LED Lights Affecting Morphogenesis and Isosteroidal Alkaloid Contents in Fritillaria cirrhosa D. Don-An Important Chinese Medicinal Herb.

Investigations were carried out to study the effects of light-emitting diode (LED) lights on growth and development of isosteroidal alkaloids in embryogenic calli of Fritillaria cirrhosa D. Don, an important traditional Chinese medicine herb. Calli were cultured in glass bottles, each containing 100 mL of Murashige and Skoog's basal medium supplemented with 2% sucrose and 0.4% gellan gum powder, a gelling agent. These bottles were incubated in a specially designed plant growth chamber equipped with eight different LED lights consisting of single or combinations of four different light spectra emitting blue (450 nm), green (525 nm), red (660 nm), and far-red (730 nm) light. After three months of incubation, morphological changes in embryogenic calli were recorded, and LC-MS/MS analysis of cultures was carried out for peimisine, sipeimine, peiminine, and peimine. The highest number of somatic embryos and the maximum fresh weight was recorded in calli incubated under red (9R), infrared (9IR), and a combination of red+blue+infrared (3R3B3IR), respectively, in decreasing order. The highest contents of peimisine, peiminine, and peimine were recorded under red (9R) and infrared (9IR) lights, respectively. Eight LED lights had significant effects on the morphogenesis of embryogenic calli of F. cirrhosa D. Don and contents of isosteroidal alkaloids.

In vitro propagation of bulblets and LC-MS/MS analysis of isosteroidal alkaloids in tissue culture derived materials of Chinese medicinal herb Fritillaria cirrhosa D. Don.

BACKGROUND: Fritillaria cirrhosa, an important Chinese medicinal herb, is a Class-III protected and highly exploited species by pharmaceutical industry. Dwindling wild populations of species are unable to meet market demand. Therefore, this study was carried out to develop an in vitro propagation method for bulblet production. Also, the study aimed to carry out LC-MS/MS analysis of tissue culture-derived bulblets and callus for the presence of isosteroidal alkaloids (peimissine, verticine, and verticinone), and compare its quantities with commercially available crude drug samples. RESULTS: In vitro seed germination (91%) of F. cirrhosa was achieved on Murashige and Skoog's basal medium (MSBM) supplemented with 6-benzylaminopurine (1 mg L-1) and α-naphthalene-acetic-acid (0.4 mg L-1). On transfer of germinated seeds from Petri-dishes to glass bottles containing hormone-free MSBM, 37.5% of seedlings developed bulblets after 3 months of incubation. Regeneration and multiplication of bulblets were achieved by culture of transverse sections of bulblets on 1/2 X MSBM. By repeated subcultures at an interval of 2 months, 3072 bulblets weighing 1270 g could be produced at the end of 5th subculture. LC-MS/MS analysis showed a significant presence of peimissine in in vitro bulblets while callus incubated in the dark showed presence of peimissine and verticine. CONCLUSION: The study reports an efficient in vitro propagation method of bulblets production of F. cirrhosa and presence of some isosteroidal alkaloids in tissue culture-derived bulblets and callus. The study could be of immense help in production of F. cirrhosa bulblets and callus under laboratory conditions round the year. Also, these results can be used further to investigate production of isosteroidal alkaloids in bioreactors at commercial scale using liquid and cell suspension cultures. Thus, we not only can reduce our dependence on collections from natural habitats, but also can help in in situ conservation of this important species.

Enhanced production of podophyllotoxin, kaempferol, and quercetin from callus culture of Dysosma pleiantha (Hance) Woodson: An endangered medicinal plant.

Dysosma pleiantha (Hance) Woodson is one of the endangered traditional Chinese medicinal herbs, highly valued for its medicinal properties by Taiwan's mountain tribes. The present study aims to develop an efficient protocol for callus biomass by optimizing suitable culture medium, carbon source culture condition, and enhanced production of pharmaceutically important podophyllotoxin, kaempferol, and quercetin from callus culture of D. pleiantha under the influence of different additives. Best callus induction was achieved in Gamborg's medium (B5) with 1 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) along with 0.2 mg/L kinetin under dark condition. Tender leaves of D. pleiantha showed the maximum of 86% callus induction among the different explants tested. Highest leaf callus proliferation was noted in B5 medium with 1 mg/L 2,4-D incubated under complete darkness. In addition, it was found that B5 medium with 1 mg/L 2,4-D along with 2 g/L peptone produced more leaf callus biomass and enhanced production of podophyllotoxin (16.3-fold), kaempferol (12.39-fold), and quercetin (5.03-fold) compared to control. Therefore, D. pleiantha callogenesis can provide an alternative source for enhanced production of secondary compounds regardless of the exploitation of its natural plant population.

