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1.
Acta Pharmaceutica Sinica ; (12): 3353-3361, 2021.
Article in Chinese | WPRIM | ID: wpr-906835

ABSTRACT

italic>Aconitum pendulum is a Tibetan medicine that is rich in bioactive compounds such as aconitine-type C19-diterpenoid alkaloids. To investigate the key enzymes in the aconitine biosynthesis pathway, roots, leaves and flowers of Aconitum pendulum were subjected to a high-throughput transcriptomic sequencing analysis by Illumina HiSeqTM2000. Trinity de novo assembly yielded 47 264 unigenes with an average length of 1 140 bp and N50 of 1 678 bp, of which 30 231 unigenes (63.96%) were annotated. In the KEGG database, 542 unigenes were implicated in 17 secondary metabolic pathways; the analysis showed that 44 genes encoded 20 key enzymes in the diterpene skeleton of aconitine biosynthesis and 12 BAHD acyltransferase genes were related to the acetylation modification, with differential expression among three organs. For example, ApTPS8 was the only committed enzyme in the upstream aconitine biosynthetic pathway. The high expression level of ApTPS8 in root indicated that it is the main tissue for the production of precursors of diterpene alkaloids. Consistent with the accumulation of aconitine, we propose that ApBAHD1/2/8 is involved in the biosynthesis of 2-hydroxyaconitine, dehydrated 14-benzoylaconitine, 8-O-methyl-14-benzoylaconine, benzoyldeoxyaconitine and benzoylaconitine, and ApBAHD10 is involved in the biosynthesis of acontine, lucidusculine, 14-O-acetylneoline and 14-O-acetylvirescenin. Comparative transcriptome analysis of A. pendulum and A. carmichaeli indicates significant gene loss in the family of diterpene synthases and acyltransferases in A. pendulum, which is in accordance with the significantly fewer type and quantity of aconitine compounds in this species. Therefore, A. pendulum has proved to be an ideal material for the study of the aconitine biosynthesis pathway. This work provides basic scientific data for further study of aconitine biosynthesis, the discussion of molecular mechanisms of toxicity, and the synthesis of genuine medicinal materials.

2.
Article in Chinese | WPRIM | ID: wpr-905076

ABSTRACT

Objective:To conduct quality evaluation of Ginkgo Folium preparations by analyzing the national evaluation sampling test results, analyze the quality differences, and put forward suggestions for the improvement of quality standards and market supervision. Method:The contents of total flavonol glycosides and terpene lactones in Ginkgo Folium tablets and Ginkgo Folium capsules were determined according to the methods of determination in the 2015 edition of Chinese Pharmacopoeia (the first volume), and the contents of free flavonoids (quercetin, kaempferide and isorhamnetin) and sophoricoside in Ginkgo Folium preparations were determined according to related supplementary testing method of Ginkgo Folium tablets and Ginkgo Folium capsules issued by National Medical Products Administration. The quality differences of Ginkgo Folium preparations from different batches and different manufacturers were compared according to the contents of total flavonol glycosides, terpene lactones, free flavonoids and sophoricoside in 328 batches of Ginkgo Folium tablets and Ginkgo Folium capsules manufactured by 48 enterprises. Result:Quality of 328 batches of Ginkgo Folium tablets and Ginkgo Folium capsules was in accordance with the standard, but the contents of terpene lactones and total flavonol glycosides were all distributed in a wide range, and the quality of samples varied greatly among different enterprises. Conclusion:It is recommended that each enterprise should optimize the production process and strictly control the raw materials to ensure the consistency between different batches of samples.

3.
Braz. j. biol ; 80(4): 790-797, Oct.-Dec. 2020. tab, graf
Article in English | LILACS | ID: biblio-1142527

ABSTRACT

Abstract Ginkgo biloba is a traditional Chinese herbal medicine containing multiple components that contribute to its notable bioactivities. Variations of seasonal, meteorological and planting soil on the phytochemicals contents in G. biloba leaves due to the effects of growth meteorological and soil parameters were investigated in this study. The leaves of G. biloba were collected from different months and place in Zhejiang province, the contents of flavones (quercetin, kaempferol and isorhamnetin) and terpene lactones (bilobalide, ginkgolides A, B and C) were quantified by high performance liquid chromatography (HPLC) and the evaporative light scattering detector (ELSD) method. The established methods were validated with good linearity, precision, repeatability, stability, and recovery. Comprehensive analysis suggested the proper harvest time for G. biloba was in October of Zhejiang province. The result of correlation analysis with meteorological factors shows that the temperature and precipitation have non-significant effect on the main components of G. biloba. In addition, the type and content (Mn and Zn) of the soil showed significantly effect on the content of flavonoids and terpene lactones. This study enriched the knowledge on the development and utilization value of the G. biloba leaves and was useful for determining the optimal harvest time and growing condition.


