Eudesmane-type sesquiterpene diols directly synthesized by a sesquiterpene cyclase in Tripterygium wilfordii

2018 ◽  
Vol 475 (17) ◽  
pp. 2713-2725 ◽  
Author(s):  
Yu-ru Tong ◽  
Ping Su ◽  
Hong-yu Guan ◽  
Tian-yuan Hu ◽  
Jin-long Chen ◽  
...  
Keyword(s):  
Molecular Model ◽  
Major Product ◽  
Pathway Engineering ◽  
Active Principle ◽  
Shake Flask Culture ◽  
Engineering System ◽  
Tripterygium Wilfordii ◽  
Sesquiterpene Cyclase ◽  
Key Residues

Cryptomeridiol, a typical eudesmane diol, is the active principle component of the antispasmodic Proximol. Although it has been used for many years, the biosynthesis pathway of cryptomeridiol has remained blur. Among terpenoid natural products, terpenoid cyclases are responsible for cyclization and generation of hydrocarbon backbones. The cyclization is mediated by carbocationic cascades and ultimately terminated via deprotonation or nucleophilic capture. Isoprene precursors are, respectively, converted into hydrocarbons or hydroxylated backbones. A sesquiterpene cyclase in Tripterygium wilfordii (TwCS) was determined to directly catalyze (E,E)-farnesyl pyrophosphate (FPP) to unexpected eudesmane diols, primarily cryptomeridiol. The function of TwCS was characterized by a modular pathway engineering system in Saccharomyces cerevisiae. The major product determined by NMR spectroscopy turned out to be cryptomeridiol. This unprecedented production was further investigated in vitro, which verified that TwCS can directly produce eudesmane diols from FPP. Some key residues for TwCS catalysis were screened depending on the molecular model of TwCS and mutagenesis studies. As cryptomeridiol showed a small amount of volatile and medicinal properties, the biosynthesis of cryptomeridiol was reconstructed in S. cerevisiae. Optimized assays including modular pathway engineering and the CRISPR–cas9 system were successfully used to improve the yield of cryptomeridiol in the S. cerevisiae. The best engineered strain TE9 (BY4741 erg9::Δ-200-176 rox1::mut/pYX212-IDI + TwCS/p424-tHMG1) ultimately produced 19.73 mg/l cryptomeridiol in a shake flask culture.

Combination of Microemulsion for Topical Deliverying Tripterygium Wilfordii Multiglycoside

2011 ◽  
Vol 694 ◽  
pp. 881-885
Author(s):  
Xin He ◽  
Xing Hua Zhao ◽  
Wu Qing Ou Yang
Keyword(s):  
Immune Suppression ◽  
Topical Delivery ◽  
Skin Permeability ◽  
Tripterygium Wilfordii ◽  
Delivery Vehicle ◽  
Control Solution ◽  
Suppression Effect ◽  
Low Viscosity ◽  
Spherical Droplets

The purpose of this study was to explore the potential application of the combination of microemulsion as a topical delivery vehicle in enhancing the absorption and efficacy of tripterygium wilfordii multiglycoside (TWM). Various microemulsion formulations were developed and an optimal microemulsion (TWM-ME)，which presented spherical droplets and consisted of RH-40, IPM and water was 27: 3.3: 69.7 by weight. It possessd an average droplet size of 23.6 nm, a low viscosity of (3.56±0.12) mm2•s-1 Zeta electric potential was (–5.35±0.42) mV, refractive index was (1.3617±0.0051) nD20, conductivity was (97.6±3.6) μs/cm. Compared to the control solution, TWM-ME provided better skin permeability in vitro. Moreover, TWM-ME has noticeable anti-inflammatory and immune suppression effect. These results indicate that the combination of microemulsion represents an effective vehicle for topical delivery of TWM.

Synthesis and turnover of proteins and mRNA in germinating wheat embryos

1982 ◽  
Vol 60 (3) ◽  
pp. 389-397 ◽  
Author(s):  
Zbyszko F. Grzelczak ◽  
Mark H. Sattolo ◽  
Linda K. Hanley-Bowdoin ◽  
Theresa D. Kennedy ◽  
Byron G. Lane
Keyword(s):  
Growth Phase ◽  
Time Course ◽  
Phase 1 ◽  
Major Product ◽  
Lag Phase ◽  
Wheat Embryo ◽  
Phase Development ◽  
Wheat Embryos

