Multi-omics analysis of the bioactive constituents biosynthesis of glandular trichome in Perilla frutescens.

BACKGROUND: Perilla frutescens (L.) Britt is a medicinal and edible plant widely cultivated in Asia. Terpenoids, flavonoids and phenolic acids are the primary source of medicinal ingredients. Glandular trichomes with multicellular structures are known as biochemical cell factories which synthesized specialized metabolites. However, there is currently limited information regarding the site and mechanism of biosynthesis of these constituents in P. frutescens. Herein, we studied morphological features of glandular trichomes, metabolic profiling and transcriptomes through different tissues. RESULTS: Observation of light microscopy and scanning electron microscopy indicated the presence of three distinct glandular trichome types based on their morphological features: peltate, capitate, and digitiform glandular trichomes. The oil of peltate glandular trichomes, collected by custom-made micropipettes and analyzed by LC-MS and GC-MS, contained perillaketone, isoegomaketone, and egomaketone as the major constituents which are consistent with the components of leaves. Metabolomics and transcriptomics were applied to explore the bioactive constituent biosynthesis in the leaves, stem, and root of P. frutescens. Transcriptome sequencing profiles revealed differential regulation of genes related to terpenoids, flavonoids, and phenylpropanoid biosynthesis, respectively with most genes expressed highly in leaves. The genes affecting the development of trichomes were preliminarily predicted and discussed. CONCLUSIONS: The current study established the morphological and chemical characteristics of glandular trichome types of P. frutescens implying the bioactive constituents were mainly synthesized in peltate glandular trichomes. The genes related to bioactive constituents biosynthesis were explored via transcriptomes, which provided the basis for unraveling the biosynthesis of bioactive constituents in this popular medicinal plant.

Microscopic and phytochemical techniques as a tool for authentication of herbal drug chiraita: Swertia cordata (G. Don) C.B. Clarke.

Swertia cordata (G. Don) C.B. Clarke is one of the potential medicinal plants extensively used in eastern traditional medicine such as Unani, Ayurveda, Siddha, and in traditional Tibetan and Chinese medicine. S. paniculata is the common adulterant of S. cordata at herbal shops and markets but S. paniculata is also used in number of herbal formulations. The present study was conducted to use microscopic, pharmacognostic, and phytochemical techniques as a tool for the authentication of herbal drug chiraita (S. cordata). In herbal markets, mixing, adulteration, and use of spurious materials as substitute have become a major concern for herbal practitioners, local user, and industry for reasons of safety and efficacy. Therefore, authentication of medicinal plants is of utmost importance at each level of drug research. In the present study, anatomical features of two species showed a great diversity, as irregular epidermal cells and nonglandular, unicellular trichomes were found in S. cordata while in S. paniculata epidermal cells were hexagonal in shape and trichomes were A-shaped. Antioxidant activity of two species showed a great variation where IC50 value recorded for S. cordata was 208 µg/mL, while for S. paniculata IC50 was 624 µg/mL. The study can serve as an important source of information to achieve the authenticity and to evaluate the quality and purity of the plant material in accordance to WHO guidelines. As this species is greatly exploited, so conservation is highly recommended.

Morpho-palynological assessment of medicinal flora of district Lahore, Pakistan based on LM and SEM.

This study was carried out to identify 20 high valued medicinal plants of District Lahore, Punjab, Pakistan with the help of the palynological studies. The distinguishing characteristics for selected species were found by systematized methods using light microscopy and Scanning Electron Microscopy. In this study, important pollen qualitative and quantitative features of these medicinal species such as polar and equatorial outline and their diameter, P/E ratio, presence or absence of pore, sculpturing of pollen, exine thickness, intine thickness, intercellular distance of exine and intine, presence or absence of spines, length of spines, width of spines, presence or absence of colpi and length of colpi, were compared. Remarkable variations in these pollen characters have been observed. Pollen grains were usually circular-semicircular (Calendula officinalis), spheroidal (Convolvulus arvensis), bilobed and trilobed (Convolvulus arvensis), spiny, (Abutilon indicum), oblate (Terminalia chebula) and elongated (Euphorbia pilulifera). In polar view Conyza bonariensis exhibited the highest pollen size [195.9 (211.6-180.2) µm]. While in equatorial view Calendula officinalis and Euphorbia pilulifera had lowest value [22.5 (20-25) µm] whereas Carissa carandas [165 (140-179) µm] appeared to be the largest. Parthenium hysterophorus showed the highest P/E ratio, that is, 1.28. Colpi were reported in only 7 species. Most of the plants showed 100% fertility but Terminalia chebula showed the least value 35%. In the present, project the diversity in pollen morphology present pollen as a valuable taxonomic tool. It is also concluded that pollen features are helpful at the specific level as well as generic level and can provide us fruitful taxonomic solutions. RESEARCH HIGHLIGHTS: Palynological flora of the medicinal plant species of district Lahore, Pakistan is investigated first time on the basis of LM and SEM and the flora is found very significant for taxonomic investigations.