Biogenic synthesis of silver nanoparticles using rhizome extract of Dysosma pleiantha and its antiproliferative effect against breast and human gastric cancer cells.

Synthesis of biogenic metal nanoparticles using plant extract has gained considerable attention in recent years. The present study aims to synthesize and investigate the cytotoxic effect of silver nanoparticles (AgNPs) from Dysosma pleiantha rhizome extract. The green biosynthesis of AgNPs was verified by ultraviolet visible spectrometer, and characterized using fourier transform infrared spectroscopy, transmission electron microscopy and scanning electron microscopy. Results of microscopic studies revealed that the synthesized AgNPs were a spherical shape with an average size of 76 nm. We also examined the anti-cancer activity of biologically synthesized AgNPs. The dose-dependent cytotoxicity was observed in the breast cancer cell lines MDA-MB-231 and MDA-MB-453 treated with biogenically synthesized AgNPs, and the IC50 was recorded at 33.521  and 36.25 µM respectively. The DNA fragmentation analysis showed that the MDA-MB-231 cells treated with increasing concentrations of AgNPs significantly triggered the fragmentation of DNA. In addition, the synthesized AgNPs exhibited dose dependent cytotoxic potential against human gastric cancer cell lines and the IC50 was recorded at 7.14 µM. Thus, the green biosynthesized AgNPs from D. pleiantha rhizome can be used in the novel development of anticancer drugs.

Andrographolide relieved pathological pain generated by spared nerve injury model in mice.

CONTEXT: Andrographolide (Andro), found in large quantities in Andrographis paniculata Nees (Acanthaceae), is anti-inflammatory, especially in the central nervous system (CNS) glia. OBJECTIVE: The objective of this study is to test Andro's ability to reduce allodynia in a spared nerve injury model. MATERIAL AND METHODS: Male 30 g BalbC mice were divided into four groups: (1) Sham-operated control (Sham-group); (2) nerve injured and treated with saline (Saline-group); (3) nerve injured and treated with Andro (Andro-group); (4) nerve injured and treated with non-steroidal anti-inflammatory drugs (NSAIDS) (NSAIDS-group). Andro or NSAIDS (diclofenac salt) were injected intraperitoneally at 5 mg/kg body weight daily. Mechanical allodynia was assessed by von Frey tests at 3, 7, and 14 d. For immunohistochemical analysis, samples were collected at 7 d. RESULTS: The threshold for inducing allodynia increased and the response percentage reduced in the Andro-group when compared with the Saline-group, as well as when compared with NSAIDS groups throughout 3-14 d. The ratio of threshold for OP-Andro/OP-saline and for OP-Andro/OP-NSAIDS groups was 20.42 and 11.67 at 14 d, respectively. The ratio of response percentage for OP-Andro/OP-saline and for OP-Andro/OP-NSAIDS was 0.32 and 0.39 at 14 d, respectively. Interleukin-1 (IL-1) immunostaining in the spinal cord was reduced in the Andro-group. Astrocytic activities were not significantly reduced in the Andro-group compared with the Saline-group at 7 d post-operation (PO) Conclusions: Andro reduced mechanical allodynia more than NSAIDS at the same concentration, and the observed behaviour was associated with a reduction in inflammatory cytokine produced in the spinal cord.

Ethyl acetate fraction from methanol extraction of Vitis thunbergii var. taiwaniana induced G0 /G1 phase arrest via inhibition of cyclins D and E and induction of apoptosis through caspase-dependent and -independent pathways in human prostate carcinoma DU145 cells.

