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Gelsemium elegans is a traditional Chinese herb of medicinal importance, with indole terpene alkaloids as its main active components. To study the expression of the most suitable housekeeping reference genes in G. elegans, the root bark, stem segments, leaves and inflorescences of four different parts of G. elegans were used as materials in this study. The expression stability of 10 candidate housekeeping reference genes (18S, GAPDH, Actin, TUA, TUB, SAND, EF-1α, UBC, UBQ, and cdc25) was assessed through real-time fluorescence quantitative PCR, GeNorm, NormFinder, BestKeeper, ΔCT, and RefFinder. The results showed that EF-1α was stably expressed in all four parts of G. elegans and was the most suitable housekeeping gene. Based on the coexpression pattern of genome, full-length transcriptome and metabolome, the key candidate targets of 18 related genes (AS, AnPRT, PRAI, IGPS, TSA, TSB, TDC, GES, G8H, 8-HGO, IS, 7-DLS, 7-DLGT, 7-DLH, LAMT, SLS, STR, and SGD) involved in the Gelsemium alkaloid biosynthesis were obtained. The expression of 18 related enzyme genes were analyzed by qRT-PCR using the housekeeping gene EF-1α as a reference. The results showed that these genes' expression and gelsenicine content trends were correlated and were likely to be involved in the biosynthesis of the Gelsemium alkaloid, gelsenicine.
Subject(s)
Genes, Essential , Gelsemium/genetics , Peptide Elongation Factor 1/genetics , Transcriptome , Gene Expression Profiling/methods , Alkaloids , Real-Time Polymerase Chain Reaction/methods , Reference StandardsABSTRACT
Three new indole alkaloids, flueindolines A-C (1-3), along with nine known alkaloids (4-12), were isolated from the fruits of Flueggea virosa (Roxb. ex Willd.) Voigt. Compounds 1 and 2 are two new fused tricyclic indole alkaloids possessing an unusual pyrido[1, 2-a]indole framework, and 3 presents a rare spiro (pyrrolizidinyl-oxindole) backbone. Their structures with absolute configurations were elucidated by means of comprehensive spectroscopic analysis, chemical calculation, as well as X-ray crystallography. Chiral resolution and absolute configuration determination of the known compounds 4, 10, and 11 were reported for the first time. The hypothetical biogenetical pathways of 1-3 were herein also proposed.
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Rauvolfia serpentine is a traditional Indian medicine and now the genus Rauvolfia becomes a commonly used south medicine in China. Rauvolfia is mainly used to treat hypertension, mental illness, cardiovascular disorders, cancer, and other diseases. The paper overviewed the main species and distribution, pharmacological properties, and the monoterpenoid indole alkaloids biosynthetic pathway of Rauvolfia in China. The paper aims to provide reference for research and clinical application in plants of the genus Rauvolfia.
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Objective: To study the chemical constituents from the stems of Uncaria scandens. Methods: The compounds were isolated and purified by silica gel column chromatography, Sephadex LH-20, RP18, and HPLC. The structures were identified by spectroscopic methods and physicochemical properties. The contents of rhynchophylline and isorhynchophylline were detected by HPLC method. Results: Seventeen compounds were isolated and identified from 95% ethanol extract of the stems of U. scandens and characterized as 5α-carboxystrictosidine (1), 3β-isodihydrocadambine (2), strictosamide (3), 3α-dihydrocadambine (4), mitraphylline (5), uncarine F (6), 3β-isodihydrocadambine 4-oxide (7), isodihydrocdambine (8), nauclefine (9), cadambine (10), calyxamine B (11), methyl chlorogenate (12), 3,4,5-trimethoxyphenol (13), α-tocopherylquinone (14), quinovic acid (15), methyl caffeate (16), and hyperoside (17). The contents of rhynchophylline and isorhynchophylline from U. scandens were 0.72 and 0.38 μg/g. Conclusion: Compounds 7-9 and 11-14 are isolated from Uncaria genus for the first time. Compounds 1-6 and 16, 17 are isolated from U.scandens for the first time. The contents of rhynchophylline and isorhynchophylline is low and the reasonable use of them as U. scandens in some areas needs to be further verified.
