Taxus Wallichiana Research Articles

Brevifoliol and its Analogs: A New Class of Anti-tubercular Agents.

Brevifoliol is an abeo-taxane isolated from the Taxus wallichiana needles; eighteen semisynthetic esters derivatives of brevifoliol were prepared by Steglich esterification and screened for their anti-tubercular potential against Mycobacterium tuberculosis H37Ra avirulent strain. The 3- [chloro (7)] and 3, 5-[dinitro (8)] benzoic acid ester derivatives were most active (MIC 25 ug/ml) against the pathogen. Further, in silico docking studies of the active derivative 7 with mycobacterium enzyme inhA (enoyl-ACP reductase) gave the LibDock score of 152.68 and binding energy of -208.62 and formed three hydrogen bonds with SER94, MET98, and SER94. Similarly, when derivative 8 docked with inhA, it gave the LibDock score of 113.55 and binding energy of -175.46 and formed a single hydrogen bond with GLN100 and Pi-interaction with PHE97. On the other hand, the known standard drug isoniazid (INH) gave the LibDock score of 61.63, binding energy of -81.25 and formed one hydrogen bond with ASP148. These molecular docking results and the way of binding pattern indicated that compounds 7 and 8 bound well within the binding pocket of inhA and showed a higher binding affinity than the known drug isoniazid. Additionally, both the derivatives (7 and 8) showed no cytotoxicity, with CC50 195.10 and 111.36, respectively towards the mouse bone marrow-derived macrophages.

Evaluation of antimicrobial, antioxidant and cytotoxic properties of bioactive compounds produced from endophytic fungi of Himalayan yew ( Taxus wallichiana) in Nepal.

Background: Endophytic fungi are largely underexplored in the discovery of natural bioactive products though being rich sources of novel compounds with promising pharmaceutical potential. In this study, Taxus wallichiana, which has huge medicinal value, was investigated for its endophytic diversity and capability to produce bioactive secondary metabolites by analyzing antioxidant, antimicrobial and cytotoxic properties. Methods: The endophytes were identified by ITS-PCR using genomic DNA samples. The secondary metabolites were extracted by solvent extraction method using ethyl acetate. The antioxidant activity was analyzed by Thin Layer Chromatography, Total Phenol Content (TPC), Total Flavonoid Content (TFC) and DPPH assay, and the antimicrobial activity was analyzed by agar-well diffusion method. Brine shrimp lethality assay was used to analyze the cytotoxicity of the fungal extracts. Results: Out of 16 different Taxus trees sampled from different locations of Dhorpatan, 13 distinctive endophytic fungi were isolated and grouped into 9 different genera: Bjerkandera, Trichoderma, Preussia, Botrytis, Arthrinium, Alternaria, Cladosporium, Sporormiella and Daldinia. The ethyl acetate extracts isolated from three endophytic fungi: Alternaria alternata, Cladosporium cladosporioides and Alternaria brassicae showed significant TPC values of 204±6.144, 312.3±2.147 and 152.7±4.958µg GAE/mg of dry extract, respectively, and TFC values of 177.9±2.911, 644.1±4.202 and 96.38±3.851µg RE/mg of dry extract, respectively. Furthermore, these three extracts showed a dose dependent radical scavenging activity with IC 50 concentration of 22.85, 22.15 and 23.001 µg/ml, respectively. The extracts of C. cladosporioides and A. brassicae also showed promising antimicrobial activity against Escherichia coli, Staphylococcus aureus and Bacillus subtilis with a minimum inhibitory concentration of 250μg/ml for all bacteria. Both the samples showed cytotoxic property against shrimp nauplii with LC 50 of 104.2 and 125.9µg/ml, respectively. Conclusions: The crude fungal extracts obtained from endophytes: A. alternata, C. cladosporioides and A. brassicae upon purification and further identification of the bioactive compounds can be a fascinating source for novel pharmaceutical agents.

