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Anatomical and Pollen Ornamentation Study on Hymenocrater species in North East of Iran

A. Jafari and F. Jafarzadeh
The present study tends to investigate the anatomy and palynology of Hymenocrater species in Northeast of Iran. To conduct the comparative study of anatomy characters, sections from stem and leaf were prepared using microtome and differential staining. In this part of investigation, arrangement of vessel in stem, stoma type and arrangement of mesophyll in leaf were studied. For the palynology study, too a comparative investigation on the species showed, the pollen was problate spheroidal, hexacolpate, bireticulate and semi- tectate. Finally a variation between the shape of lumina in eu-reticulate and supra reticulate of pollen was observed.
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A. Jafari and F. Jafarzadeh, 2008. Anatomical and Pollen Ornamentation Study on Hymenocrater species in North East of Iran. Pakistan Journal of Biological Sciences, 11: 2149-2153.

DOI: 10.3923/pjbs.2008.2149.2153



Hymenocrater Fisch and Mey. from Lamiaceae family, Stachyioideae subfamily, having numerous variety of species, it is expanded from Iran to Iraq, Pakistan, Afghanistan (Parsa, 1949; Groshkova, 1954; Jalas, 1972; Boissier, 1975; Davis, 1982; Rechingr, 1982; Hedge, 1990). Hymenocrater has nine species in Iran (Rechingr, 1982), about four of which are found in north-east of Iran. About 55% species of Hymenocrater is endemic for Iran (Naghibi et al., 2005). This genus has aromatic essential oils and antimicrobial effects (Groshkova, 1954; Firoznia et al., 2005; Zadidi and Crow, 2005). The purpose of present study was to investigate variation of internal structure and identification of Hymenocrater species on the basis of micromorphology and internal structure because morphologically identification of this species is difficult. This respect we collected fresh plants from their localities. Then we prepared some cross section of stem and leaf of H. platystegius, H. calycinus, H. elegans and H. bitominosus. In terms of comparative anatomic studies on Hymenocrater, there has been a report about H. bituminosus (Satil et al., 2007). The other reports are about Salvia and Cyclotrichum from Lamiaceae (Metcalfe and Chalk, 1983; Kaya et al., 2000). In palynological studies, the comparison were made among the pollen grains of Hymenocrater. In this part of study, pollen grains were extracted and acetolysed , to study shape and ornamentation of the pollen through SEM and LM. The previous palynology investigation had been done in Salvia, Origanum and Lycopus (Moon and Hong, 2003; Akyalcin, 2003). So, comparative palynology of Hymenocrater carried out for first time in Iran.


As for the anatomic study, the examined species were collected from the localities in north-east of Iran during May- June 2005-2006 (Table 1). For preparing of cross section of stem and leaf, base of stem and basal leaves were selected from 8-9 specimens. The fresh specimens were fixed in FAA then, dehydrated with ethanol and later, some slices prepared with microtome. The section- 12 micron thick-stained with Safranine and Fast-green (Johnson, 1940; Chamberlain, 1990). For the stem maceration, this organ was placed in Jeffery solution for four hours to soften In the palynological study, the pollen was extracted from the anther and dehydrated by glacial acetic acid, then acetolised, coated with sputter finally studied by LM Olympus and SEM LEO1450VP. (Erdtman, 1971: Moore et al., 1991). The pollen terminology was adapted from Punt et al. (1994). Finally, clustering analysis carried out with using JUMP.4 software for identifying close species.

Table 1: The localities of studied Hymenocrater species

Fig. 1: Stem cross section of H. elegans; a: epidermis layer, b: collenchyma layer, c: parenchyma layer, d: sclerenchyma layer, e: suber laye and f: phloem tissue

Fig. 2: Stem cross section of H. platystegius. The arrow showing solitary vessel (x 416)

Fig. 3: Stem cross section of H. calycinus. The arrow showing radial chain pore (x 416)
Fig. 4: Stem cross section of H. bituminosus. The arrow showing solitary and cluster (x 416)

Fig. 5: Dorsi- ventral mesophyll in leaf cross section of H. bituminosus (x 940)

Fig. 6: Isolateral mesophyll In leaf cross section of H. calycinus (x 416)


Anatomic results: The results from the anatomic studies demonstrated the stem with following tissues:

Some angular collenchymatous layers with small cells under the epidermis.