Identification of Medicinally Used Flora Using Pollen Features Imaged in the Scanning Electron Microscopy in the Lower Margalla Hills Islamabad Pakistan.

The present study planned to assess the medicinally important plants in the lower Margalla Hills of Islamabad, Pakistan using scanning electron microscopy of their pollens. Both quantitative and qualitative characters of the pollens were analyzed. The shapes of the pollen were prolate, spheroidal, elliptical, and oblate. Exine sculpturing showed rugulate, scabrate, psilate gemmate, and reticulate ornamentation. The people residing in the rural areas are thought to depend, primarily, on medicinal plant resources for many health care needs. A sample of 140 respondents was selected and interviewed by using a stratified random sampling technique. Results showed that the local communities of lower Margalla Hills are highly dependent on plants to treat various ailments, for example, respiratory (28%), digestive (46%) nervous, circulatory and dermatological problems (6% each), and reproductive problems (8%). The continuous dependence of the local community on ethnomedicinally important plants has led to serious degradation of the natural forest of lower Margalla Hills. Priority-based conservation and sustainable use of these forest resources are necessary, and use of alternative resources will also help in the forest conservation.

Comparison of the Morphology, Anatomy, and Chemical Profile of Mikania glomerata and Mikania laevigata.

Mikania glomerata and Mikania laevigata are medicinal plants popularly used in Brazil for colds and asthma. Although several studies report differences in their composition, they are frequently used indistinctly, which may be due to misidentification of the species or the plant part used. Herein, the chemical, anatomical, and morphological profiles of both species were evaluated, seeking to differentiate them. Due to the pattern of secondary metabolites found in M. glomerata and M. laevigata, the chemical profile of the leaves was the characteristic that best differentiated the two species. Coumarin was present in all the leaf samples of M. laevigata but absent or in low concentration in leaf samples of M. glomerata. The stems presented coumarin in both species, albeit in different concentrations. The quantification of chlorogenic and dicaffeoylquinic acids was crucial for the separation of the two species, as higher concentrations are found in M. glomerata. The anatomic structure of the leaves, stem, and petioles of both species was conserved, even when exposed to different environments, but presented only subtle differences between species. The morphology of the leaves presented different characteristics for each species but altered when exposed to shade. Therefore, the chemical analysis together with the morphological characteristics of plants grown in full sunlight may be used for the correct identification of these species.

Organogenesis and Ultrastructural Features of In Vitro Grown Canna indica L.

An efficient protocol for micropropagation of Canna indica L., an economically and pharmaceutically important plant, was standardized using rhizome explants, excised from two-month-old aseptic seedlings. Complete plant regeneration was induced on MS medium supplemented with 3.0 mg/L BAP plus 1.5 mg/L NAA, which produced the highest number of shoots (73.3 ± 0.5%) and roots (86.7 ± 0.4%) after 2 weeks. Furthermore, the optimum media for multiple shoots regeneration were recorded on MS enriched with 7.0 mg/L BAP (33.0 ± 0.5%). Plantlets obtained were transplanted to pots after two months and acclimatized in the greenhouse, with 75% survival. In addition, ultrastructural studies showed that rhizomes of in vitro grown specimens were underdeveloped compared to the in vivo specimens, possibly due to the presence of wide spaces. Meanwhile, the leaves of in vivo specimens had more open stomata compared to in vitro specimens, yet their paracytic stomata structures were similar. Hence, there were no abnormalities or major differences between in vitro regenerants and mother plants.

Biofertilization with Azospirillum brasilense improves in vitro culture of Handroanthus ochraceus, a forestry, ornamental and medicinal plant.