Tree diversity and community composition in sacred forests are superior than the other community forests in a human-dominated landscape of Meghalaya

Tree diversity and community composition in six sacred forests (SF) of Meghalaya were compared with those of six adjacent community forests (CF) to understand how protection provided to sacred forests by the communities impacts their diversity and community characteristics. The SFs harboured a greater number of tree species (187) compared to the CFs (56). Tree density in the SFs ranged from 750 trees ha−1 (MKSF) to 1470 trees ha−1 (MTSF) while in CFs, it was 875 trees ha−1 (ILCF) to 1236 trees ha1 (NBCF). Among the SFs, the highest basal area was recorded in MNSF (56.89 m2 ha−1) and least in NBSF (40.75 m2 ha−1), while among the CFs tree basal area ranged from 23.7 m2 ha−1 in MKCF to 36.0 m2 ha−1 in MNCF. The canopy cover in SFs ranged from 73.3 to 86.2% while in CFs it ranged from 54.3 to 70.8%. Species rank abundance distribution (RAD) curve or dominance-diversity curve showed high equitability and low dominance in all the six SFs while the RAD curve showed high dominance and low equitability in all the six CFs. The dominance-diversity curve in the SFs followed a log-normal distribution whereas the CFs exhibited a broken stick distribution pattern. Among the CFs, Pinus kesiya was the dominant tree species in five out of six study sites. The dominant species were different for different SFs, which were Microtropis discolour (ILSF), Citrus latipes (MNSF), Castanopsis tribuloides (MTSF), Castanopsis armata (NBSF), Myrica esculenta (NKSF) and Taxus wallichiana (MKSF). SFs were characterized by high diversity and low dominance as seen in old-growth forests. Shannon’s diversity index (H′) in the SFs ranged from 2.06 to 4.28 indicating high tree diversity, while in CFs it was 0.76–2.61 reflecting relatively lower diversity. Simpson’s dominance index for trees was higher in CFs (0.09–0.73) than the SFs (0.02–0.26).

Ethnobotanical and pharmacological importance of Taxus wallichiana Zucc.

Taxus wallichiana Zucc. or the Himalayan Yew is a gymnosperm growing along the Himalayan region of Indian and adjoining countries. The plant is extensively used by local people for treatment of various diseases such as fever, headache, diarrhea, fractures, problems of nervous system etc. It also finds usage in Unani system of medicine. The plant is rich in various bioorganic compounds natural products such as hydrocarbons, terpene alcohols, terpenoids (including taxoids), organic acids etc. The plant has been explored for anti-inflammatory, analgesic, antipyretic, anticonvulsant, immunomodulatory, hepatoprotective and anticancer activity with satisfactory outcome. The pharmacological activity of the plant is largely due to the presence of large number of terpenoids. The bioactive constituents present in the plant interacts with a large number of biochemical pathways involved in inflammatory processes, cell division cycles and inhibits a number of enzymes to bring about its protective action against various diseases. In this review, an attempt have been made to highlight the beneficial properties of Taxus wallichiana in various levels of usage starting from its fundamental ethnobotanical use to pharmacological use involving both in-vitro and in-vivo studies. Insights into the molecular mechanisms of action of the active constituents in bringing about the beneficial activity have also been illustrated. The plant can very well become a source of medicine for better management of a large number of diseases including cancer.

Comparative Genomic Analysis Reveals the Mechanism Driving the Diversification of Plastomic Structure in Taxaceae Species.

Inverted repeat (IR) regions in the plastomes from land plants induce homologous recombination, generating isomeric plastomes. While the plastomes of Taxaceae species often lose one of the IR regions, considerable isomeric plastomes were created in Taxaceae species with a hitherto unclarified mechanism. To investigate the detailed mechanism underpinning the IR-independent genesis of plastomic diversity, we sequenced four Taxaceae plastomes, including Taxus cuspidata Siebold & Zuccarini, Taxus fauna Nan Li & R. R. Mill, and two individuals of Taxus wallichiana Zuccarini. Then we compared these structures with those of previously reported Taxaceae plastomes. Our analysis identified four distinct plastome forms that originated from the rearrangements of two IR-flanking inverted fragments. The presence of isomeric plastomes was then verified in T. cuspidata individuals. Both rearrangement analyses and phylogenetic results indicated that Taxaceae were separated into two clades, one including Taxus and Pseudotaxus and another formed by Amentotaxus and Torreya. Our reconstructed scenario suggests that the minimum number of inversion events required for the transformation of the plastome of Cephalotaxus oliveri Masters into the diversified Taxaceae plastomes ranged from three to six. To sum up, our study reveals a distinct pattern and the mechanism driving the structural diversification of Taxaceae plastomes, which will advance our understanding of the maintenance of plastomic diversity and complexity in conifers.

