Sumet Kongkiatpaiboon

Structural relationships of stemona alkaloids: assessment of species-specific accumulation trends for exploiting their biological activities.

On the basis of a comparison of 42 Stemona samples, representing eight different species collected and cultivated in Thailand, species-specific accumulation trends of Stemona alkaloids were analyzed. An overview was achieved by comparative HPLC analyses of methanolic crude extracts of underground parts coupled with diode array or evaporative light scattering detectors. All major compounds were isolated and their structures elucidated by NMR and MS analyses. Protostemonine- and stichoneurine-type derivatives dominated, from which the latter characterize S. tuberosa and S. phyllantha accumulating species-specific isomers of tuberostemonine (3). The widespread S. curtisii and S. collinsiae clearly deviate by protostemonine-type derivatives dominated by stemofoline (10) and/or didehydrostemofoline (11). Further diversification within this structural type results from a mutual accumulation of derivatives with a pyrrolo- or pyridoazepine nucleus, leading to chemical variability in S. curtisii and S. aphylla.

HPLC quantitative analysis of insecticidal didehydrostemofoline and stemofoline in Stemona collinsiae root extracts.

INTRODUCTION Stemona spp. have been traditionally used as natural pesticides and medicinal plants. Stemona collinsiae Craib has been of interest for its insecticidal activity, which has been supported by many scientific research studies. The roots contain didehydrostemofoline and stemofoline alkaloids as active components. OBJECTIVE To develop and validate a HPLC method for the quantitative analysis of didehydrostemofoline and stemofoline in S. collinsiae root extracts. METHODOLOGY HPLC was carried out using a Hypersil BDS C₁₈-column eluted with methanol:1 m m ammonium acetate (55:45) with a flow rate of 1 mL/min and detection at 295 nm. Method validation was performed to assure its linearity, precision, accuracy and limits of detection and quantitation. RESULTS Didehydrostemofoline and stemofoline showed a linear relationship within the range of 0.5-432.4 and 0.5-188.4 µg/mL, respectively. The method was shown to be precise with RSD < 2%. The average recovery of didehydrostemofoline and stemofoline were 98.80 and 99.97%, respectively. Eight samples of S. collinsiae root extracts were analysed and the average contents of didehydrostemofoline and stemofoline were 0.78 and 0.048% w/w, respectively. CONCLUSION The HPLC method developed was appropriate for the analysis of didehydrostemofoline and stemofoline in S. collinsiae root extracts. This work would be useful as a guide for the standardisation of S. collinsiae root extract raw materials and their products as natural pesticides.

Determination of Morin in Maclura cochinchinensis Heartwood by HPLC

The HPLC-DAD method was developed to determine morin content in Maclura cochinchinensis Corner heartwood extract. The chromatographic separation was performed using a Hypersil BDS C18 column, isocratic solvent system of 0.5% acetic acid in water:acetonitrile (80:20) with 1.0 mL/min flow rate and detected at 355 nm. The standard curve of morin was linear in the range of 7-905 μg/mL. The method was precise with intra-day relative standard deviation (RSD) of lower than 1% and 2.06% for inter-day RSD. The method accuracy represented by percent recover was 99.58%. The highly efficient HPLC system developed from this study could detect morin contents in M. cochinchinensis heartwood samples collected from various locations in Thailand in the range of 0.74-1.57% w/w. This developed method provided a useful standardization procedure of M. cochincihinesis materials for further application in pharmacy and other commercial developments.

Microscopic characterization as a tool for separation of Stemona groups.

Introduction: Stemona plants have been traditionally used as an insecticide, scabicide and pediculocide, and for the treatment of skin and respiratory diseases. Stemona can be separated into two groups according to their morphological characters and bioactive components i.e. stichoneurine and protostemonine groups. Protostemonine group contains alkaloids that possess potent insecticidal activity while stichoneurine group accumulates alkaloids with antitussive activity. In Thailand, a vernacular name “Non Tai Yak” refers to the roots of different species of Stemona, making it confusing to discern different species. The purposes of this study are to investigate the microscopic characteristics of the roots of seven species of Stemona growing in Thailand and to distinguish and identify these groups of Stemona. Methods: Cross-sectional histology of fresh root samples and powdered drug characteristics of 7 species of Stemona were studied under a microscope. Results: The roots of Stemona in the stichoneurine group (S. tuberosa and S. phyllantha) contained a non-lignified large pith while the roots of protostemonine group (S. burkillii, S. cochinchinensis, S. collinsiae, S. curtisii and S. kerrii) had a small lignified one. The powder of stichoneurine group contained numerous thin-walled parenchyma, but only few thick-walled parenchyma and lignified fibers and vessels were present. In contrast, thick-walled parenchyma and lignified fibers and vessels were frequently found in the powdered roots of protostemonine Stemona. These characteristics could be used to discern between Stemona in the stichoneurine and protostemonine groups. Conclusions: The microscopic characterizations can be used as a primary tool to categorize and separate 2 main Stemona groups.