Biofertilization with plant growth-promoting rhizobacteria is a potential alternative to plant productivity. Here, in vitro propagation of Handroanthus ochraceus (yellow lapacho), a forest crop with high economic and environmental value, was developed using the Azospirillum brasilense strains Cd and Az39 during rhizogenesis. Epicotiles of in vitro plantlets were multiplied in Woody Plant Medium (WPM). For rooting, elongated shoots were transferred to auxin-free Murashige-Skoog medium with Gamborg's vitamins and WPM, both at half salt concentration (½MSG and ½WPM), and inoculated with Cd or Az39 at the base of each shoot. Anatomical studies were performed using leaves cleared and stained with safranin for optical microscopy and leaves and roots metalized with gold-palladium for scanning electron microscopy (SEM). In ½WPM auxin-free medium, A. brasilense Cd inoculation produced 55% of rooting, increased root fresh and dry weight (45% and 77%, respectively), and led to lower stomata size and density with similar proportion of open and closed stomata. Both strains selectively increased the size or density of glandular trichomes in ½MSG. Moreover, bacteria were detected on the root surface by SEM. In conclusion, the difference in H. ochraceus response to A. brasilense inoculation depends on the strain and the plant culture media. Cd strain enhanced rooting in auxin-free ½WPM and produced plantlets with features similar to those expected in ex vitro plants. This work presents an innovative in vitro approach using beneficial plant-microorganism interaction as an ecologically compatible strategy in plant biotechnology.

The foliar anatomy and micromorphology of Aloe ferox Mill. (Asphodelaceae).

BACKGROUND: The genus Aloe has been the subject of several chemotaxonomic and phytochemical investigations. Aloe ferox is an important source of biologically active compounds like anthrones, chromones, pyrones, and flavonoids. It is a plant used extensively in traditional medicine. Despite the myriad of studies on the pharmacological profile of the plant, there still exists a dearth of scientific literature on the anatomical and morphological attributes of this important medicinal plant. MATERIALS AND METHODS: Hence, the objective of this study was to examine the foliar anatomy and micromorphological characteristics of A. ferox using light microscope and scanning electron microscope and to determine the elemental composition of the leaf sections by energy dispersive x-ray spectroscopy. RESULTS: Epidermal cells varied from round, square to hexagonal, with distinctive anticlinal walls and sunken stomata, equally distributed on both the adaxial and abaxial surfaces were observed. There were vertically elongated palisade cells with numerous chloroplasts in the palisade mesophyll layer. The EDXS analysis revealed Ca, K, S, Al, Mg, and Si as the major constituents of crystals present in the mesophyll. CONCLUSION: Information on the foliar anatomy and micromorphology can further enlightened our perceptions on the inherent interrelationships between structure and function as regards the synthesis and secretion of bioactive secondary metabolites by plants.

Ultrastructure and elemental analysis of Hypoxis hemerocallidea: a multipurpose medicinal plant.

BACKGROUND: Herbal medicine is a popular means of medical management in some parts of the world especially in Africa. Hypoxis hemerocallidea Fisch.C.A.Mey. & Avé-Lall, also known as African potato of the Hypoxidaecae family, is one of the medicinal plants that have enjoyed long usage as an herbal medicine in South Africa. In this study, the morphology and elemental constituents of H. hemerocallidea leaf was investigated to correlate the functional role of the ultrastructure in the production of therapeutic compounds. MATERIALS AND METHODS: Fresh leaves of H. hemerocallidea were prepared for analysis using standard methods. The ultrastructure and crystal deposits of the plant were assessed using scanning electron microscopy (SEM), and energy dispersive x-ray (EDX). RESULTS: It was observed that the leaves were characterised by multicelullar glandular and non glandular trichomes which are sparsely distributed over the entire surfaces. The glandular trichomes (GTs) in H. hemerocallidea leaf have boulbous heads which are probably filled with secretions, while the non glandular trichomes were long, fibrous and sparse. EDX-SEM of Hypoxis hemerocallidea leaf revealed that carbon, oxygen, nitrogen and silicon are the major components of the deposits, while other elements such as iron, sulphur, sodium, calcium, magnesium, potassium, manganese, iodine, chromium and iodine were present in small but variable amounts. CONCLUSION: The presence of these elements which are crucial to maintaining good health, in addition to other bioactive constituents might be accountable for the multipurpose therapeutic uses of Hypoxis hemerocallidea in the treatment of cancers, HIV/AIDS related diseases, urinary tract infections, cardiovascular disorders, diabetes and other chronic ailments of humans.

Foliar micro-morphology of Gasteria bicolor haw. (Asphodelaceae) from South Africa.