Vitis thunbergii var. taiwaniana (VTT) is a wild grape native to Taiwan, belonging to the Vitaceae family and Vitis genus, and widely used as folk herbal medicine. It is traditionally used for the treatment of diarrhea, hypertension, neuroprotection, jaundice, and arthritis. We used the wild-collected VTT and sterilized them to establish the plant tissue culture, and then took the leaves for DNA sequencing to determine its original base. We use methanol to extract VTT in four different solvents: 1-butanol, n-hexane, ethyl acetate, and water. These four preliminary extracts were used to treat human prostate cancer DU145 cells in vitro. We use the flow cytometry to check the cell survival situation. Finally, we found the ethyl acetate layer roughing product (referred VTEA) in human prostate cancer apoptotic effects of cell line DU-145. In the present studies, we use the crude extract of VTT to examine whether or not it can induce apoptosis of DU145 cells in vitro. Viability assays for extracts of VTT treatment showed that it had dose-dependent effect on human prostate cancer DU145 cells. We also found that the extract of VTT induces time-dependent mitochondrial and intrinsic-dependent apoptosis pathways. The in vitro cytotoxic effects were investigated by cell cycle analysis and the determination of apoptotic DNA fragmentation in DU145 cells. The cell cycle analysis showed that extracts of VTT induced a significant increase in the number of cells in G0 /G1 phase. The extract of VTT induced chromatin changes and apoptosis of DU145 cells also were confirmed by DAPI and PI staining that were measured by fluorescence microscopy and flow cytometry, respectively. Finally, the expression of relevant proteins was analyzed by Western blot analysis. These results promoted us to further evaluate apoptosis associated proteins and elucidate the possible signal pathway in DU-145 cells after treated with the extract of VTT.

Biocompatible 3D SERS substrate for trace detection of amino acids and melamine.

A novel, low-cost and biocompatible three-dimensional (3D) substrate for surface-enhanced Raman spectroscopy (SERS) is fabricated using gold nanoparticles (AuNPs) loaded on cellulose paper for detection of amino acids and melamine. Dysosma pleiantha rhizome (Dp-Rhi) capped AuNPs (Dp-Rhi_AuNPs) were prepared by in situ using aqueous extract of Dp-Rhi and in situ functionalized Dp-Rhi on AuNPs surface was verified by Fourier transform infrared spectroscopy and zeta potentials analysis shows a negative (-18.4mV) surface charges, which confirm that presence of Dp-Rhi on AuNPs. The biocompatibility of Dp-Rhi_AuNPs is also examined by cell viability of FaDu cells using MTS assay and compared to control group. In conclusion, the SERS performance of AuNPs@cellulose paper substrates were systematically demonstrated and examined with different excitation wavelengths (i.e. 532, 632.8 and 785nm lasers) and the as-prepared 3D substrates provided an enhancement factor approaching 7 orders of magnitude compared with conventional Raman intensity using para-nitrothiophenol (p-NTP), para-aminothiophenol (p-ATP) and para-mercaptobenzoic acid (p-MBA) as probe molecules. The strong electromagnetic effect was generated at the interface of AuNPs and pre-treated roughened cellulose paper is also investigated by simulation in which the formation of possible Raman hot-spot zone in fiber-like microstructure of cellulose paper decorated with AuNPs. Notably, with optimized condition of as-prepared 3D AuNPs@cellulose paper is highly sensitive in the SERS detection of aqueous tyrosine (10-10M) and melamine (10-9M).

Influence of LED light spectra on in vitro somatic embryogenesis and LC-MS analysis of chlorogenic acid and rutin in Peucedanum japonicum Thunb.: a medicinal herb.

