Hsin-Sheng Tsay

Studies on the Production of Some Important Secondary Metabolites from Medicinal Plants by Plant Tissue Cultures

Plants are a tremendous source for the discovery of new products of medicinal value for drug development. Today several distinct chemicals derived from plants are important drugs currently used in one or more countries in the world. Many of the drugs sold today are simple synthetic modifications or copies of the naturally obtained substances. The evolving commercial importance of secondary metabolites has in recent years resulted in a great interest in secondary metabolism, particularly in the possibility of altering the production of bioactive plant metabolites by means of tissue culture technology. Plant cell culture technologies were introduced at the end of the 1960s as a possible tool for both studying and producing plant secondary metabolites. Different strategies, using an in vitro system, have been extensively studied to improve the production of plant chemicals. The focus of the present review is the application of tissue culture technology for the production of some important plant pharmaceuticals. Also, we describe the results of in vitro cultures and production of some important secondary metabolites obtained in our laboratory.

Biological Screening of Medicinal Plants Collected from Eastern Ghats of India Using Artemia salina (Brine Shrimp Test)

Medicinal plants constitute important components of flora and are widely distributed in different regions of India. Based on ethnomedical significance, we have collected several medicinal plants used in traditional medicine from Eastern Ghats of India and evaluated for their biological activity. In the present study, a method utilizing brine shrimp (Artemia salina Leach) lethality was used to screen medicinal plants for their biological activity. Aqueous extracts from 118 Indian medicinal plants were screened by the brine shrimp lethality assay and found eleven out of the 118 extracts showed significant toxicity to the brine shrimp (＜60 μg/ml). Polygonum cuspidatum and Syzygium cumini extracts have exhibited potent activity with LC50 13.5 and 20, respectively. The results were analyzed within the context of the available traditional knowledge and uses for these plants. Present study could be useful in the search for new antitumor compounds from the Indian flora.

Anticancer effects of tanshinone I in human non-small cell lung cancer

Tanshinones are the major bioactive compounds of Salvia miltiorrhiza Bunge (Danshen) roots, which are used in many therapeutic remedies in Chinese traditional medicine. We investigated the anticancer effects of tanshinones on the highly invasive human lung adenocarcinoma cell line, CL1-5. Tanshinone I significantly inhibited migration, invasion, and gelatinase activity in macrophage-conditioned medium-stimulated CL1-5 cells in vitro and also reduced the tumorigenesis and metastasis in CL1-5-bearing severe combined immunodeficient mice. Unlike tanshinone IIA, which induces cell apoptosis, tanshinone I did not have direct cytotoxicity. Real-time quantitative PCR, luciferase reporter assay, and electrophoretic mobility shift assay revealed that tanshinone I reduces the transcriptional activity of interleukin-8, the angiogenic factor involved in cancer metastasis, by attenuating the DNA-binding activity of activator protein-1 and nuclear factor-κB in conditioned medium-stimulated CL1-5 cells. Microarray and pathway analysis of tumor-related genes identified the differentially expressed genes responding to tanshinone I, which may be associated with the Ras-mitogen-activated protein kinase and Rac1 signaling pathways. These results suggest that tanshinone I exhibits anticancer effects both in vitro and in vivo and that these effects are mediated at least partly through the interleukin-8, Ras-mitogen-activated protein kinase, and Rac1 signaling pathways. Although tanshinone I has a remarkable anticancer action, its potential anticoagulant effect should be noted and evaluated. [Mol Cancer Ther 2008;7(11):3527–38]

Cytokinin-induced somatic embryogenesis and plant regeneration in Corydalis yanhusuo (Fumariaceae) - a medicinal plant.

An efficient method has been developed for regeneration of complete plants via somatic embryogenesis in Corydalis yanhusuo (Fumariaceae), an important medicinal plant, using tuber-derived callus. Primary callus was induced by culturing mature tuber pieces on Murashige and Skoogs (MS) medium supplemented with 2.0 mg l(-1) N(6)-benzyladenine (BA) and 0.5 mg l(-1) alpha-naphthaleneacetic acid (NAA) in darkness. Somatic embryos were induced by subculturing the primary callus on MS medium supplemented with 0.5-4.0 mg l(-1) BA, kinetin, or zeatin, within 2 weeks of culture in light. Embryos with well-developed cotyledonary leaves were transferred in half-strength liquid MS medium supplemented with 1.0 mg l(-1) zeatin riboside for the development of roots. Converted somatic embryos were cultured on half-strength MS medium supplemented with 6% sucrose, and with 0.5-10.0 mg l(-1) abscisic acid (ABA), paclobutrazol, or ancymidol, 0.5-5.0 mg l(-1) GA(3) and 15-100 mg l(-1) polyethylene glycol (PEG) 4000 for further development of plantlets and in vitro tuber formation. The development of somatic embryos over the surface of tuber and/or cotyledonary leaf base region of the converted primary somatic embryo was observed. Before ex vitro establishment of somatic embryo-derived plants, plants with well-developed tubers were cultured on half-strength MS medium with 2% sucrose and 0.1 mg l(-1) GA(3) for 3 weeks.