BACKGROUND: The succulent genus, Gasteria, which comprises 16 species, is endemic to South Africa and has its main centre of distribution in the Savanna Region of the Eastern Cape. Whereas G. bicolor has been investigated phyto-chemically and pharmacologically, not much data concerning the anatomical and micro-morphological features can be found in literature. MATERIALS AND METHODS: This study was undertaken, using light and scanning electron microscopy to obtain information on the micro-morphological features of this important medicinal plant to facilitate its identification and authentication. The elemental composition of the leaf was determined by energy dispersive X-ray spectroscopy (EDXS). RESULTS: The epidermal cells are either hexagonal or pentagonal in form, and are compactly arranged with undulate anti-clinal cell walls. The epidermal cell width was approximately 50 µm. Stomata apertures are elliptical and the upper epidermis of the leaf has paracytic stomata which are slightly raised above the epidermal surface with 4 to 5 subsidiary cells surrounding each stoma. Based on the EDXS microanalysis, the mineral crystals present at the level of the mesophyll of G. bicolor were probably mixtures of calcium oxalate, calcium sulphate and silica. CONCLUSION: The co-occurrence of aluminum suggests the potential role of the crystals in detoxification of aluminum and heavy metals, as reported previously.

Cytological evaluation of Apiaceae Lindl. from Western Himalayas.

The present paper deals with cytological studies on 31 populations covering 17 species belonging to 10 genera of Apiaceae from Western Himalayas. The chromosome numbers in the two species as Chaerophyllum capnoides (n = 11) and Heracleum brunonis (n = 11), along with additional cytotypes for Pimpinella acuminata (n = 9) and Sium latijugum (n = 12) have been reported for the first time on world-wide basis. The genus Pleurospermum, although cytologically worked out earlier from outside India, its species densiflorum (n = 11) makes first representation of the genus from India. Besides, the chromosome number in Chaerophyllum aromaticum (n = 11) have been worked out for the first time from India. The course of meiosis varies from normal to abnormal in different populations of Chaerophyllum villosum, Pimpinella achilleifolia and Sium latijugum while abnormal meiotic course has been observed in all the studied populations of Chaerophyllum acuminatum, C. aromaticum, C. capnoides, Pimpinella acuminata, P. diversifolia, Pleurospermum densiflorum and Vicatia coniifolia. Such taxa are marked with meiotic abnormalities in the form of cytomixis, chromatin stickiness, formation of laggards and bridges resulting into abnormal microsporogenesis. The occurrence of structural heterozygosity has been recorded in the Chaerophyllum acuminatum and C. aromaticum. The effect of these abnormalities is clearly seen on the pollen size and fertility.

Avaliação da qualidade de amostras de plantas medicinais comercializadas no Brasil / Quality assessment of samples of medicinal plants marketed in Brazil

Apart from constituting the raw material used to manufacture phytomedicines, plant drugs are commonly used by people as a therapeutic resource. Thus, the market in plant natural products has become an attractive target for investments of pharmaceutical companies. The aim of this study was to test the quality of commercial plant drugs in Brazil, employing simple and low-cost methods. Anatomical and microchemical tests were performed on commercial samples of "centela" (Asian pennywort or centella), "chá verde" (green tea) and "espinheira santa", to assess their quality and check their identity. The anatomical study revealed that all 3 samples of centella consisted of Centella asiatica leaves, but some were poorly conserved. The majority of contaminants consisted of other parts of C. asiatica, leaves of Poaceae and other species, and unidentified stalks. Two samples of green tea revealed leaves of the correct species (Camellia sinensis), with twigs of the same as contaminants, while the third consisted mainly of Ilex paraguariensis (mate tea) with some Bambusoideae (Poaceae) leaves. One of the 3 samples of "espinheira santa" contained Sorocea bonplandii leaves (cincho), and the others revealed leaves and stem fragments of Maytenus ilicifolia. The 3 samples of centella showed triterpene saponins. All samples of "green tea" revealed methylxanthines, but only those with C. sinensishad flavonoids. The samples of "espinheira santa" showed condensed tannins. Thus, the proposed analytical methods provided complementary results, which may be applied to quality control of plant drugs...

