Anatomical Structure Study of Aerial Organs of Crataegus ambigua C . A . Mey

The purpose of the present research was to study the peculiarities of the anatomical structure of some vegetative and genesic organs of Crataegus ambigua C.A.Mey. growing in the territory of Mangistau region. The findings of investigation allowed determining that the annual sprouts and pedicel of this species have a primary anatomical structure with the transition to secondary growth. Leaf of the Crataegus ambigua is of a dorsoventral type of light mesophytic structure, hypostomatic in terms of stomata arrangement. The fetal epidermis is presented by polygonal cells of the epidermis without additional elements. The pulp consists of large and loosely-arranged parenchymal cells. The epidermis of the corolla is represented by oval thick-walled cells. The pericarp of the seed consists of three well-defined layers (exocarp, mesocarp and endocarp). The corcule occupies the entire internal cavity. In terms of the cover and pulp structure, Crataegus ambigua has a structure similar to other species. Characteristic features for this species are the shape and the arrangement of the main cells of the epidermis, stomata and simple unicellular trichomes.

Anatomical features of Crataegus genus plants are underinvestigated.Thus, there are just single works on wood structure and stamens of the flowers 11,12 .The fruit structure is a more studied issue as a pharmacognostic character of medicinal raw material 13,14 .However, the anatomical structure of the leaves and young sprouts are completely absent.
Crataegus ambigua C.A.Mey. is a rare and endemic plant of the flora of Mangystau region.The habitat of this species is confined to the grounds and gentle slopes of the gorges of the Mangyshlak Peninsula and Tubkaragan 15 .Anatomic features of this plant are to date unexplored.
The purpose of the present research was to study the peculiarities of the anatomical structure of some vegetative and genesic organs of Crataegus ambigua growing in the territory of Mangistau region.

Methodology
The investigation objects were aerial vegetative (leaf and annual sprouts) and genesic (flowers and fruit) organs of middle-age Crataegus ambigua plants growing in Mangystau (Akmysh, Samal and Kezym mountain gorges).
Sampling was carried out during the vegetation periods of the years 2015-2016 in the phase of large flowering and fruiting.Freshly harvested organs were preserved in a mixture of alcohol (70%), glycerin and distilled water in 1:1:1 ratio (Strauss-Fleming solution).When determining the anatomical features of the leaf blade of the studied species, we selected the most developed intact leaves in the middle part of the sprouts, as well as analyzed the fragments (surface preparations and transverse sections) in the middle part between the main nervure and the edge.When studying flowers, we investigated the surface preparation of the corolla and cross-section of the pedicle.Transverse sections of annual sprouts were made throughout their length every 2-3 cm.For fruit, we investigated the peel surface preparation and transverse sections of pulp and seeds.Not less than 10-15 micro specimens were prepared for each organ.
Anatomical study of the plant was conducted according to the recommended practices described by M.N.Prozina 16 , A.A. Dolgova and E.Ya.Ladygina 17 , V.N.Vachov and L.I. Lotov 18 .Fabrication of temporary preparations (surface and pressure preparations, transverse sections) were performed according to standard techniques [19][20][21] using "MZP-01 Tehnokom" freezing microtome.Bleaching of the preparations was performed using the glycerol.To obtain surface preparations, leaves were boiled in 10% potassium hydroxide solution.
The obtained preparations were studied by means of Melji-Techno "MT 4310 L" scanning microscope and BisionCamV 500B camera.Digital photographs were obtained at the magnification of eyeglass and lens by 10E4, 10x10, 10x40, employing the VisualBio program.When studying leaf epidermis, drawings were performed from photos manually.

RESULTS AND DISCUSSION
Crataegus ambigua is a shrub or small tree [26].Sprouts are glabrous, inerm, rarely with spines, branches are brown with gray patches of preserved peel, leaves are bright green, the lowermost leaves are brevipetiolate, cuneate obovate, trilobate or incisodentate at the apex, upper leaves are petiolate, ovate, with broadly cuneate or rounded base, 5-7-lobed.Inflorescences are dense with up to 20 flowers.Fruits are globose or broadly elliptical, purple-black with light dots, fleshy; bones in outline are broadly elliptical.