Strategic engineering for detecting antimicrobial compounds from <i>Taxus wallichiana Zucc.</i> (Himalayan yew)

Strategic engineering for detecting antimicrobial compounds from <i>Taxus wallichiana Zucc.</i> (Himalayan yew)

Quantification of taxol by high-performance thin layer chromatography in Taxus wallichiana callus cultivated in vitro

ENWEndNote BIBJabRef, Mendeley RISPapers, Reference Manager, RefWorks, Zotero AMA Mujib A, Tonk D, Gulzar B, Maqsood M, Ali M. Quantification of taxol by high-performance thin layer chromatography in Taxus wallichiana callus cultivated in vitro. BioTechnologia. 2020;101(4):337-347. doi:10.5114/bta.2020.100425. APA Mujib, A., Tonk, D., Gulzar, B., Maqsood, M., & Ali, M. (2020). Quantification of taxol by high-performance thin layer chromatography in Taxus wallichiana callus cultivated in vitro. BioTechnologia, 101(4), 337-347. https://doi.org/10.5114/bta.2020.100425 Chicago Mujib, Abdul, Dipti Tonk, Basit Gulzar, Mehpara Maqsood, and Muzamil Ali. 2020. "Quantification of taxol by high-performance thin layer chromatography in Taxus wallichiana callus cultivated in vitro". BioTechnologia 101 (4): 337-347. doi:10.5114/bta.2020.100425. Harvard Mujib, A., Tonk, D., Gulzar, B., Maqsood, M., and Ali, M. (2020). Quantification of taxol by high-performance thin layer chromatography in Taxus wallichiana callus cultivated in vitro. BioTechnologia, 101(4), pp.337-347. https://doi.org/10.5114/bta.2020.100425 MLA Mujib, Abdul et al. "Quantification of taxol by high-performance thin layer chromatography in Taxus wallichiana callus cultivated in vitro." BioTechnologia, vol. 101, no. 4, 2020, pp. 337-347. doi:10.5114/bta.2020.100425. Vancouver Mujib A, Tonk D, Gulzar B, Maqsood M, Ali M. Quantification of taxol by high-performance thin layer chromatography in Taxus wallichiana callus cultivated in vitro. BioTechnologia. 2020;101(4):337-347. doi:10.5114/bta.2020.100425.

Assessment of risk, extinction, and threats to Himalayan yew in Pakistan.

Himalayan yew (Taxus wallichiana) is in high demand due to the presence of taxol in its bark, needles, and seeds. This metabolite is used for the treatment of breast and ovarian cancer. In addition, Himalayan yew wood is used to prepare slabs (Tabai), coffins (Taabut), for graveyards. Due to illegal cutting of plant parts and other anthropogenic pressures, Himalayan yew is endangered, and threatened with extinction, in Himalaya. This species grows slowly and regenerates poorly, primarily due to low production and delayed germination (1.5–2 years) of its seeds. The study being reported here was conducted to assess the factors (natural and anthropogenic) threatening this species. Nine valleys (Miandam, Kalam, Shinko, Beha, Lalku, Shahgram, Bishigram, Gurnai, and Daral) in the Swat district of Khyber Pakhtunkhwa (KP), Pakistan, have stands of Himalayan yew that were selected for the study. Before the survey was conducted, five informal discussions were carried out to identify people to be interviewed. A survey was conducted with 225 key informants in these valleys concerning the threats associated with this species. Nineteen percent of the respondents felt that the main problem was lack of awareness, while 17% indicated over-harvesting (peeling bark, lopping branches, etc.), and 13% thought it was slow growth. Other reasons for Himalayan yew decline included various anthropogenic pressures, such as: overgrazing, 15%; agriculture, 11%; roof construction, 9%; fuelwood, 7%; decoration, 5%; medicinal use, 3%; and other, 1% (e.g., utility poles, as blades in water turbine because of its hard nature). The results of this study suggest that there is an immediate need to protect T. wallichiana by increasing awareness of its importance and the threats from over-grazing; cuttings (peeling bark, lopping branches, etc.); and other damaging, anthropogenic activities. Biotechnological tools, such as vegetative propagation and in-vitro regeneration, could be practiced in nurseries and laboratories to produce large numbers of healthy, juvenile plants. In addition to in-situ and ex-situ conservation and management, there is a need for local community involvement in the large-scale reforestation efforts.