Drogas vegetais constituem uma das matérias-primas utilizadas na fabricação de fitoterápicos, além de serem largamente utilizadas pela população como recurso terapêutico. O mercado de produtos derivados de matéria-prima vegetal, com isso, se tornou alvo de investimentos de empresas do setor farmacêutico. O presente trabalho objetivou verificar a qualidade de drogas vegetais comercializadas no mercado brasileiro, utilizando-se conjuntamente métodos de análise simples e de baixo custo.Para tanto , foram usadas amostras de "centela", "chá verde" e "espinheira santa", obtidas em estabelecimentos comerciais, sendo sua identidade e qualidade avaliadas por meio de análises anatômicas e microquímicas. O estudo anatômico demonstrou que as três amostras de centela continham Centella asiatica, estando algumas em mau estado de conservação. Os contaminantes desta amostra eram principalmente outras partes do corpo vegetativo de C. asiatica, além de folhas de Poaceae e de outras espécies vegetais. Duas amostras de chá verde foram identificadas como Camellia sinensis e apresentavam caules da mesma espécie como contaminantes. A terceira amostra de chá verde era constituída por Ilex paraguariensis, sendo que folhas de Bambusoideae (Poaceae) também foram encontradas. Uma das amostras de espinheira santa era constituída de Sorocea bonplandii. As demais continham folhas e fragmentos de caule de Maytenus ilicifolia. As três amostras de centela apresentaram saponinas triterpênicas. Todas as amostras de chá verde possuíam metilxantinas. Dessas, apenas aquelas constituídas por C. sinensis demonstraram a presença de flavonoides. As amostras de espinheira-santa apresentaram taninos condensados. Desse modo, as metodologias propostas forneceram resultados complementares que podem ser empregados no controle de qualidade de drogas vegetais...

Microscopic characters of the leaf and stem of Lavandula dentata L. (Lamiaceae).

Lavandula dentata L. is an aromatic plant used in folk medicine for different purposes and, for this reason, phytochemical surveys have been carried out in the search for bioactive substances aiming to support its uses. Since there is little knowledge on the structural aspects of L. dentata, this work has studied the anatomical characters of the leaf and stem using light and scanning electron microscopy, in order to assist the species identification. As a result, there are different types of trichomes: capitate glandular with uni- or bicellular head, peltate glandular with multicellular head, and branched non-glandular. The leaf is hypostomatic showing diacytic stomata. The epidermis is uniseriate and coated with striate cuticle. The mesophyll is dorsiventral and the midrib is concave-convex and traversed by a single collateral vascular bundle. The stem is quadrangular and has alternating strands of collenchyma and cortical parenchyma as well as a typical endodermis in the cortex. The phloem and xylem cylinders are traversed by narrow rays and there is an incomplete sclerenchymatic sheath adjoining the phloem. These results are a novelty for the species and contribute to distinguish it from other lavenders.

Comparative authentication of three "snow lotus" herbs by macroscopic and microscopic features.

"Snow lotus" is a famous Chinese Materia Medica derived from species of the genus Saussurea (Compositae). To differentiate three representative easily-confused snow lotus herbs, namely, Saussurea involucrata (Kar. et Kir.) Sch.-Bip, Saussurea laniceps Hand.-Mazz., and Saussurea medusa Maxim., macroscopic features of the three herbs were systemically observed, and microscopic features were compared by using ordinary light microscopy, polarized light microscopy and scanning electron microscopy (SEM). The results indicate that, as for macroscopic identification, capitula situation and arrangement, and as for microscopic identification, pollen grains, nonglandular hairs, glandular hairs, and cells of inner surface of the microdiodange can be used to authenticate the three snow lotus herbs. Comprehensive table comparing the characteristics were presented in this study. SEM has been found to provide a number of unique characteristics of pollen grains. Based on the observation of pollen grains, evolution sequence of the three species was speculated. The present method was proven to be efficient, convenient, simple, and reliable, which was successfully applied to the authentication of three snow lotus herbs.

[Macroscopic and microscopic identification of Mongolian medicine Potentilla glabra].

OBJECTIVE: To provide the identification basis for Mongolian medicine Potentilla glabra. METHODS: Macroscopic and microscopic identification were applied to observe macroscopic, histological, superficial, and powder characteristics of its stems and leaves. RESULTS: The following characteristics were observed: The cork layer sandwiched with sclerenchyma ring which was main composed of fibers, accompanied by stone cells and large cell layer; Catheter with fiber existed in xylem; Beaded thickened anticlinal wall in leaf epidermal cell; Stomatal infinitive; Small clusters of calcium oxalate crystal and different type of leaf transverse section. CONCLUSION: These characteristics can provide evidences for the identification and quality control of Potentilla glabra.