Resumo Ginkgo biloba é um fitoterápico tradicional da medicina chinesa que contém vários componentes que contribuem para suas notáveis bioatividades. Variações sazonais, meteorológicos e de plantio do solo sobre os teores fitoquímicos em folhas de G. biloba, devido aos efeitos do crescimento meteorológico e parâmetros do solo, foram investigadas neste estudo. As folhas de G. biloba foram coletadas em diferentes meses e na província de Zhejiang, os teores de flavonas (quercetina, kaempferol e isorhamnetina) e lactonas terpênicas (bilobalida, ginkgolídeos A, B e C) foram quantificados por Cromatografia Líquida de Alta Eficiência (CLAE) e pelo método do detector de dispersão da luz por evaporação. Os métodos estabelecidos foram validados com boa linearidade, precisão, repetibilidade, estabilidade e recuperação. Uma análise abrangente sugeriu que o tempo de colheita adequado para G. biloba foi em outubro na província de Zhejiang. O resultado da análise de correlação com fatores meteorológicos mostra que a temperatura e a precipitação não têm efeito significativo sobre os principais componentes de G. biloba. Além disso, o tipo e composição (Mn e Zn) do solo apresentaram efeito significativo sobre o teor de flavonoides e lactonas terpênicas. Este estudo enriqueceu o conhecimento sobre o valor de desenvolvimento e utilização das folhas de G. biloba e foi útil para determinar o melhor tempo de colheita e condição de crescimento.


Subject(s)
Soil , Ginkgo biloba , Seasons , Plant Extracts , Chromatography, High Pressure Liquid , Plant Leaves
4.
Article in Chinese | WPRIM | ID: wpr-846180

ABSTRACT

Objective: To clone and analyze the cDNA sequence of 3-Hydroxy-3-methylglutaryl coenzyme-a reductase (HMGR1) of Cornus officinalis. Methods: In this study, specific primers were designed at both ends of the open reading frame (ORF) based on the unigene (c100572_g1) in the transcriptome data from C. officinalis. Subsequently, the cDNA sequence of CoHMGR1 gene was amplified by RT-PCR, cloned into the pTOPO-T vector and sequenced. This gene and its encoded protein were analyzed using bioinformatics methods. Results: The results suggested that CoHMGR1 was 2 116 bp in length andthe ORF was 1 338 bp in length, which encodes 445 amino acids and is a hydrophobic protein. Multiple sequence alignment and phylogenetic analysis showed that the amino acid sequence encoded by this gene has a high similarity with that of Camptotheca acuminate, reaching 79.56%. Based on the first comparison of transcriptome sequencing from leaves and fruits of C. officinalis, the CoHMGR1 gene was successfully cloned and analyzed. Conclusion: This study laid a foundation for studying the function of CoHMGR1 protein and the molecular mechanism of terpene biosynthesis pathway of C. officinalis.

5.
Article in Chinese | WPRIM | ID: wpr-828376

ABSTRACT

The components of traditional Chinese medicine(TCMCs) are the basic unit of raw materials for Chinese medicines, and their physical and chemical properties directly affect the choice of dosage forms and the optimization of prescriptions. However, most of TCMCs are multi-component complex systems, and the characterization of their overall properties is still in the exploration stage. On the basis of biological activity, the representative components are determined, and then the individual characteristics are fitted with the weight coefficient of efficacy contribution rate, which may provide reference for characterizing the overall properties of TCMCs. In this study, with the pharmacological effects of isoproterenol(ISO)-induced myocardial ischemia in rats as the indicators, the pharmacodynamic contribution rates of three representative components of chishao terpene glucoside components(CSTGCs) were evaluated by the normalization weighting method. The contribution rates of paeoniflorin, paeoniflorin and benzoylpaeoniflorin were 54.87%, 32.46% and 12.67%, respectively. The oil-water partition coefficients of paeoniflorin, albiflorin, benzoylpaeoniflorin in water and buffer solutions with different pH values were measured, and the oil-water partition coefficients of CSTGCs were characterized by the weight of their pharmacodynamics contribution rate. The results showed that the apparent oil-water partition coefficient(log P) of CSTGCs in the phosphate buffer system such as n-octanol-water(pH 2.0, 2.5, 5.0, 5.8, 6.8) were 0.18-0.22, indicating that CSTGCs have common absorption and low permeability, providing basis for the preparation of CSTGCs.