A study of the effect of Paclitaxel, Docetaxel and Tamoxifen citrate coumpounds found in Taxus wallichiana on the Beta-tubulin and Estrogen Receptor (ESR) proteins in humans using a bioinformatics approach

Taxus wallichiana, an indigenous plant of North East India contains bioactive compounds such as Tamoxifen citrate and taxol (paclitaxel and docetaxel) which inhibit the growth of cancerous cells. Tamoxifen citrate in its active form competitively inhibits the binding of estradiol or estrogen with estrogen receptor and results in tumor suppression whereas the taxol compounds stabilize the tubulin protein in microtubules and inhibit cell division in cancerous cells. In this approach, important domains and motifs of the protein receptors were studied and analyzed followed by homology modeling and its validation. Molecular docking studies were carried out via tubulin and estrogen receptor with paclitaxel, docetaxel, and endoxifen (an active form of tamoxifen citrate). Hence it was verified that tubulin and estrogen receptor contain binding sites for the bioactive compounds found in T. wallichiana and these bioactive compounds after binding with these receptors result in the prevention of the proliferation of the tumorous cells by stabilizing the microtubules and preventing the intake of steroidal hormones by cells respectively.

In-vitro and in-silico anticancer potential of taxoids from Taxus wallichiana Zucc.

Natural products derived from medicinal plants provide beneficial cancer chemotherapeutic drugs. Bioactive constituents from plants are explored for their anticancer properties. Three known compounds (deacetylbaccatin III, tasumatrol B, and taxawallin J) were isolated from Taxus wallichiana. Compounds were screened against four cancer cell lines, such as eA498, HepG2, NCI-H226, and MDR 2780AD. Cytotoxic activity was evaluated using MTT assay against cancer cell lines. Tasumatrol B showed good cytotoxic activity conducted for the improvement of inhibiting potential of these compounds against the cancer drug target protein (EGFR tyrosine kinase enzyme). The docking study showed that all compounds have binding affinities and interaction profile with the receptor tyrosine kinase. The study suggests that these compounds could be used for the discovery of novel inhibitors against the target receptors for the treatment of cancer.

Efficient Heated Ultrasound Assisted Extraction and Clean-Up Method for Quantifying Paclitaxel Concentrations in Taxus Wallichiana

ABSTRACT A simple method was developed using heated ultrasound-assisted extraction (UAE) with ethanol and an ENVI-Carb clean-up step for quantifying paclitaxel from various parts (leaf, stem, bark and roots) of T. wallichiana with standard liquid chromatography using a Waters Novapak Phenyl column with ultraviolet detection (HPLC-UV). Method effectiveness and accuracy were evaluated by quantifying paclitaxel independently in multiple sequential extractions, comparing HPLC-UV quantitation with quadrapole time-of-flight mass spectrometry (QToF/MS) time-of-flight to confirm the adequacy of HPLC-UV and assessing the use of docetaxel as a paclitaxel surrogate. Nearly identical trends for % recovered with each sequential extract was observed for both compounds. ENVI-Carb clean-up removed most of the colours in the extract, did not affect the recovery of paclitaxel or docetaxel in the presence of the sample matrix, and minimised compounds that may interfere with paclitaxel quantitation. For both docetaxel and paclitaxel, most of the mass was extracted in the first two extractions with 93.1% ± 4.4% and 92.6% ± 3.9%, respectively. The mass sum recovered in the first three extractions was similar to analysis of the combined extracts with 95–99% extracted relative to the total recovered from four sequential extractions. Paclitaxel concentrations in ENVI-Carb cleaned extracts were similar between HPLC-UV and uPLC-ToF/MS (using the 551 product ion) supporting the reliability of the method. The similar recovery trend between docetaxel and paclitaxel indicates that extractions could be limited to a single step if docetaxel is used to correct for recovery.

Distribution of paclitaxel and its precursors in different parts of Taxus wallichiana var. Mairei

To understand the distributions of paclitaxel and its precursors in the different parts of yew this study was conducted. The contents of paclitaxel and its precursors in the different parts of Taxus wallichiana var. mairei were determined with RP-HPLC. The contents of paclitaxel and its precursors in yew root are higher than those in other parts. The content of paclitaxel in the bark of Taxus wallichiana var. mairei is higher than that in leaf. But there was very little paclitaxel in the xylem and tender stem of Taxus wallichiana var. mairei. There were considerable paclitaxel precursors in the leaf of Taxus wallichiana var. mairei, but low contents in 2 - 3 years stem. The root of Taxus wallichiana var. mairei can be utilized also. It is reasonable measures of satisfying the demand of people on paclitaxel and preventting people cutting yew that artificially synthesize paclitaxel with the precursors extracted from yew leaf.