BACKGROUND: Peucedanum japonicum Thunb, an important medicinal herb is reported to possess pharmacological properties such as anti-obesity, anti-oxidant, anti-inflammatory, anti-bacterial, anti-diabetic and anti-platelet aggregation. The present study aimed to develop an in vitro plant regeneration system of P. japonicum via somatic embryogenesis and to analyse chlorogenic acid and rutin contents in a few commercially available plant products of P. japonicum in Japan and Taiwan markets, and tissue culture plants derived from somatic embryos. RESULTS: Induction of somatic embryogenesis could be achieved when root derived calli after three subcultures were transferred from Murashige Skoog's salts and vitamins (MS basal) medium with 2,4-dichlorophenoxyacetic acid (2,4-D) (0.1-5 mg/L) to a medium with abscisic acid (ABA) (0.5-4 mg/L), or exposed to eight different light spectra provided by light-emitting diode (LED) sources. Concentrations of ABA and LED light spectra had an influence on number of somatic embryos induced and proliferation of callus. Development of secondary somatic embryos and conversion of embryos to plantlets was achieved on a medium with ABA, or their exposure to red or blue lights in a special incubation chamber. Four months old tissue culture plants derived from somatic embryos showed significantly higher levels of chlorogenic acid (10.5 mg/g dw) compared to commercial product sold in Japanese market (0.55 mg/g dw). However, rutin was absent in tissue culture plants in contrast to commercial sample (0.33 mg/g dw). CONCLUSION: In this report, we describe in vitro plant regeneration system in P. japonicum via somatic embryogenesis and production of chlorogenic acid in tissue culture plants. The present study has application in further tissue culture propagation of elite plant material with high chlorogenic acid content, and identification of high yielding plants with the LC-MS method.

Therapeutic values, chemical constituents and toxicity of Taiwanese Dysosma pleiantha--a review.

Dysosma pleiantha (Hance) Woodson also called as Bajiaolian belongs to the family Berberidaceae, is widely used in Taiwan as traditional Chinese herbal medicine for more than thousands of years. It is usually recommended by various traditional Chinese medical doctors and herbal pharmacies for general remedies including postpartum recovery, treatment of weakness, neck mass, acne, hepatoma, lumbago, snakebite, tumor growth and dysmenorrhea. In the textbooks of traditional Chinese medicine, there is limited information about the toxicity of Bajiaolian. Podophyllotoxin, a lignan is the main toxic ingredient of Bajiaolian rhizome. Therefore, Bajiaolian is documented as the fifth highest cause of poisoning among the herbal medicine in Taiwan. Since the therapeutic and toxic doses are very close, Bajiaolian poisoning cases are frequently reported in Taiwan. Moreover, Dysosma poisoning cases are difficult to diagnosis because physicians are unfamiliar with this medicine's multiple clinical presentations in different stages of intoxication. Therefore, the objective of this review is to represent the collective information available in literatures regarding D. pleiantha, a cytotoxic lignan containing medicinal plant. Specifically, the literatures have been reviewed for articles pertaining to chemical constituents, properties, therapeutical benefits, toxicity, poisoning symptoms, toxic as well as therapeutic dose and medical management.

In vitro culture and production of syringin and rutin in Saussurea involucrata (Kar. et Kir.) - an endangered medicinal plant.

BACKGROUND: Saussurea involucrata (Kar. et Kir.) commonly known as 'snow lotus' or 'Xue Lian' is an important plant in the traditional Chinese system of medicine. The plant contains flavonoids such as syringin and rutin. These compounds have been reported to be anti-rheumatic, anti-inflammatory and dilate blood vessels, lower blood pressure, prevent cardiovascular diseases, enhance immunity, and act as anti-aging, anti-cancer, and anti-fatigue agents. The species has become endangered due to the excessive collection of S. involucrata plants in the wild, slower plant growth and ecological destruction of natural habitats. There is a severe shortage of plant material, while the market demand is ever increasing. Hence, it is very important to apply tissue culture technique for plant propagation and production of the bioactive compounds of this species. RESULTS: Multiple shoot induction and proliferation in shoot base explants derived from in vitro raised seedlings of S. involucrata was achieved on 3/4 strength of Murashige and Skoog's (MS) basal medium (MSBM) supplemented with 1.0 mg/L-1 BA and 1.5 mg/L-1 NAA. Rooting was induced in 100 % shoots cultured on 1/2X MSBM supplemented with 1.0 mg/L-1 IBA for one week and then transfer to auxin free medium. The plantlets could be acclimatized successfully by sachet technique and established in the greenhouse. Maximum callus induction and proliferation in leaf segments was achieved on 1/2X MSBM supplemented with 0.5 mg/L-1 BA, 0.5 mg/L-1 NAA, 0.4 % gelrite and on incubation at 20 °C. Container closures had an influence on the quality and quantity of callus and production of the active compounds. The HPLC analysis showed much higher syringin content in in vitro shoots and callus as compared to commercially available market crude drug. CONCLUSION: The present study describes an in vitro culture protocol of Saussurea involucrata. The bioactive compounds, syringin and rutin could be produced through tissue culture technique without sacrificing the endangered Saussurea involucrata plants in the wild.