The most prominent methionine-labeled protein made when cell-free systems are programmed with bulk mRNA from dry wheat embryos has been identified with what may be the most abundant protein in dry wheat embryos. The protein has been brought to purity and has a distinctive amino acid composition, Gly and Glx accounting for almost 40% of the total amino acids. Designated E because of its conspicuous association with early imbibition of dry wheat embryos, the protein and its mRNA are abundant during the "early" phase (0–1 h) of postimbibition development, and easily detected during "lag" phase (1–5 h), but they are almost totally degraded soon after entry into the "growth" phase of development, by about 10 h postimbibition.The most prominent methionine-labeled protein peculiar to the cell-free translational capacity of bulk mRNA from "growth" phase embryos is not detected as a product of in vivo synthesis. Its electrophoretic properties and its time course of emergence, after 5 h postimbibition development, suggest that this major product of cell-free synthesis may be an in vitro counterpart to a prominent methionine-labeled protein made only in vivo, by "growth" phase embryos. Designated G because of its conspicuous association with "growth" phase development, the cell-free product does not comigrate with any prominent dye-stained band in electrophoretic distributions of wheat proteins. The suspected cellular counterpart to G, also, does not comigrate with a prominent dye-stained wheat protein during electrophoresis, and although found in particulate as well as soluble fractions of wheat embryo homogenates it is not concentrated in either nuclei or mitochondria, as isolated.

scholarly journals In VitroAnti-Plasmodial Activity ofTrigonella foenum–graecumL.

2010 ◽  
Vol 7 (4) ◽  
pp. 441-445 ◽  
Author(s):  
M. Palaniswamy ◽  
B. V. Pradeep ◽  
R. Sathya ◽  
J. Angayarkanni
Keyword(s):  
Ethanol Extract ◽  
Traditional Healers ◽  
Active Principle ◽  
Test Plant ◽  
Moderate Activity ◽  
Inhibition Assay ◽  
Phytochemical Screening ◽  
Solvent Systems ◽  
Fenugreek Leaves

Developing countries, where malaria is one of the most prevalent diseases, still rely on traditional medicine as a source for the treatment of this disease. For the present study,Trigonella foenum-graecumL. (fenugreek) were collected from Coimbatore, Tamilnadu, India. The test plant has been used in India by traditional healers for the treatment of fever as well as other diseases. The active principle was extracted out in different solvent systems to assess the anti-plasmodial potential, with an aim that they can further be utilized to formulate drugs.In vitroanti-plasmodial assay of the extracted fractions of fenugreek leaves was carried out using laboratory adapted chloroquine sensitive and resistantPlasmodium falciparumisolates. Schizont maturation inhibition assay was adopted to analyze the potential of the extracts. Ethanol extract (50%) seemed to possess profound anti-plasmodial activity with IC50value of 8.75 ± 0.35 µg ml−1and 10.25 ± 0.35 µg ml−1against chloroquine sensitive and resistantP. falciparumisolates, respectively. Among the investigated six fractions of the plant extracts, two were found to have significant anti-plasmodial activity with IC50values <10 µg ml−1, namely ethanol and butanol extracts. Two extracts chloroform and ethyl acetate showed moderate activity with IC50values ranging from 10 to 20 µg ml−1, and the other two extracts, hexane and water appeared to be inactive with IC50values >85 µg ml−1. In addition, preliminary phytochemical screening of the various extracts indicated the presence of alkaloids, saponin, tannin like phenolic compounds, flavonoids and steroids.

scholarly journals Synthesis and In Vitro Screening of Novel Heterocyclic β-d-Gluco- and β-d-Galactoconjugates as Butyrylcholinesterase Inhibitors

Molecules ◽  
2019 ◽  
Vol 24 (15) ◽  
pp. 2833
Author(s):  
Krešimir Baumann ◽  
Lorena Kordić ◽  
Marko Močibob ◽  
Goran Šinko ◽  
Srđanka Tomić
Keyword(s):  
Active Site ◽  
Kinetic Studies ◽  
Inhibitory Effect ◽  
In Vitro Screening ◽  
Sugar Moiety ◽  
Brain Barrier Permeability ◽  
The Central Nervous System ◽  
Key Residues ◽  
Micromolar Range

The development of selective butyrylcholinesterase (BChE) inhibitors may improve the treatment of Alzheimer’s disease by increasing lower synaptic levels of the neurotransmitter acetylcholine, which is hydrolysed by acetylcholinesterase, as well as by overexpressed BChE. An increase in the synaptic levels of acetylcholine leads to normal cholinergic neurotransmission and improved cognitive functions. A series of 14 novel heterocyclic β-d-gluco- and β-d-galactoconjugates were designed and screened for inhibitory activity against BChE. In the kinetic studies, 4 out of 14 compounds showed an inhibitory effect towards BChE, with benzimidazolium and 1-benzylbenzimidazolium substituted β-d-gluco- and β-d-galacto-derivatives in a 10–50 micromolar range. The analysis performed by molecular modelling indicated key residues of the BChE active site, which contributed to a higher affinity toward the selected compounds. Sugar moiety in the inhibitor should enable better blood–brain barrier permeability, and thus increase bioavailability in the central nervous system of these compounds.