Chloroplast population genetics reveals low levels of genetic variation and conformation to the central-marginal hypothesis in Taxus wallichiana var. mairei, an endangered conifer endemic to China.

The central–marginal hypothesis predicts that geographically peripheral populations should exhibit reduced genetic diversity and increased genetic differentiation than central populations due to smaller effective population size and stronger geographical isolation. We evaluated these predictions in the endangered conifer Taxus wallichiana var. mairei. Eight plastid simple sequence repeats (cpSSRs) were used to investigate plastid genetic variation in 22 populations of Taxus wallichiana var. mairei, encompassing nearly its entire distribution range. Low levels of plastid genetic variation and differentiation were detected in the populations, and the findings were attributed to low mutation rates, small population sizes, habitat fragmentation and isolation, and effective pollen or seed dispersal. Hunan and Hubei were identified as major refugia based on the number of private haplotypes and species distribution modeling. Trends in plastid genetic diversity and genetic differentiation from central to peripheral populations supported the predictions of the central–marginal hypothesis. In scenarios wherein the future climate becomes warmer, we predict that some peripheral populations will disappear and southern and southeastern regions will become significantly less habitable. Factors that include the levels of precipitation during the driest month, annual precipitation level, and annual temperature range will be decisive in shaping the future distribution of these populations. This study provides a theoretical basis for the conservation of T. wallichiana var. mairei.

Integration of multiple climate models to predict range shifts and identify management priorities of the endangered Taxus wallichiana in the Himalaya\u2013Hengduan Mountain region

Taxus wallichiana Zucc. (Himalayan yew) is subject to international and national conservation measures because of its over-exploitation and decline over the last 30 years. Predicting the impact of climate change on T. wallichiana’s distribution might help protect the wild populations and plan effective ex situ measures or cultivate successfully. Considering the complexity of climates and the uncertainty inherent in climate modeling for mountainous regions, we integrated three Representative Concentration Pathways (RCPs) (i.e., RCP2.6, RCP4.5, RCP8.5) based on datasets from 14 Global Climate Models of Coupled Model Intercomparison Project, Phase 5 to: (1) predict the potential distribution of T. wallichiana under recent past (1960–1990, hereafter “current”) and future (2050s and 2070s) scenarios with the species distribution model MaxEnt.; and (2) quantify the climatic factors influencing the distribution. In respond to the future warming climate scenarios, (1) highly suitable areas for T. wallichiana would decrease by 31–55% at a rate of 3–7%/10a; (2) moderately suitable areas would decrease by 20–30% at a rate of 2–4%/10a; (3) the average elevation of potential suitable sites for T. wallichiana would shift up-slope by 390 m (15%) to 948 m (36%) at a rate of 42–100 m/10a. Average annual temperature (contribution rate ca. 61%), isothermality and temperature seasonality (20%), and annual precipitation (17%) were the main climatic variables affecting T. wallichiana habitats. Prior protected areas and suitable planting areas must be delimited from the future potential distributions, especially the intersection areas at different suitability levels. It is helpful to promote the sustainable utilization of this precious resource by prohibiting exploitation and ex situ restoring wild resources, as well as artificially planting considering climate suitability.

Relationship of plant abundance, air temperature and humidity with negative ions based on control experiment

Previous studies on negative air ion (NAI), an important index for evaluating atmospheric quality, has been focused on field observation, and less on NAI under controlled condition. In this study, the NAI concentrations of different individual abundance of Liquidambar formosana and Taxus wallichiana were continuously monitored under the same climatic conditions in Hushan Experimental Base of Qianjiangyuan Forest Ecosystem Research Station, Zhejiang Province from September to October 2018. Changes of NAI concentration were monitored under different levels of air temperature and relative humidity to explore the effects of forest vegetation and meteorological factors on NAI. The results showed that both species significantly increased the NAI concentration. Plant abundance was positively correlated with the NAI concentration, and the relationship between them fitted the quadratic function with the plant abundance ranging from 0 to 50. The fitting equations for L. formosana and T. wallichiana were as follows: y=-0.0484x2+4.7005x+345.7 (R2=0.62), y=-0.0207x2+1.9189x+365.91 (R2=0.34). There was a significant positive correlation between NAI concentration and air temperature in the range of 5-30 ℃ with a fitting equation of y=0.4139x2-9.2229x+89.919 (R2=0.92). The NAI concentration and the relative humidity of air in the range of 56%-87% were positively correlated with a fitting equation of y=3.6508e0.0526x(R2=0.94).