The suppressive activities of six sources of medicinal ferns known as gusuibu on heat-labile enterotoxin-induced diarrhea.

Diarrheal disease is one of the most important worldwide health problems. Enterotoxigenic Escherichia coli (ETEC) is the most frequently isolated enteropathogen in diarrheal diseases. In developing countries, a very large number of people, especially children, suffer from diarrhea. To combat this problem, World Health Organization has constituted the Diarrhea Diseases Control Program which guides studies on traditional medicinal practices and preventive measures. Gusuibu, a traditional folk medicine, has been claimed to heal certain types of diarrhea. However, so far no scientific study has been carried out on the anti-diarrheal mechanism of Gusiubu. The present study was performed to examine the suppressive activities of ethanol extracts of six sources of folk medicinal ferns used as Gusuibu on heat-labile enterotoxin (LT)-induced diarrhea. Inhibitory effects of six sources were evaluated on the ETEC LT subunit B (LTB) and monosialotetrahexosylganglioside (GMI) interaction by GM1-enzyme linked immunosorbent assay and patent mouse gut assay. Our results indicated that Drynaria fortunei had no anti-diarrheal effect, while, among the remaining five folk medicinal ferns, four belonging to family Davalliaceae had significant abilities on both the blocking of LTB and GM1 interaction and the inhibition of LT-induced diarrhea. In conclusion, these findings suggested the potential application of Gusuibu as an anti-diarrheal remedy.

Inhibitory effects of cultured Dendrobium tosaense on atopic dermatitis murine model.

Dendrobium tosaense is one of the most valuable Chinese medicines and well developed health food. Atopic dermatitis (AD) is a chronic skin disease that occurs mainly in childhood. The pathogenesis of atopic dermatitis had been studied in BALB/c mice modeling by skin-inoculated ovalbumin (OVA) with 2,4,6-trinitro-1-chrolobenzene (TNCB). These mice exhibit features of chronic dermatitis, including skin rash, mast cells infiltration, and elevated serum anti-OVA specific IgE and cytokines modulation. In this study, a standardized ethyl acetate extract of D. tosaense (DtE) was used to protect these mice from the OVA/TNCB-induced skin lesions of atopic dermatitis. The results indicated an increased population of natural T regulatory cell was accompanied by immunosuppression in cytokine profiles and anti-OVA IgE level to significantly reduce Th2 polarization. Finally, toluidine blue staining indicated mast cell infiltration and degranulation was reduced in skin lesion. Our results were shed light on the usage of D. tosaense in AD.

Establishment of hairy root lines and analysis of iridoids and secoiridoids in the medicinal plant Gentiana scabra.

BACKGROUND: Gentiana scabra is commonly known as 'Longdan' is an important herb in traditional Chinese medicines, commonly used for the treatment of inflammation, anorexia, indigestion and gastric infections. Iridoids and secoiridoids are main bioactive compounds which attributed to the pharmacological properties of this plant. The use of hairy root cultures as an excellent alternative for the production of pharmaceutically important metabolites in less time period with ensured quality of raw materials. RESULTS: An efficient hairy root culture system of Gentiana scabra and influence of different plant growth regulators (PGRs) on the production of gentiopicroside, swertiamarin and loganic acid constituents were described. Leaf explants were infected with Agrobacterium rhizogenes, which induced hairy roots up to 21%. The transformed hairy root lines were confirmed by PCR using rolB and rolC gene-specific primers. Among various solid and liquid media, B5 liquid medium resulted maximum root biomass (36- fold higher) in 4-weeks. Quantitative analysis showed loganic acid was 6.6- fold higher in the presence of zeatin (1 mg/l) and gentiopicroside accumulation was 1.8- fold higher in the presence of naphthaleneacetic acid (NAA, 1 mg/l), as compared to the roots of plants grown in greenhouse. On the other hand, 1.4- and 2.5- fold higher gentiopicroside and swertiamarin were observed in the presence of 1.0 mg/l NAA as compared to commercial Gentiana herb No. 2. The result also showed iridoid and secoiridoid contents affected greatly by age, physiology and growing environment of the plant. CONCLUSIONS: The use of hairy root cultures is an excellent alternative to harvesting natural or in vitro grown plants to produce pharmaceutically important metabolites in less time with ensured quality.