[Macroscopic and microscopic identification of Ligularia przewalskii].

OBJECTIVE: To provide macroscopic and microscopic identification basis for Ligularia przewalskii. METHODS: Macroscopic and microscopic identification of roots, stems and leaves of Ligularia przewalskii were carried out with the methods of paraffin section, leaves epidermal section and powder transdermal section. RESULTS: The microscopic characteristics included: Open collateral vascular bundles in stem were not in the same size and arranged in two rings; Lots of fiber bundles scattered in the column parts; There were two vascular bundles in principal vein of leaf; Anticlinal wall of upper epidermis cells was thickened like moniliform, lower epidermis were like waves with irregular; The type of stoma was anomocytic; Calcium oxalate acicular crystal could be seen in the powder. CONCLUSION: These features can provide references for identification of Ligularia przewalskii.

[Macroscopic and microscopic identification of Zhuang medicine Tetrastigma planicaule].

OBJECTIVE: To study the macroscopic and microscopic characteristics of Zhuang medicine Tetrastigma planicaule. METHODS: Macroscopic and microscopic identification were studied. RESULTS: The microscopic characteristics were significant. The root had more than 10 layers of cork cells arranging in line. The starch grain and calcium oxalate cluster crystals distributed in the phloem parenchymas, and mucilage cells scattered in the cortex of the stem. There were 8 vascular bundles in the vein of leaves. CONCLUSION: This study provides a scientific basis for the identification and utilizing of Tetrastigma planicaule.

[Microstructure characteristics and identification significance of leaf epidermis of medicinal plants from Sabia genus].

OBJECTIVE: To study the feasibility and significance of the method to identify medicinal plants through the observation and statistics of 9 species of Sabia medical plants from Guizhou province. METHODS: Leaf epidermis characteristics were observed, measured by optical microscope and analyzed by SPSS 17.0 software. RESULTS: All of these plants had some differences in indumentum, cell morphology and size, and had significant difference in the length and circumference of lower epidermis cells. CONCLUSION: The method combining microscopic observation with statistics can be used as the classification and identification basis of medicinal plants and materials of Sabia genus.

Diversity of foliar endophytic fungi from the medicinal plant Sapindus saponaria L. and their localization by scanning electron microscopy.

Endophytic fungi inhabit vegetable tissues or organs, without causing them any harm. Endophytes can co-evolve with plant hosts and possess species-specific interactions. They can protect the plant from insect attacks and diseases, and are also able to produce substances of biotechnological interest. In folk medicine, the bark, roots and fruits of Sapindus saponaria is used to produce substances with anxiolytic, astringent, diuretic and expectorant properties, as well as tonics, blood depuratives and cough medicine. This study evaluated the diversity of endophytic fungi present in the leaves of S. saponaria L. and observed the colonization of host plants by endophytes, using light and scanning electron microscopy. We verified that these fungi are found in intercellular and intracellular spaces. The genera of some isolates of S. saponaria were identified mainly by sequencing of ITS region of rDNA and, when possible, also by their microscopic features, as follows: Cochliobolus, Alternaria, Curvularia, Phomopsis, Diaporthe and Phoma. Phylogenetic analysis showed the existence of genetic variability of the genera Phomopsis and Diaporthe and interspecific variation among the Curvularia, Alternaria and Phoma, belonging to family Pleosporaceae.

[Pollen morphological characteristics, viability test and storage of endangered medicinal plant Atractylodes lancea].

OBJECTIVE: To study the pollen morphological characteristics, viability test and storage character of the endangered plant Atractylodes lancea. METHOD: Pollen grains morphologies of A. lancea were observed by scanning electron microscope. The optimum culture medium and viability determination methods were screened out by liquid culture and dyeing methods, and then the pollen germination capacities in different storage conditions were detected. RESULT: The pollen grains are quasi-spherical, with tricolpate and spinous sculpture. The optimal culture medium was ME3 + 16% PEG4000 + 10% sucrose, in which the pollen germination capacity reached to 62.1%, while the other three dyeing methods were not able to be applied to detecting the pollen viability of A. lancea. The low storage temperature could significantly prolong the storage time of pollen of A. lancea. At -80 degrees C, pollen viability could be maintained for 60 days. CONCLUSION: Liquid culture method is suitable for the determination of pollen germination of A. lancea, and the rate of pollen germination is closely related to the storage time and temperature. At last, this study provides a foundation for the artificial pollination and cultivating in wildness of A. lancea.