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Objective:To obtain the information of alkaloids in Evodia rutaecarpa by HPLC-Q-TOF-MS/MS. Method:Inter Sustain-C18 column (4.6 mm×250 mm,5 μm) was used with 0.2% formic acid water-acetonitrile as the mobile phase for gradient elution. The column temperature was 25℃,the volume flow rate was 1.0 mL·min-1,and the sample volume is 5 μL. The detection wavelength was 245 nm,and the chromatographic effluent was detected and analyzed by using both positive and negative ions. Result:According to molecular ion peaks and secondary mass spectrometry characteristic fragment ions,as well as the mass spectrometry information of reference substances and relevant literature reports,more than 40 major peaks were analyzed,and 21 alkaloids were identified from the methanol extract of E. rutaecarpa, including 10 kinds of indole alkaloids,10 kinds of quinolone alkaloids,and 1 kind of ephedrine. Main types of alkaloids in E. rutaecarpa were basically clarified. And the research found that the alkaloids have a good response mainly in the positive mode. Conclusion:Based on HPLC-Q-TOF-MS/MS technology, high-performance liquid chromatography (HPLC) separation,mass spectrometry determination of molecular mass,pyrolysis data,literature analysis and retrieval were performed to quickly,accurately and comprehensively identify alkaloids in E. rutaecarpa, so as to provide a scientific basis for the further extraction and separation of the chemical constituents of E. rutaecarpa.
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Uncaria genus( Rubiaceae) contains 34 species all over the world,of which 11 species and one variant are present in China. Five species,namely U. rhynchophylla,U. macrophylla,U. hirsuta,U. sinensis and U. sessilifructus,are documented in Chinese Pharmacopoeia as the raw materials of Uncariae Ramulus Cum Uncis. Indole alkaloids are the characteristic constituents of Uncaria plants,in addition to triterpenes,lignans and flavones. This paper reviews the progress of indole alkaloids and their distribution within the five Uncaria species documented in Chinese Pharmacopoeia for better understanding the active constituents of Uncariae Ramulus Cum Uncis.
Subject(s)
Alkaloids , China , Drugs, Chinese Herbal , Indole Alkaloids , Medicine, Chinese Traditional , UncariaABSTRACT
Five alkaloids were isolated from a decoction of Uncaria rhynchophylla by a combination of various chromatographic techniques, including macroporous adsorbent resin, MCI resin, silica gel, Sephadex LH-20, and reversed phase HPLC. Their structures were characterized by comprehensive analyses of spectroscopic data as monoterpene indole alkaloids (+)-(7R)-3-oxo-7-hydroxy-3,7-seco-dihydrorhynchohylline (1), (+)-(7S)-3-oxo-7-hydroxy-3,7-seco-dihydrorhyncho-hylline (2), (+)-(7R)-3-oxo-7-hydroxy-3,7-seco-rhynchohylline (3) and (+)-(7S)-3-oxo-7-hydroxy-3,7-seco-rhynchohylline (4), and a β-carboline alkaloid 1,2,3,4-tetrahydro-1-oxo-β-carboline (5). Among them, 1 and 2 are new compounds, 3 and 4 are new natural products that were semi-synthesized from isorhynchohylline with incorrect specific rotations, and 5 is isolated for the first time from the genus Uncaria.
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Objective To study the chemical constituents from the stems of Uncaria lancifolia. Methods The isolation and purification were carried out by silica gel column chromatography, RP18, Sephadex LH-20, and preparative HPLC. The structures of the isolated compounds were elucidated by physical and chemical properties, and spectroscopic methods. Results Eighteen compounds were isolated and identified from 95% ethanol extract from the stems of U. lancifolia and characterized as uncarine A (1), uncarine E (2), isomitraphylline (3), tetrahydroalstonine (4), strictosidine (5), cadambine (6), glabratine (7), strictosamide (8), (13R)-hydroxy- octodeca-(9Z,11E,15Z)-trien-oic acid (9), (6S,9R)-roseoside (10), periplanetin (11), integracin A (12), integracin B (13), 6β,19α- dihydroxyurs-3-oxours-12-en-28-oic aicd (14), ursolic acid (15), oleanic acid (16), β-sitosterol (17), and β-daucosterol (18). Conclusion This is the first report for the chemical constituents from U. lancifolia. All compounds are obtained from this plant for the first time, and compounds 9-13 are isolated from Uncaria genus for the first time. Compounds 1-8 are monoterpene indole alkaloids, which are characteristic constituents in Uncaria genus.