In vitro propagation and analysis of secondary metabolites in Glossogyne tenuifolia (Hsiang-Ju) - a medicinal plant native to Taiwan.

BACKGROUND: Glossogyne tenuifolia Cassini (Hsiang-Ju in Chinese) is a perennial herb native to Penghu Islands, Taiwan. The herb is a traditional anti-pyretic and hepatoprotective used in Chinese medicine. Several studies on G. tenuifolia have demonstrated its pharmacological values of antioxidation, anti-inflammation, immunomodulation, and cytotoxicity on several human cancer cell lines. Active compounds, oleanolic acid and luteolin in G. tenuifolia are affected by several factors, including climatic change, pathogens and agricultural practices. Plant population of G. tenuifolia has been severely affected and reduced considerably in natural habitat due to the use of herbicides by farmers. Also, collection of plant material from the natural habitat is restricted to a few months in a year. Therefore, the objective of the present study was to develop an efficient micropropagation protocol for G. tenuifolia. The study also aimed to investigate the influence of in vitro growth environment on the active compounds in in vitro shoots, tissue culture raised greenhouse plants; compare the values with wild plants and commercially available crude drug. RESULTS: Half-strength MS (Murashige and Skoog) basal medium supplemented with 0.1 mg/L 6-benzyladenine (BA) and 0.1 mg/L α-naphthaleneacetic acid (NAA) induced the maximum average number of shoots (7.3) per shoot tip explant excised from in vitro grown seedlings. Induction of rooting in cent percent in vitro shoots with an average number of 6.6 roots/shoot was achieved on ½ strength MS medium supplemented with 3.0 mg/L indole-3-acetic acid (IAA). The rooted plantlets acclimatized successfully in the greenhouse with a 100% survival rate. HPLC analysis revealed that the quantity of oleanolic acid and luteolin in in vitro shoots, tissue culture plants in the greenhouse, wild type plants and commercial crude drug varied depending upon the source. The oleanolic acid and luteolin contents were found to be significantly higher (16.89 mg/g and 0.84 mg/g, respectively) in 3-month old tissue culture raised plants in greenhouse compared to commercially available crude drug (6.51 mg/g, 0.13 mg/g, respectively). CONCLUSIONS: We have successfully developed an in vitro propagation protocol for G. tenuifolia which can expedite its plant production throughout the year. The contents of oleanolic acid and luteolin in the tissue culture raised plants in the greenhouse were significantly higher than the marketed crude drug demonstrating the practical application of the tissue culture technology. These findings may be very useful in micropropagation, germplasm conservation and commercial cultivation of G. tenuifolia. So far, there is no published report on tissue culture propagation of this important medicinal plant species.

In vitro propagation of Gentiana scabra Bunge - an important medicinal plant in the Chinese system of medicines.

BACKGROUND: Gentiana scabra Bunge commonly known as 'Long dan cao' in China has been used in traditional Chinese medicines for more than 2000 years. Dry roots and rhizome of the herb have been used for the treatment of inflammation, anorexia, indigestion and gastric infections. Iridoids and secoiridoids are the main bioactive compounds which attribute to the pharmacological properties of this plant. The species is difficult to mass propagate by seed due to the low percentage of germination and limited dormancy period. Wild populations in some locations are considered to be in the endangered category due to over exploitation. RESULTS: In the present study, we report an efficient micropropagation system. Shoot apices of six weeks old in vitro grown G. scabra plants were used as explants for the in vitro propagation. Induction of multiple shoots (9.1/explant) was achieved on the culture of shoot apices on half strength Murashige and Skoog's basal medium (MSBM) containing 2.0 mg/L-1 6-benzylaminopurine (BA), 3% sucrose and 0.9% Difco agar. In vitro shoots induced profuse rooting on half strength of MSBM supplemented with 0.1 mg/L-1 1-naphthaleneacetic acid (NAA), 3% sucrose and 0.3% gelrite. A two-stage ventilation closure procedure during the in vitro culture, and transparent sachet technique enhanced the survival rate of G. scabra plantlets to 96% in the greenhouse. Tissue culture plants flowered after 5 months of transfer to pots. CONCLUSIONS: A simple and an efficient in vitro propagation protocol of Gentiana scabra Bunge by optimizing the medium composition and ventilation closure treatments has been developed. The protocol can be very useful in germplasm conservation and commercial cultivation of G. scabra plants.

