| PART ⅠThe windmill palm (Trachycarpus fortunei) is a rich source of pharmaceutically important compounds. These chemicals include polyphenols, flavonoids and steroidal saponin found abundantly in seeds and leaves. Previously random anti-bacterial screening from a list of plants showed that T. fortunei possess remarkable activity against Escherichia coli. In this research,further studies were performed to explore the antibiotic properties of seed coat of T. fortunei.In the first phase of study, dried seed coat material of T. fortunei was extracted with aqueous and ethanolic solution. In this experiment extraction was performed by common Soxhlet apparatus. Further fractionation of obtained extracts was performed using macroporous resin D101 in column chromatography and four different fractions was made i.e. 0%, 20%, 40% and 60% ethanol (EtOH). In these preliminary experiments,the highest yield was obtained from 20% EtOH fraction as compared to the rest of the fractions. Material obtained from these fractions was used in-vitro antibacterial studies against human pathogenic bacterial strains including Staphylococcus aureus,Staphylococcus epidermidis, Pseudomonas aeruginosa, Bacillus cereus, Bacillus subtilis and E. colil Based on the findings of this experiment, maximum antibacterial activity was observed in case of 20% EtOH fraction against most of the tested bacterial strains.Minimum Inhibitory Concentration (MIC) analysis of this fraction was performed to ensure its use as plant derived antimicrobial drug for in-vivo use. The lowest possible MIC value of this fraction was recorded as156.25ug/mL against S. aureus and E. coli.Along with that, cytotoxicity analysis was performed by using human cell lines (HMEC-1). The 20% EtOH fraction at the concentration of 250ug/mL showed the closest range of cell survival percentage level.Second phase of this study was aimed on identification of bioactive compounds in best performing fraction as provided by previous experimental procedure. For that purpose 20% EtOH fraction was further analyzed on Ultra-Performance Liquid Chromatography/ mass spectroscopy (UPLC-MS) system to identify the compounds present in that fraction along with the rest of fractions and a list of compounds were identified. Bioassays and principle component analysis provided that among these identified compounds, 2,4,5-triacetoxybiphenyl (10) and 1-(4-Fluorophenyl)-2-(methylthio)-1H-imidazole-5-carboxylic acid (9) actively inhibited the growth of the test bacterial strains whereas stearamide (7) was unable to show inhibition potential against any of bacterial strains. Further experiment suggested that compounds 9 and 10 have antibacterial activity against gram-negative bacteria. Whereas, the lowest minimum inhibitory concentration value (39.06 μg/mL) was obtained by compound 10 against S.epidermidis. The antibacterial activity of 20% EtOH fraction may be attributed to these two compounds i.e. compound 9 (1-(4-Fluorophenyl)-2-(methylthio)-1H-imidazole-5-carboxylic acid) and compound 10 (2,4,5-triacetoxybiphenyl).However antibacterial potential of pure compounds was found to be just comparable to parent 20% EtOH fraction. This might be due to synergetic effect from other compounds. Our findings suggest that seed coat material of T. fortunei possess antibacterial properties and can serve for the development of novel antimicrobial agents for pharmaceutical industries.PART ⅡAtractylodes lancea (Thunb.) DC., named "Cangzhu" in China, which belongs to the Asteraceae family. In some countries of Southeast Asia (China, Thailand, Korea,Japan etc.) its rhizome, commonly called rhizoma atractylodis, is used to treat many diseases as it contains a variety of sesquiterpenoids and other components of medicinal importance. Despite its medicinal value, the information of the sesquiterpenoid biosynthesis is largely unknown.In this study, we investigated the transcriptome analysis of different tissues of non-model plant A. lancea by using short read sequencing technology (Illumina).We found that there were 62,352 high quality unigenes with an average sequence length of 913 bp in the transcripts of A. lancea. Among these, 43,049 (69.04%), 30,264 (48.53%),26,233 (42.07%), 17,881 (28.67%) and 29,057(46.60%) unigenes showed significant similarity (E-value<le-5) to known proteins in Nr, KEGG, SWISS-PROT, GO, and COG databases, respectively. Of the total 62,352 unigenes, 43,049 (Nr Database) open reading frames were predicted. On the basis of different bioinformatic tools we identified all the enzymes that take part in the terpenoid biosynthesis as well as five different known sesquiterpenoids via cytosolic mevalonic acid (MVA) pathway and plastidal methylerythritol phosphate (MEP) pathways. In our study, 6, 864 Simple Sequence Repeats (SSRs) were also found as great potential markers in A. lancea.This transcriptomic resource of A. lancea provides a great contribution in advancement of research for this specific medicinal plant and more specifically for the gene mining of different classes of terpenoids and other chemical compounds that have medicinal as well as economic importance. |
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