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Bufonis Venenum, as a precious Chinese materia medica, has complex chemical constituents and has been widely used in clinical treatment with significant effects. The chemical constituents in Bufonis Venenum mainly included bufadienolides, indole alkaloids, and steroids, etc. Modern pharmacological research has demonstrated its antitumor, cardiac, anti-inflammatory, and narcotic analgesic, etc. This paper reviewed the chemical constituents, pharmacological activity, and artificial synthesis of Bufonis Venenum, providing theoretical reference for material basic research, pharmacodynamic mechanism interpretation, development, and utilization.
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The chemical constituents of the fruits of Chaenomeles sinensis (Thouin) Koehne were investigated using chromatographic methods, including Diaion HP-20, Toyopearl HW-40, MCI Gel CHP-20, ODS, Silica gel chromatography and semi-preparative-HPLC. Three compounds were isolated and their structures were elucidated with spectral data and physicochemical properties, which were identified as chaenomeles alkaloid A (1), ginsenine (2) and 1,2,3,4-tetrahydro-1-methyl-β-carboline-3-car-boxylic acid (3). Among those, compound 1 is a new alkaloid, compound 2 and 3 were isolated from this plant for the first time. To investigate the protective effect of compounds 1-3 on Rat adrenal pheochromocytoma (PC-12) injury induced by the β-amyloid protein (Aβ25-35). The results show that compounds 2 and 3 have a significant protective effect on the PC12 cells exposed to Aβ25-35.
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Objective To establish an high perfermance liquid chromatography method for determination of five indole alkaloids in Rauvolfia.Methods The Diamonsil C18 column was used at 25 ℃ with methanol-water (gradient elution) as the mobile phase,the flow rate was 1.0 mL·min-1,the detective wavelength was 280 nm,and the injection volume was 5 μL.Results In the range of 1.56-200 μg· mL-1,the correlation coefficients of regression equations of sarpagine,yohimbine,ajmaline,ajmalicine,reserpine were higher than 0.999 0.The average recovery (n =9) of five indole alkaloids was between 95.0％-105.0％.Conclusion This method is simple with good accuracy and repeatability.It can be used for quality control of Rauvolfia.
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Objective: To study the alkaloids from Ervatamia hainanensis. Methods: The alkaloids were isolated and purified by silica gel, MCI, Sephadex LH-20 column chromatography, and semi-preparative HPLC, and their structures were elucidated by physical and spectroscopic analysis. Results: Twelve alkaloids were obtained and identified as coronaridine (1), 19-epi-heyneanine (2), 9,10-dimethoxycoronaridine (3), vobasine (4), vobasine N(4)-oxide (5), 3-oxo-19-epi-heyneanine (6), strictamine (7), deacetylakuammiline (8), pandine (9), rhazicine (10), rhazicine N(4)-oxide (11), and rhazimine (12). Conclusion: Compounds 8 and 10-12 are isolated from the genus Ervatamia Stapf for the first time, while compounds 3 and 5-7 are firstly obtained from E. hainanensis.
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Toad venom (Chansu) is prepared from the dried secretion of Bufo bufo gargarizans or B. melanostictus. It is not only one of the famous and expensive traditional Chinese medicines(TCMs) from animal origin, but also one of 28 kinds of toxic TCMs to be required for special management issued by the State Council of the People's Republic of China. Chansu contains the rich bufadienolides and indole alkaloids, and displays various bioactivity including cardiotonic, anti-tumor, analgesia, and local anesthesia. Based on the published references in the recent years, the advance on the identification of adulterants and quality evaluation as well as the influence factors on the quality of toad venom was summarized to improve the quality standards and promote the level of quality control of toad venom and its preparations.