Anti-metastatic activity of biologically synthesized gold nanoparticles on human fibrosarcoma cell line HT-1080.

Plants are exploited as a potential source for the large-scale production of noble gold nanoparticles in the recent years owing to their various potential applications in nanobiotechnology and nanomedicine. The present work describes green biosynthetic procedures for the production of gold nanoparticles for the first time by using an aqueous extract of the Dysosma pleiantha rhizome. The biosynthesized gold nanoparticles were confirmed and characterized by ultraviolet-visible spectroscopy, Fourier transform infrared spectroscopy, transmission electron microscopy, and scanning electron microscopy equipped with energy dispersive spectroscopy. The results revealed that aqueous extract of D. pleiantha rhizome has potential to reduce chloroauric ions into gold nanoparticles and the synthesized gold nanoparticles were showed spherical in shape with an average of 127nm. Further, we investigated the anti-metastatic activity of biosynthesized gold nanoparticles against human fibrosarcoma cancer cell line HT-1080. The results showed that the biosynthesized gold nanoparticles were non-toxic to cell proliferation and, also it can inhibit the chemo-attractant cell migration of human fibrosarcoma cancer cell line HT-1080 by interfering the actin polymerization pathway. Thus, the usage of gold nanoparticles biosynthesized from D. pleiantha rhizome can be used as a potential candidate in the drug and gene delivery to metastatic cancer.

Activation tagging in Salvia miltiorrhiza can cause increased leaf size and accumulation of tanshinone I and IIA in its roots.

BACKGROUND: Salvia miltiorrhiza Bunge (Danshen), an important herb in traditional Chinese medicine, is commonly used for treatment of cardiovascular diseases. One of the major bioactive constituents of Danshen, diterpenoid tanshinone, has been proved with pharmacological properties and have the potential to be a new drug candidate against various diseases. In our previous study, we have established an activation tagging mutagenesis (ATM) population of callus lines of S. miltiorrhiza Bunge by Agrobacterium- mediated transformation. RESULTS: In the present study, we have identified ATM transgenic Salvia plant (SH41) with different leaf morphology and more tanshinones in its roots. The transgenic background of SH41 was identified by PCR (using hpt II primers) and Southern blots. PCR analysis showed a single band of hpt II gene and Southern blot analysis showed single insertion in SH41. External appearance of ATM transgenic SH41 was observed with broader leaves comparing to non-transformed plants. More healthy trichomes as well as bigger and wobbly guard cells and stomata were observed in SH41 by scanning electron microscopy (SEM). Quantitative analysis of active compounds in SH41 roots revealed a significant increase in tanshinone I (3.7 fold) and tanshinone IIA (2 fold) contents as compared to the wild plant. CONCLUSIONS: We have generated an activation tagged transgenic Salvia plant (SH41) with different leaf morphology and high diterpenes content in its roots. The increased amount of tanshinones in SH41 will definitely offer a route for maximizing the benefits of this plant in traditional Chinese herbal medicines. The present report may also facilitate the application of ATM for genetic manipulation of other medicinal crops and subsequent improved metabolite contents.

Comparative analysis among three Taiwan-specific Gentiana species and Chinese medicinal plant Gentiana scabra.