Fig. 7: Diacytc stoma type in H. bituminosus

Fig. 8: Anemocytic stoma type in H. platystegius

Fig. 9: Anisocytic stoma type in H. calycinus

Some parenchymatous layers with large cells
Some sclerenchymatous layers with small cells.
Suber layer under the sclerenchymatous layers. (Fig. 1 ).
Vascular bundles which the arrangement of vessel was variable for example solitary in H. platystegius (Fig. 2), radial chain pore in H. calycinus (Fig. 3), solitary and cluster in H. bitunimosus (Fig. 4) and cluster- radial chain pore in H. elegans. In all of studied species, sclerid and fiber observed. The results from stem maceration confirmed the presence of vessel with simple perforation plate.

Fig. 10: The pollen of H. elegans: (a) apocolpium m: mesocolpium (c) colpium (x 5000)

Fig. 11: The pollen of H. elegans (x 20000)

Fig. 12: The pollen of H. platystegius (x 2000)

Fig. 13: The pollen of H. calycinus (x 2000)

Table 2: The anatomic and pollen characters of Hymenocrater studies species

Fig. 14: The pollen of H. bituminosus (x 5000)

Fig. 15: Dendrogram on the basis of anatomy and palynology characters

The results from the anatomic study of the leaf showed, the arrangement of mesophyll was dorsi-ventral except H. calycinus (Fig. 5, 6). The stoma type was diacytic in H. bituminosus, diacytic with anemocytic in H. platystegius, diacytic with anemocytic and anisocytic in H. calycinus (Fig. 7-9).

Palynology analysis: The pollen was problate spheroidal, bireticulate, hexacolpate, semi-tectate and agranulate (Fig. 10). The lumina of supra reticulate in H. elegans was uniseriate and oblong (Fig. 11), in H. platystegius rectangular (Fig. 12). The lumina of eu-reticulate was circular except H. calycinus which narrow and very oblong was observed (Fig. 13, 14). The shape of lumina in apocolpium was circular but in H. elegans was tetrahedral.

Finally using 46 character of anatomy and the pollen clustering analysis carried out and prepared dendrogram (Fig. 15, Table 2).


As the results from the anatomic studies about Hymenocrater suggest, we can state the anatomic structure of the stem in all species under the investigation, were similar. Also, their internal structures were similar to Salvia and Cyclotrichum (Metcalf and Chalk, 1983; Kaya et al., 2000). Satil observed just radial chain pore the arrangement of vessel in H. bituminosus and he didn`t report suber layer under the sclerenchymatous tissue. Also he identified only diacytic stoma type (Satil et al., 2007) but present research results showed cluster- radial chain pore, suber layer, stoma type as diacytic and anemocytic. Also, only, the arrangement of mesophyll in H. calycinus was isolateral because, growing in stony mountains and rest of them were dorsiventral. The morphology of pollen in Hymenocrater was similar to Lycopus i.e., hexacolpate and bireticulate (Moon and Hong, 2003). Just differences observed among the species as size, shape of lumina.

Clustering analysis on the basis of anatomic and palynology results showed the closest species were H. platystegius and H. bituminosus because both of them were similar as arrangement of mesophyll, vessel and the shape of lumina. This results demonstrated, H. calycinus is different from the others, due having isolateral arrangement of mesophyll, compound stoma type, radial chain pore arrangement of vessel and circular in lumina at apocolpium. In conclusion using of micromorphology and anatomic characters can be useful for identifying of species.

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