ASYMBIOTIC GERMINATION OF IMMATURE SEEDS, PLANTLET DEVELOPMENT AND EX VITRO ESTABLISHMENT OF PLANTS OF DENDROBIUM TOSAENSE MAKINO - A MEDICINALLY IMPORTANT ORCHID

SummaryShi-hu (Dendrobium spp. or Dendrobii Herba) is one of the important traditional Chinese medicines. The commercially available crude drug in the traditional medicine market is composed mainly of three species: Dendrobium tosaense, D. nobile, and D. moniliforme. An efficient method of propagation has been developed via asymbiotic germination of seeds in vitro for the medicinally important D. tosaense. Seeds from capsules of D. tosaense collected 8–14 wk after artificial pollination germinated after being cultured on full-strength or half-strength Murashige and Skoog (MS) medium devoid of plant growth regulators and with 3% sucrose. Germination of seeds varied with the medium type and seed maturity. Germinated seedlings after transfer to MS medium with 1.5% sucrose and 8% banana homogenate or potato juice or coconut water and 20 wk of incubation developed into healthy plantlets. Well-developed plantlets were transplanted to moss or moss and tree fern or tree fern as substrates in plastic trays and transferred to a greenhouse for hardening. All plants survived, attained maturity, and developed normal flower and capsule after one and a half years. This protocol of successful plant regeneration by asymbiotic seed germination should permit rapid propagation and conservation of this medicinally important Dendrobium species.

Withanolide sulfoxide from Aswagandha roots inhibits nuclear transcription factor-kappa-B, cyclooxygenase and tumor cell proliferation.

Investigation of the methanol extract of Aswagandha (Withania somnifera) roots for bioactive constituents yielded a novel withanolide sulfoxide compound (1) along with a known withanolide dimer ashwagandhanolide (2) with an S‐linkage. The structure of compound 1 was established by extensive NMR and MS experiments. Compound 1 was highly selective in inhibiting cyclooxygenase‐2 (COX‐2) enzyme by 60% at 100 µm with no activity against COX‐1 enzyme. The IC50 values of compound 1 against human gastric (AGS), breast (MCF‐7), central nervous system (SF‐268) and colon (HCT‐116) cancer cell lines were in the range 0.74–3.63 µm. Both S‐containing dimeric withanolides, 1 and 2, completely suppressed TNF‐induced NF‐κB activation when tested at 100 µm. The isolation of a withanolide sulfoxide from W. somnifera roots and its ability to inhibit COX‐2 enzyme and to suppress human tumor cell proliferation are reported here for the first time. In addition, this is the first report on the abrogation of TNF‐induced NF‐κB activation for compounds 1 and 2. Copyright

IN VITRO PROPAGATION OF SOME IMPORTANT CHINESE MEDICINAL PLANTS AND THEIR SUSTAINABLE USAGE

SummaryMedicinal plants are valuable sources of medicinal and many other pharmaceutical products. The conventional propagation method is the principal means of propagation and takes a long time for multiplication because of a low rate of fruit set, and/or poor germination and also sometimes clonal uniformity is not maintained through seeds. The plants used in the phyto-pharmaceutical preparations are obtained mainly from the natural growing areas. With the increase in the demand for the erude drugs, the plants are being overexploited, threatening the survival of many rate species. Also, many medicinal plant species are disappearing at an alarming rate due to rapid agricultural and urban development, uncontrolled deforestation, and indiscriminate collection. Advanced biotechnological methods of culturing plant cells and tissues should provide new means for conserving and rapidly propagating valuable, rare, and endangered medicinal plants. The purpose of the present review is to focus the application of tissue culture technology for in vitro propagation via somatic embryogenesis and organogenesis and the cell suspension culture with suitable examples reported earlier. An overview of tissue culture studies on important Chinese medicinal plants and related species is presented.

IN VITRO PROPAGATION OF THE CHINESE MEDICINAL PLANT, DENDROBIUM CANDIDUM WALL. EX LINDL., FROM AXENIC NODAL SEGMENTS

SummaryDendrobium candidum Wall. Ex Lindl. is an important species used in the formulation of Shih-hu, a Chinese traditional medicine. An efficient protocol for in vitro propagation of D. candidum using the axenic nodal segments of the shoots, originated from the in vitro germinated seedlings, was developed. The seeds from 120-d-old capsules after pollination were first germinated on half-strength Murashige and Skoog (MS) basal medium supplemented with 30 g l−1 sucrose. After 4 mo., the seedlings were subcultured on a similar medium supplemented with 1 ml l−1 HYPONeX, 80 g l−1 potato homogenate and 2 g l−1 activated charcoal for further growth. Axenic nodal segments excised from 9-mo.-old seedlings were cultured on the medium in the presence of 2 mg l−1 benzyladenine (BA) and 0.1 mg l−1 naphthaleneacetic acid (NAA). After 75 d, 73.2% of the explants gave rise to buds/shoots. The elongated shoots were rooted on the medium containing 0.2 mg l−1 NAA and the plantlets were successfully acclimatized in soil.

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.

Wild bitter gourd improves metabolic syndrome: A preliminary dietary supplementation trial

BackgroundBitter 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.MethodsA 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.ResultsAfter 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.ConclusionThis 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.