipophilic compounds. The determination of gland size was based on the measurement of height, length, width and area of ten different glands on the lower surface of the leaf, in triplicate. The results of the analysis showed that both species contain glandular hairs on both upper and lower surface of the leaf, which are sunk in pits in the epidermis. The glands are built of a short stalk cell and a bicellular head, as described in the literature [2]. In G. meridionalis glands were found to be approximately 34, 5µm high compared to 20, 5µm in G. punctata. The observed surface area of the gland head was about 664, 6µm2 in G. meridionalis and 530, 5µm2 in G. punctata. The results of all performed measurements are expressed as mean value ± standard error and are shown in Table I. The Mann-Whitney test showed that there was a significant difference (P<0, 05) between the height and surface area of glands in the investigated species (P<0, 0001 ; P=0, 0007), as well as their length measured in superficial section (P=0, 0029) and width measured in transverse section (P=0, 0412). Stomata are widely present on both leaf surfaces and are anomocytic, which corresponds to the literature [2]. The epidermal cells have sinuous shape, with an extremely thick outer cell wall, especially in G. meridionalis. Staining with Sudan III showed the presence of a thin cuticle layer on the epidermal surface, as well as the presence of oily substances produced by the glands. The ESEM analysis showed a difference between the lower leaf surface of the two species, which was smooth in G. meridionalis and striated in G. punctata. Further research considering the inner structure of the leaves as well as other plant organs of the two species are in progress.

electron microscopy; ESEM; Globularia; leaf; glands

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