Transcriptome Sequencing Reveals Regulatory Mechanisms of Taxol Synthesis in Taxus wallichiana var. Mairei.

Taxol is one of the most potent and effective anticancer drugs and is originally isolated from Taxus species. To investigate the specific regulatory mechanisms of taxol synthesis in Taxus wallichiana var. mairei, RNA-seq was conducted to reveal the differences in transcriptional levels between wild type (WT) and “Jinxishan” (JXS), a cultivar selected from a population of Taxus mairei that shows about 3-fold higher taxol content in the needles than WT. Our results indicated that high expressions of the genes taxadienol acetyltransferase (TAT), taxadiene 5-alpha hydroxylase (T5H), 5-alpha-taxadienol-10-beta-hydroxylase (T10OH), and 2-debenzoyl-7,13-diacetylbaccatin III-2-O-benzoyl-transferase (DBBT), which catalyze a series of key acetylation and hydroxylation steps, are the main cause of high taxol content in JXS. Moreover, in the present study, the activation of jasmonic acid (JA) signal transduction and its crosstalk with gibberellin (GA), auxin, and ethylene (ET) explained the elevation of differentially expressed genes (DEGs) from the taxol biosynthesis pathway. This also indicates that taxol biosynthesis in T. mairei is associated with the balance of cell development and defense. TF-encoding (transcriptional factor) genes, represented by the ethylene-responsive transcription factor (ERF), basic/helix-loop-helix (bHLH), MYB, and WRKY families, were detected as differentially expressed between JXS and WT, further indicating that the regulation of hormone signaling on taxol biosynthesis genes was mediated by transcription factors (TFs). To our knowledge, this is the first study to illustrate the regulatory mechanisms of taxol synthesis in a new cultivar of T. mairei with a high taxol content in its needles. These transcriptome data provide reasonable explanations for the variation of taxol content between WT and JXS.

Modelling the vulnerability of Taxus wallichiana to climate change scenarios in South East Asia

Taxus wallichiana (Himalayan yew) has been subjected to over-exploitation for its anti-cancer ingredient Taxol. As a consequence, the species has already become endangered, facing enhanced threats of regional extinction. To compound this, the species shows high vulnerability to climate change impacts that may significantly reduce its present climatic niche. In spite of the ecological and enormous commercial importance of the species, the direct or indirect impacts of climate change on the species are not yet well-researched and well-documented. To develop sustainable adaptation pathways to conserve the surviving populations of the species, it is important to have precise assessments of the future distribution ranges of the species. The present study models the potential current and future distribution ranges of Taxus wallichiana based on its suitable climatic envelop developed under a baseline scenario (1960–1990) and climate change scenarios centred on representative concentration pathways (RCPs) for the year 2070, as provided in the Fifth Assessment Report (AR5) of the UNO’s Intergovernmental Panel on Climate Change (IPCC). The projected shrink in climatic niche of Taxus wallichiana by 28% (RCP 4.5) and 31% (RCP 8.5) highlights the vulnerability of the endangered species to climate change impacts and the perturbations on the structure of mountain ecosystem. It raises an alarm for immediate conservation of geographic areas that can provide potential refuge to the species under the adverse climate change impacts.

Phosphate solubilization potential of endophytic fungi isolated from Taxus wallichiana Zucc. roots