The anatomical structure of Crataegus ambigua annual sprouts
Cross-sections of young sprouts of Crataegus ambigua have a primary anatomical structure with the transition to secondary growth (Fig. 1).Outside, the stem is covered with onelayered epidermis 19-23 µm thick consisting of the cells with rounded-rectangular structure, which are covered on top with thick cuticle.Under the epidermis there is a three-layer lamellar collenchyme 83-98 µm thick stained in a dark color.
The area between the endoderm and the cortex is filled with 5-6 rows of loose parenchymal cell.The thickness of this zone ranged from 148 to 240 µm.Endoderm is arranged in a single row and its cells are tangentially thickened There are numerous conducting bundles    upper and lower epidermis are well seen in a crosssection (Fig. 2).They are thin-walled, covered from outside with a thin layer of cutin.The thickness of the upper epidermis was 23-25 µm, while that of the lower one was 18-20 µm.
The mesophyll is differentiated into palisade and spongy tissues.Cells of palisade mesophyll are arranged in 1-2 rows 45-51 µm thick, the intercellular ducts are not expressed.Palisade mesophyll over midrib is interrupted by angular collenchyma.The transition to the spongy mesophyll is clear, intercellular ducts are of aerenchyme type 78-90 µm thick.Collateral bundles of closed type (cambium is absent) are formed only along the nervures and consist of 10-15 rows of xylem and 5-8 rows of phloem.From the phloem side, conducting bundle is lined with 2-3 rows of sclerenchyma as a cap.The bundles area made up 405-453 10 -3 mm 2 .The hawthorns are characterized by variability in shape of the lower part of the main nervure.Some leaves are rounded in cross section, while others have projections in the shape of the "ears".
The cells of the upper epidermis have slightly sinuous membranes (Fig. 3) with expressed thickened walls.Cells length is 0.46-0.70µm, width -0.30-0.35µm.Some rare simple single-celled trichomes about 4 µm in length with the base thickness of 0.38-0.42µm are effused over the surface Cells of the lower epidermis are more elongated with strongly sinuous membranes 0.47-0.59µm in length and 0.19-0.30µm in width.Walls are thin and surface is covered with a cuticle layer.On the underside of the leaf there are anomocytic stomata, 0.40-0.47µm in length and 0.16-0.22µm in width.The number of stomata is 9-10 pieces/mm 2 .Trichomes are absent.Thus, Doubtful Hawthorn is characterized by hypostomatic type of leaf.
Above the nervure, the cells of epidermis have prosenchymatous shape.Nervures on both sides of the leaf are surrounded by the rows of sheath, whose cells contain crystals of calcium oxalate.

Anatomical structure of Crataegus ambigua fruits
Drupe of the hawthorn contains 2 to 3 seeds, 5-6 mm long and 3-4 mm wide.The weight of 1,000 seeds ranges from 12 to 16.5 g.The seed is oblong with the slightly recurved tip, bluntly cut at upper end.It has 1-2 vague ribs with color ranged from yellow to light brown (Fig. 4).
In the structure of the seed one can clearly distinguish the pericarp, endosperm and cotyledons.Pericarp of the seeds consists of 3 layers (exocarp, mesocarp and endocarp) 10-16 µm thick.
Exocarp is thin, covered with cuticle.It consists a single layer of dead cells.Cells of mesocarp are thin-walled, densely arranged, parenhimy.Endocarp is very weak, comprises of thin-walled living cells arranged in a single layer.The cells of the endosperm 109-116 µm thick have the same rounded shape arranged in 8-10 rows.
The epidermal cells of the cotyledon are elongated, large, arranged in a single layer 570-575 µm thick.Embryo is 2-cotyledonous, elongated, disk-shaped, the cut completely captures the internal cavity.
The rind of the hawthorn fruit (Fig. 5) consists of polygonal cells with thickened walls and yellow-brown contents, tightly adjacent to each other.
The cells length was 0.20-0.37µm, width -0.19-0.26µm.The surface of epidermis is covered with a layer of cuticle.
The pulp of the fruit (Fig. 6) consists of round or oval cells, 0.96-1.15µm in length and 0.61-0.90µm in width, containing inclusions of orangered or brownish-yellow color (carotenoids), small druses and prismatic crystals of calcium oxalate The inner part of the fruit pulp is shot through with collateral bundles and solitary sclereids.Layers of stone cells are arranged near the large bundles; calcium oxalate crystals form in patches crystalliferous sheath.

The anatomical structure of Doubtful Hawthorn flower
The corolla of the flower on the surface preparation consists of oval cells of 0.28-0.36µm in length and 0.15-0.22µm in width (Fig. 7).Walls are thick and surface is covered with a layer of cuticle.Stomata and trichomes were not identified At cross-section, pedicel of hawthorn is rounded, 9-10 µm in diameter (Fig. 8).Cells of the epidermis, rounded at cross section, 0.16-0.18µm in diameter are arranged peripherally.Beneath are 3-4 layers of collenchyma laid after 3-5 layers of primary cortex 1.22-1.49µm thick.Cells are of parenchymal type with large lumens and loose arrangement Primary cortex is isolated from the central cylinder by 0.20-0.41µm thick endodermis layer, whose cells are tangentially thickened.Conducting system is of a bundle type.The bundles are collateral, closed, with area of 0.63-0.88 10 -3 mm 2 .Each beam is covered by the sclerenchyma "cap" 0.26-0.32µm thick.The central part is filled with medullar parenchyma with the cells diameter of 0.28-0.32µm.

CONCLUSION
Thus, on the basis of conducted research we can draw the following conclusions: 1.
The stem and pedicel of Crataegus ambigua are of primary anatomical structure, while transition to secondary growth was observed for stem.

2.
The leaf of Crataegus ambigua is of a dorsoventral type of light mesophytic structure, hypostomatic in terms of stomata arrangement.Its characteristic features are the shape and arrangement of the main epidermal cells, stomata and single-cellular trichomes.

3.
The fetal epidermis is represented by polygonal cells of the epidermis without additional elements.The pulp consists of large and loosely-arranged parenchymal cells.

5.
Epidermis of the corolla is represented by oval thick-walled cells without additional structures.