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Rauwolfia species (Apocynaceae) are medicinal plants well known worldwide due to its potent bioactive monoterpene indole alkaloids (MIAs) such as reserpine, ajmalicine, ajmaline, serpentine and yohimbine. Reserpine, ajmalicine and ajmaline are powerful antihypertensive, tranquilizing agents used in hypertension. Yohimbine is an aphrodisiac used in dietary supplements. As there is no report on the comparative and comprehensive phytochemical investigation of the roots of Rauwolfia species, we have developed an efficient and reliable liquid chromatography-tandem mass spectrometry (LC–MS/MS) method for ethanolic root extract of Rauwolfia species to elucidate the fragmentation pathways for dereplication of bioactive MIAs using high-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry (HPLC–ESI–QTOF–MS/MS) in positive ion mode. We identified and established diagnostic fragment ions and fragmentation pathways using reserpine, ajmalicine, ajmaline, serpentine and yohimbine. The MS/MS spectra of reserpine, ajmalicine, and ajmaline showed C-ring-cleavage whereas E-ring cleavage was observed in serpentine via Retro Diels Alder (RDA). A total of 47 bioactive MIAs were identified and characterized on the basis of their molecular formula, exact mass measurements and MS/MS analysis. Reserpine, ajmalicine, ajmaline, serpentine and yohimbine were unambiguously identified by comparison with their authentic standards and other 42 MIAs were tentatively identified and characterized from the roots of Rauwolfia hookeri, Rauwolfia micrantha, Rauwolfia serpentina, Rauwolfia verticillata, Rauwolfia tetraphylla and Rauwolfia vomitoria. Application of LC–MS followed by principal component analysis (PCA) has been successfully used to discriminate among six Rauwolfia species.
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Indole alkaloid family has been the biggest so far among the various alkaloids,which contains pharmaceutical and effective constituents of various plants featuring diverse biological activities.Thanks to the development of metabonomics,to reveal the biosynthetic pathway of active components for the molecular mechanism of indole alkaloids and the regulation research of plant metabolism present a growing importance and significantly direct the researches of improving biological production.This paper reviewed the biosynthetic pathways of some indole alkaloids in accordance with the structure classification of indole alkaloids to lay a foundation for the further studies on the biosynthetic pathways of indole alkaloids and provide a reference for the biosynthetic pathways of other indole alkaloids.
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Catharanthus roseus can produce a variety of terpenoid indole alkaloids (TIA), most of which exhibit strong pharmacological activities. Hence, biosynthesis and regulation of TIA have received recent attention. 3α (S)-strictosidine is an important node in TIA biosynthesis, which is a condensation product of secologanin and tryptamine. The former is produced in iridoid pathway, and the latter is produced in indole pathway. Vindoline and catharanthine, which are produced respectively by 3α (S)-strictosidine via multi-step enzymatic reaction, can form α-3, 4-anhydrovinblastine by the condensation reaction. Then, vinblastine and vincristine are generated from α-3, 4-anhydrovinblastine. Many transcription factors are involved in the regulation of TIA synthesis, such as AP2/ERF and WRKY. Illumination of biosynthetic pathway has laid a foundation for the study of synthetic biology. Today, 3α (S)-strictosidine and vindoline have been synthesized in heterologous hosts Saccharomyces cerevisiae.Research about synthetic biology and the regulation mechanisms will provide a guidance for the production and development of TIA drugs in C. roseus.
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This study aimed to provide guidance for the heterogenous gene expression, gene prediction and species evolution by analyzing codon usage bias of Catharanthus roseus.The codon composition and usage bias of 30 437 high-confidence coding sequences from C.roseus were analyzed and the proportion of rare codons of Escherichia coli and Saccharomyces cerevisiae in 25 genes involved in the biosynthesis of terpenoid indole alkaloids (TIAs) in C.roseus were calculated.The results showed that the average GC content of the genes was 42.47%; the average GC content of the third bases in codon was 35.89%.The relative synonymous codon usage (RSCU) of 28 codons were greater than 1 and 26 of them ended with A or T.The above 25 genes involved in TIA biosynthesis contained much more rare condons of E.coli than that of S.cerevisiae.It was concluded that C.roseus mainly prefered the codons ending with A or T and the rule of codon usage was more different to E.coli than S.cerevisiae.Thus, S.cerevisiae may be more suitable host for heterologous expression of these genes.