Endophytic microorganisms live inside the host plant and contribute in various biological processes, without causing any harmful effect. Inorganic phosphate solubilization, through microorganisms, is one of the major mechanisms involved in plant growth. The present study highlights the potential of endophytic fungi for their ability to solubilize insoluble phosphates in presence of tricalcium (TCP), aluminium (AlP), and iron phosphate (FeP) at different temperatures through production of phosphatases, phytases and organic acids. Five endophytic fungi, isolated from the roots of Taxus wallichiana, were identified following their phenotypic and molecular characters. Three fungal isolates showed maximum similarity with species of Penicillium (GBPI TWR_F1, GBPI TWR_F2, and GBPI TWR_F3) and two with species of Aspergillus (GBPI TWR_F4 and GBPI TWR_F5). All the endophytes solubilized phosphate by utilizing the substrates namely calcium, aluminium and iron phosphate along with the production of phosphatase and phytase enzymes. Maximum phosphate solubilization and phytase activity was recorded in case of the fungal isolate GBPI TWR_F2 (P. daleae) being 83.42 ± 3.41 µg/ml TCP, 57.63 ± 0.79 µg/ml AlP, and 57.76 ± 1.70 µg/ml FeP at 15 °C. GBPI TWR_F2 and GBPI TWR_F5 (Aspergillus sp.) produced maximum calcium phytase at 25 and 15 °C, 10.33 ± 0.13 and 10.37 ± 0.37 µM/ml, respectively. Phosphatase production was higher in acidic conditions in comparison to alkaline. In quantification of organic acids through HPLC, malic and succinic acids were determined in maximum quantity 0.97 ± 0.003 and 0.92 ± 0.008 µg/ml, respectively, followed by oxalic (0.71 ± 0.006 µg/ml) and lactic acid (0.61 ± 0.005 µg/ml). Citric acid was estimated in minimum quantity.

The Research Progress of Chalcone Isomerase (CHI) in Plants.

Chalcone isomerase (CHI) is the second rate-limiting and the first reported enzyme involved in the biosynthetic pathway of flavonoids. It catalyzes the intramolecular cyclization reaction, converting the bicyclic chalcone into tricyclic (2S)-flavanone. In this paper, we obtained and analyzed 916 DNA sequences, 1310 mRNA sequences, and 2403 amino acid sequences of CHI registered in NCBI by Jan 2018. The full length of CHI DNA sequences ranges from 218 to 3758bp, CHI mRNA sequences ranges from 265 to 1436bp, and CHI amino acid sequences ranges from 35 to 465 amino acid residues. Forty representative species were selected from each family to construct the maximum likelihood tree and analyze the evolutionary relationship. According to the medicinal and agricultural use, 13 specific species were selected, and their physicochemical properties were analyzed. The molecular weight of CHI ranges from 23 to 26kD, and the isoelectric point of CHI ranges from 4.93 to 5.85. All the half-life periods of CHI are 30h in mammalian reticulocytes in vitro, 20h in yeast, and 10h in E. coli in vivo, theoretically. The consistency of the 13 CHI amino acid sequences is 63.55%. According to the similarity between each sequence, we selected four CHI sequences of Paeonia suffruticosa, Paeonia lactiflora, Taxus wallichiana, and Tradescantia hirsutiflora for secondary structure, three-dimensional protein models, conserved domains, transmembrane structure, and signal peptide prediction analysis. It was found that CHI sequences of Paeonia suffruticosa and Paeonia lactiflora owned a higher similarity; they both share the template 4doi.1.A. The four CHI all have no signal peptides, and they exert their activities in cytoplasm. Then, PubMed, Web of Science, Science Direct, and Research Gate were used as information sources through the search terms 'chalcone isomerase', 'biosynthesis', 'expression', and their combinations to get the latest and comprehensive information of CHI, mainly from the year 2010 to 2018. More than 300 papers were searched and 116 papers were reviewed in the present work. We summarized the classification of CHI, catalytic reaction mechanism of CHI, and progress of genetic engineering regarding CHI clone, expression, and exogenous stimulator regulation. This paper will lay a foundation for further studies of CHI and other functional genes involved in flavonoids biosynthetic pathway.

Analysis of Phytochemical Constituents and Biological Activity of Taxus Wallichiana Zucc. Dolakha District of Nepal

Phytochemical and biological activities of methanolic extract of Taxus wallichiana Zucc. (Leaf, stem) were carried out. The brine shrimp bioassay showed T. wallichiana is pharmacologically active. The antibacterial potential was studied against one gram positive bacteria (Staphylococcus aureus) and one gram negative bacteria (Escherichia Coli) using Agar Well Diffusion Method. Stem of T. wallichiana showed significant zone of inhibition against gram positive bacteria while the leaf of T. wallichiana did not show significant zone of inhibition against both gram positive and gram negative bacteria. Antioxidant activity was evaluated by 2, 2-diphenyl-1-picryl hydrazyl (DPPH) free radical scavenging activity and FRAP assay. Both assay showed that T. wallichiana leaves has high antioxidant activities.Int. J. Appl. Sci. Biotechnol. Vol 6(2): 110-114