Simultaneous quantification of stemocurtisine, stemocurtisinol and stemofoline in Stemona curtisii (Stemonaceae) by TLC-densitometric method.

Stemona curtisii Hook. F. (Family Stemonaceae), a prominent species distributed in the south and southwest of Thailand, has widely been used as a natural pesticide and as treatment for head lice and skin diseases. This study developed a thin-layer chromatography (TLC)-densitometric method for the simultaneous quantification of major components--stemocurtisine, stemocurtisinol and stemofoline--in the extracts from the roots of S. curtisii collected from 10 locations in Thailand. Components were found in the ranges of 0.0353-0.1949, <0.0121-0.0859 and 0.0733-0.1689 percent dry weight, respectively. The method was validated for linearity, precision, accuracy, robustness, limit of detection and limit of quantitation. The linearity was found over the range of 40-320 ng/spot with a good correlation coefficient (r > 0.9866). Intra-day and inter-day precision showed a relative standard deviation of less than 6%. The accuracy of the method was determined by a recovery study, and the average recoveries were 100.4, 100.2 and 100.3% for stemocurtisine, stemocurtisinol and stemofoline, respectively. The proposed TLC-densitometric method was found to be simple, precise, specific and inexpensive, and can be used simultaneously for the routine quality control of raw materials of S. curtisii roots, extracts and their products, and also other products containing these markers.

Conserving threatened widespread species: a case study using a traditional medicinal plant in Asia

Overharvesting and loss of habitat arising from human activities are the most important threats to plants and animals. Conservation efforts targeting medicinal plant species usually focus on endemic, economic and endangered taxa, typically Plant Species with Extremely Small Populations (PSESP) under high extinction risk. However, comparatively little attention has been focused on traditional medicinal plants with widespread distribution, although some of these species, where the whole plant or the tubers are raw materials for medicines, may be on the edge of local extinction. In this study, we appeal to conservationists to pay close attention to threatened widespread species. To this end, a traditional medicinal plant, Stemona tuberosa, which has a distribution that covers more than ten countries in Asia, is studied in detail. The results suggest that destructive overharvesting of the underground tuberous roots of S. tuberosa is the key threat to this species. In addition, pollination limitation, loss of potential seed dispersers, habitat loss, and a scattered distribution pattern may further reduce the survival chances of S. tuberosa. Through multidisciplinary conservation efforts conducted by researchers and local resident, we witnessed the preliminary recovery of S. tuberosa in locations where it has been reintroduced. We think that threatened widespread species should not be overlooked in the field of conservation. Local participation in natural resource management may help to make resources more relevant locally and hence sustainable in remote regions. We suggest that developing multi-stakeholder coalitions should be encouraged to save threatened medicinal plant species in underdeveloped areas.

Antioxidant Activity and Antibacterial Effects on Clinical Isolated Streptococcus suis and Staphylococcus intermedius of Extracts from Several Parts of Cladogynos orientalis and Their Phytochemical Screenings

The in vitro antioxidant and antibacterial assays against clinically isolated Streptococcus suis and Staphylococcus intermedius of the extracts prepared by decoction and ethanolic reflux of different parts of Chettaphangki (Cladogynos orientalis Zipp. ex Span), including the leaves, roots, and stems, using 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging assay and disc diffusion method were conducted. Quantitative analysis of total phenolic and total flavonoid contents in the extracts using spectrophotometric methods was also performed. Finally, phytochemical screening by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) was conducted. Leaf ethanolic reflux extract (100 g) contained the highest total phenolic and total flavonoid contents of 7.21 ± 0.28 μg gallic acid equivalent (GAE) and 11.51 ± 2.02 μg rutin equivalent (RE), respectively. Chettaphangki extracts promoted low antioxidant activity with EC50 values in the range of 0.27–0.48 mg/mL. Extracts and fractions from the roots and stems of this plant promoted low to intermediate antibacterial activity against S. intermedius with the inhibition zones between 7 and 14 mm. The chromatographic data suggested that the leaf extracts of C. orientalis contained rutin while the root and stem extracts contained scopoletin and chettaphanin I. Rutin promoted strong antioxidant activity while chettaphanin I showed low antibacterial activity against Staphylococcus intermedius.