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AbstractZ-Vallesiachotamine is a monoterpene indole alkaloid that has a β-N-acrylate group in its structure. This class of compounds has already been described in different Psychotriaspecies. Our research group observed that E/Z-vallesiachotamine exhibits a multifunctional feature, being able to inhibit targets related to neurodegeneration, such as monoamine oxidase A, sirtuins 1 and 2, and butyrylcholinesterase enzymes. Aiming at better characterizing the multifunctional profile of this compound, its effect on cathecol-O-methyltransferase activity was investigated. The cathecol-O-methyltransferase activity was evaluated in vitro by a fluorescence-based method, using S-(5′-adenosyl)-l-methionine as methyl donor and aesculetin as substrate. The assay optimization was performed varying the concentrations of methyl donor (S-(5′-adenosyl)-l-methionine) and enzyme. It was observed that the highest concentrations of both factors (2.25 U of the enzyme and 100 µM of S-(5′-adenosyl)-l-methionine) afforded the more reproducible results. The in vitro assay demonstrated that Z-vallesiachotamine was able to inhibit the cathecol-O-methyltransferase activity with an IC50 close to 200 µM. Molecular docking studies indicated that Z-vallesiachotamine can bind the catechol pocket of catechol-O-methyltransferase enzyme. The present work demonstrated for the first time the inhibitory properties of Z-vallesiachotamine on cathecol-O-methyltransferase enzyme, affording additional evidence regarding its multifunctional effects in targets related to neurodegenerative diseases.
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OBJECTIVE: To investigate the application of near infrared spectroscopy in rapid and quantitative determination of multiple quality indicators in concentration process of Bufo bufo gargarizans Cantor extract. METHODS: NIR transmission spectra of concentrated solutions were collected in rectangular quartz cuvette with different optical pathlengths (2 and 5 mm). Calibration models of density, water content and indole alkaloids were developed by partial least squares regression (PLSR). The established models were applied to predict the unknown samples to test the performance of the models. RESULTS: The correlation coefficients of the density, water content and indole alka. The residual predictive deviations (RPD) were all above 3, and the relative standard errors of predictions(RSEP) were less than 10%, indicating the satisfactory model performance and predictive ability. Moreover, better calibration results were obtained with an optical path length of 2 mm for density and water content, and 5 mm for indole alkaloids. The results revealed that the optical path length has influence on the NIR analysis results, and the appropriate path length for the NIR analysis should be determined by comparison analysis. CONCLUSION: NIR spectroscopy technique provides a novel efficient and environmental friendly approach for the fast simultaneous determination of three key quality indicators (density, water content and indole alkaloids) in the concentration process of Bufo bufo gargarizans Cantor extract. Copyright 2012 by the Chinese Pharmaceutical Association.
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O presente trabalho descreve o isolamento e a identificação de nove alcaloides indólicos monoterpênicos das cascas das raízes e folhas de Tabernaemontana salzmannii (Apocynaceae). As estruturas dos alcaloides foram identificadas através de métodos espectroscópicos uni (RMN ¹H, 13C, APT) e bidimensionais (¹H-¹H-COSY, ¹H-¹H-NOESY, HMQC e HMBC) e espectrometria de massas (EM), além da comparação com dados de literatura. Um screening in vitro da atividade antileucêmica foi feito com os alcaloides majoritários isolados. Dentre os nove alcaloides isolados, a isovoacangina e voacangina mostraram-se capazes de induzir morte celular por apoptose em células leucêmicas humanas THP-1.
This work describes the isolation and structural determination of nine monoterpenic indole alkaloids from the roots bark and leaves of Tabernaemontana salzmannii. The alkaloids were identified by spectroscopic methods uni (NMR ¹H, 13C, APT) and two-dimensional (¹H-¹H-COSY, ¹H-¹H-NOESY, HMQC and HMBC) and mass spectra besides comparison with literature data. An in vitro screening was done with the isolated major alkaloids. Among the nine alkaloids isolated, isovoacangine and voacangine alkaloids were able to induce apoptosis cell death in human leukemic cells line THP-1.