scholarly journals Preliminary cytotoxic activity of different extracts of Averrhoa bilimbi (fruits)

2013 ◽  
Vol 2 (3) ◽  
pp. 83-84 ◽  
Author(s):  
Md. Ramjan Ali ◽  
Marjan Hossain ◽  
Jannatul Ferdous Runa ◽  
Md. Hasanuzzaman
Keyword(s):  
Carbon Tetrachloride ◽  
Room Temperature ◽  
Methanolic Extract ◽  
Pharmaceutical Journal ◽  
Artemia Salina ◽  
Active Principle ◽  
Cytotoxic Potential ◽  
Potential Source ◽  
Soluble Fractions

The crude methanolic extract of Averrhoa bilimbi Linn. (Oxalidiaceae) fruits and its different fractions have been investigated for the evaluation of in vitro cytotoxic potential. The dried and powder fruits were extracted with methanol at room temperature and the concentrated methanolic extract was fractionated by the modified Kupchan partitioning method to provide pet-ether, carbon tetrachloride, chloroform and aqueous soluble fractions. Brine shrimp (Artemia salina) lethality bioassay was used to investigate the cytotoxic potential of A. bilimbi. Compared to vincristine sulfate (with LC50 of 0.839 µg/ml) methanolic extract, carbon tetrachloride and pet-ether soluble fractions demonstrated a significant cytotoxic potential (having LC50 of 0.005µg/ml, 1.198µg/ml and 0.781µg/ml, respectively). The LC50 values of chloroform and aqueous soluble fractions were 5.691µg/ml and 6.123µg/ml, respectively. This study reveals that A. bilimbi possesses effective cytotoxic properties and hence can be a potential source for the isolation of active principle(s) for cancer therapy.DOI: http://dx.doi.org/10.3329/icpj.v2i3.13634 International Current Pharmaceutical Journal, February 2013, 2(3): 83-84 

scholarly journals Import of the carboxy-terminal portion of acyl-CoA oxidase into peroxisomes of Candida tropicalis.

1987 ◽  
Vol 105 (1) ◽  
pp. 247-250 ◽  
Author(s):  
G M Small ◽  
P B Lazarow
Keyword(s):  
Cdna Clone ◽  
Candida Tropicalis ◽  
In Vitro Transcription ◽  
Major Product ◽  
Sufficient Information ◽  
Terminal Portion ◽  
Peroxisomal Protein ◽  
Acyl Coa Oxidase ◽  
Carboxy Terminal

We report the sequence of a cDNA clone that codes for the carboxy-terminal portion of the peroxisomal protein, acyl-CoA oxidase, from the yeast, Candida tropicalis. This is a newly identified acyl-CoA oxidase sequence, most likely a second allele of POX4. The cDNA clone was expressed by in vitro transcription followed by translation. The major product, a 43-kD protein, associated with isolated peroxisomes in an in vitro import assay. More than half of the peroxisome-associated protein was protected from added protease, implying that it was internalized within the organelle. These findings indicate that there is sufficient information in the carboxy-terminal portion of the protein to target it to peroxisomes.

scholarly journals Feiba Immuno: A Preparation with Factor Eight Inhibitor Bypassing Activity

1977 ◽  
Author(s):  
F. Elsinger
Keyword(s):  
Factor Viii ◽  
Factor Xa ◽  
Coagulation Factors ◽  
Active Principle ◽  
Factor Viii Inhibitor ◽  
Factor V ◽  
In Vitro Experiments ◽  
Factor X ◽  
Viii Inhibitor

FEIBA IMMUNO is a preparation in which a new activity is generated capable of bypassing factor VIII. The preparation which is used to treat patients with inhibitors (especially inhibitors to factor VIII) is standardized in FEIBA units, i.e. in terms of its in vitro capacity to shorten the activated PTT of a factor VIII inhibitor plasma.It could be concluded from different in vitro experiments that none of the classic’ activated coagulation factors is responsible for the factor VIII bypassing reaction; FEIB-activity seems to be correlated to a new complex of coagulation factors.To get an answer to the question which coagulation factors are essential for FEIB-activity, we tried to generate this activity from different deficient plasmas; from these experiments the following conclusions could be drawn:, the presence of at least factors VII, IX, and X is essential for the generation of the molecular species responsible for factor VIII as well as factor X bypassing activity, but factor V is not bypassed. This activity is not factor Xa itself. Factors VIII and V are not necessary for the generation of this active principle, but factor V is finally needed for its bypassing action.