BACKGROUND: The root of Gentiana scabra is commonly known as Longdan in Chinese herbal medicines and has been used in the treatment of inflammation, anorexia, indigestion and gastric infections for over 2000 years. High market demand had made G. scabra (GS) plants not to be the only source of Longdan in China, other Gentiana spp., G. triflora, G. manshurica and G. rigescens, were also recognized as Longdan in China now. RESULTS: In this study, we identified three Taiwan-specific Gentiana spp., G. davidii var. formosana (GDF) and G. arisanensis (GA) and G. scabrida var. punctulata (GSP) that are phylogenetically different from GS (main source of Longdan). However, the active compounds of Longdan, gentiopicroside and swertiamari, were found in GSP and GDF showed higher antioxidant ability and free radical scavenging activities than Chinese Longdan. This discovery might explore the medicinal potential of GDF. Meanwhile, another Taiwan-specific Gentiana spp., GSP, was found to have the strongest antioxidant ability and free radical scavenging activities which might suggest a possible use of GSP as a source of natural antioxidant agents for industrial purpose. CONCLUSIONS: The finding of this study indicated that ITS analysis can be used to identify Taiwan-specific Gentiana spp. Also the Taiwan-specific Gentiana spp. which has strongest antioxidant and free radical scavenging activities among others could be a better choice for industrial purpose.

Development of pollen mediated activation tagging system for Phalaenopsis and Doritaenopsis

In the present study, a novel plant transformation system for Doritaenopsis and Phalaenopsis has been developed. The pollen-mediated activation tagging system was established by artificial pollination. The pollens, co-cultured with Agrobacterium tumefaciens strain EHA105 harbouring an activation tagging vector (pTAG-8), were used for pollination. In order to optimize the transformation efficiency, several factors (concentration of A. tumefaciens, concentration of acetosyringone during co-cultivation and the duration of co-cultivation) known to influence Agrobacterium-mediated DNA transfer were examined. A concentration of 0.5-1 x 10(8) CFU/ml for A. tumefaciens, 0.1 mM acetosyringone, and 6 hrs of co-culture period were found to be the optimal condition for high transformation efficiency. Integration of T-DNA into the genome of putative transgenic plants was confirmed by PCR and DNA blot analyses. Single copy of the transgene was observed in all transgenic plants analyzed. Most of the transgenic plants had a morphologically normal phenotype and the overall capsule formation efficiency was similar to control plant. Our results showed a new approach of genetic transformation in orchids and this method can be employed for genetic improvement of the orchids.

Wild bitter gourd improves metabolic syndrome: a preliminary dietary supplementation trial.

BACKGROUND: Bitter gourd (Momordica charantia L.) is a common tropical vegetable that has been used in traditional or folk medicine to treat diabetes. Wild bitter gourd (WBG) ameliorated metabolic syndrome (MetS) in animal models. We aimed to preliminarily evaluate the effect of WBG supplementation on MetS in Taiwanese adults. METHODS: A preliminary open-label uncontrolled supplementation trial was conducted in eligible fulfilled the diagnosis of MetS from May 2008 to April 2009. A total of 42 eligible (21 men and 21 women) with a mean age of 45.7 ± 11.4 years (23 to 63 years) were supplemented with 4.8 gram lyophilized WBG powder in capsules daily for three months and were checked for MetS at enrollment and follow-up monthly. After supplementation was ceased, the participants were continually checked for MetS monthly over an additional three-month period. MetS incidence rate were analyzed using repeated-measures generalized linear mixed models according to the intention-to-treat principle. RESULTS: After adjusting for sex and age, the MetS incidence rate (standard error, p value) decreased by 7.1% (3.7%, 0.920), 9.5% (4.3%, 0.451), 19.0% (5.7%, 0.021), 16.7% (5.4%, 0.047), 11.9% (4.7%, 0.229) and 11.9% (4.7%, 0.229) at visit 2, 3, 4, 5, 6, and 7 compared to that at baseline (visit 1), respectively. The decrease in incidence rate was highest at the end of the three-month supplementation period and it was significantly different from that at baseline (p = 0.021). The difference remained significant at end of the 4th month (one month after the cessation of supplementation) (p = 0.047) but the effect diminished at the 5th and 6th months after baseline. The waist circumference also significantly decreased after the supplementation (p < 0.05). The WBG supplementation was generally well-tolerated. CONCLUSION: This is the first report to show that WBG improved MetS in human which provides a firm base for further randomized controlled trials to evaluate the efficacy of WBG supplementation.