scholarly journals Regulation of Carbon and Electron Flow inClostridium butyricum VPI 3266 Grown on Glucose-Glycerol Mixtures

2001 ◽  
Vol 183 (5) ◽  
pp. 1748-1754 ◽  
Author(s):  
Sylvie Saint-Amans ◽  
Laurence Girbal ◽  
Jose Andrade ◽  
Kerstin Ahrens ◽  
Philippe Soucaille
Keyword(s):  
Electron Flow ◽  
Sharp Decrease ◽  
Major Product ◽  
Glycerol Dehydrogenase ◽  
Hydrogenase Activity ◽  
Specific Rate ◽  
Oxygen Sensitive ◽  
High Level

ABSTRACT The metabolism of Clostridium butyricum was manipulated at pH 6.5 and in phosphate-limited chemostat culture by changing the overall degree of reduction of the substrate using mixtures of glucose and glycerol. Cultures grown on glucose alone produced only acids (acetate, butyrate, and lactate) and a high level of hydrogen. In contrast, when glycerol was metabolized, 1,3-propanediol became the major product, the specific rate of acid formation decreased, and a low level of hydrogen was observed. Glycerol consumption was associated with the induction of (i) a glycerol dehydrogenase and a dihydroxyacetone kinase feeding glycerol into the central metabolism and (ii) an oxygen-sensitive glycerol dehydratase and an NAD-dependent 1,3-propanediol dehydrogenase involved in propanediol formation. The redirection of the electron flow from hydrogen to NADH formation was associated with a sharp decrease in the in vitro hydrogenase activity and the acetyl coenzyme A (CoA)/free CoA ratio that allows the NADH-ferredoxin oxidoreductase bidirectional enzyme to operate so as to reduce NAD in this culture. The decrease in acetate and butyrate formation was not explained by changes in the concentration of phosphotransacylases and acetate and butyrate kinases but by changes in in vivo substrate concentrations, as reflected by the sharp decrease in the acetyl-CoA/free CoA and butyryl-CoA/free CoA ratios and the sharp increase in the ATP/ADP ratio in the culture grown with glucose and glycerol compared with that in the culture grown with glucose alone. As previously reported for Clostridium acetobutylicum (L. Girbal, I. Vasconcelos, and P. Soucaille, J. Bacteriol. 176:6146–6147, 1994), the transmembrane pH of C. butyricum is inverted (more acidic inside) when the in vivo activity of hydrogenase is decreased (cultures grown on glucose-glycerol mixture). For both cultures, the stoichiometry of the H+ ATPase was shown to remain constant and equal to 3 protons exported per molecule of ATP consumed.

scholarly journals Evaluation of Antileishmanial Activity of Selected Brazilian Plants and Identification of the Active Principles

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Valdir Cechinel Filho ◽  
Christiane Meyre-Silva ◽  
Rivaldo Niero ◽  
Luisa Nathália Bolda Mariano ◽  
Fabiana Gomes do Nascimento ◽  
...  
Keyword(s):  
Active Principle ◽  
Chloroform Fraction ◽  
Methanolic Extracts ◽  
Ethanolic Extracts ◽  
Stems And Leaves ◽  
Isolated Compound ◽  
Antiparasitic Agents ◽  
Brazilian Medicinal Plants ◽  
Solanum Sisymbriifolium

This study evaluated extracts, fractions, and isolated compounds from some selected Brazilian medicinal plants against strains of promastigotes ofLeishmania amazonensisandL. brasiliensis in vitro. The cell viability was determined, comparing the results with reference standards. The dichloromethane fractions of the roots, stems, and leaves ofAllamanda schottiishowed IC50values between 14.0 and 2.0 μg/mL. Plumericin was the main active compound, with IC50of 0.3 and 0.04 μg/mL against the two species ofLeishmaniaanalyzed. The hexane extract ofEugenia umbelliflorafruits showed IC50of 14.3 and 5.7 μg/mL againstL. amazonensisandL. brasiliensis, respectively. The methanolic extracts of the seeds ofGarcinia achachairuand guttiferone A presented IC50values of 35.9 and 10.4 μg/mL, againstL. amazonensis, respectively. The ethanolic extracts of the stem barks ofRapanea ferrugineaand the isolated compound, myrsinoic acid B, presented activity againstL. brasiliensiswith IC50of 24.1 and 6.1 μg/mL. Chloroform fraction ofSolanum sisymbriifoliumexhibited IC50of 33.8 and 20.5 μg/mL, and cilistol A was the main active principle, with IC50of 6.6 and 3.1 μg/mL againstL. amazonensisandL. brasiliensis, respectively. It is concluded that the analyzed plants are promising as new and effective antiparasitic agents.

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