Subject(s)
Animals , Coronary Artery Disease , Glucosides , Medicine, Chinese Traditional , Myocardial Ischemia , Rats , Terpenes , Water
6.
Article in Chinese | WPRIM | ID: wpr-851223

ABSTRACT

Objective To analyze the volatile components of Magnoliae Flos, Magnoliae Flos kernels, and Magnoliae Flos bracts and compare the differences of these components in order to provide a theoretical basis for the usage of kernels of Magnolia Flos in classical famous prescriptions. Methods The volatile components were extracted by steam distillation and analyzed by gas chromatography-mass spectrometry (GC-MS). The volatile components in different parts of Magnoliae Flos were identified by searching the retention index in the mass spectrometry database. The relative content of volatile components was calculated by peak area normalization method, and the data of GC-MS was further proceeded by principal component analysis (PCA). Results The average volatile components content in Magnoliae Flos, Magnoliae Flos kernels, and Magnoliae Flos bracts was 1.70%, 6.79%, and 0.45%, respectively. The numbers of compounds identified by GC-MS in those three different parts were 87, 82, and 85, respectively. These compounds mainly included terpene olefins and alcohols type chemicals. PCA method was then applied to process the GC-MS data and distinguish different parts and habitats of Magnoliae Flos. Conclusion The volatile components in the kernels of Magnoliae Flos were generally the same as those of Magnoliea Flos, while the content of which was much higher than those of whole flower bud of Magnoliae Flos. The results indicated that it was rational to use the kernels of Magnolia Flos in classical famous prescriptions.

7.
Article in Chinese | WPRIM | ID: wpr-851218

ABSTRACT

Objective To study the chemical constituents from the flowers of Bombax malabaricum. Methods The compound was separated and purified by silica gel and gel column choromatography. The structure of the new compound was identified by NMR, HRMS (EIS), IR, UV, and single crystal X-ray diffraction. Results A new compound was isolated from the flowers of B. malabaricum and identified as 3β-acetoxy-22α,30-dihydroxyurs-20-ene. Conclusion Compound 1 was a ursane-type triterpenoid and named as mumian terpene A.

8.
Acta Pharmaceutica Sinica B ; (6): 242-251, 2018.
Article in English | WPRIM | ID: wpr-690914

ABSTRACT

In order to clarify regions of production and to discriminate processing methods, quantitative and qualitative analyses for saccharides and terpenes in 35 batches of Alismatis Rhizoma were performed. Methodologies included HPLC-PDA, HPLC-VWD and UHPLC-MS , combined with principal component analysis (PCA) and partial least squares regression techniques (PLSR). The inhibitory effects of triterpenes and Alismatis Rhizoma extracts on lipase activity were evaluated . PLSR analysis revealed significant positive correlations ( = 0.5795) between the contents of triterpenes , , , and and the inhibitory effects of Alismatis Rhizoma. The present study establishes an effective method for simultaneous determination of multiple components, and identifies key bioactive triterpenes. These results can be used for systematic and novel analytical strategies for the quality control of Alismatis Rhizoma production.

9.
Chinese Journal of Biotechnology ; (12): 862-875, 2018.
Article in Chinese | WPRIM | ID: wpr-687730

ABSTRACT

Santalene and santalol are the main components of valuable perfume sandalwood essential oil, and have good antibacterial, anti-oxidation and anti-tumor activities. Commercial sandalwood essential oil is mainly extracted from sandalwood tree that grows slowly and is difficult to cultivate. In addition, the extraction recovery of sandalwood essential oil from sandalwood tree is too low to meet the market demand. These factors make sandalwood essential oil expensive. An option is to use genetic engineering and molecular biological methods to heterologously express related synthase of santalene and santalol in microbial host. In this paper, the biosynthesis progress of santalene and santalol synthase, as well as the optimization of mevalonate metabolic pathways in the hosts are summarized. Furthermore, the strategies of applying protein engineering technology to carry out orthomutation of santalene synthase were also discussed, to provide reference for the optimal biosynthesis of santalene and santalol.

10.
Article in Chinese | WPRIM | ID: wpr-687286

ABSTRACT

Ginkgo terpene lactones, as an important active ingredient from Ginkgo leaves, has high medicinal values and has been widely used in clinics. This article would review the researches both at home and abroad, including chemical composition, structure-activity relationship, analytical methods, pharmacological effects, pharmacokinetic characteristics, and so on, providing a reference for further development and utilization of ginkgo terpene lactones.

11.
Acta Pharmaceutica Sinica ; (12): 1954-1962, 2018.
Article in Chinese | WPRIM | ID: wpr-780077

ABSTRACT

The discovery and verification of components are prerequisites for developing of component preparations. The molecular docking technique and pharmacodynamic activity evaluation provide effective methods for the discovery and verification of the representative components of Chishao terpene glucoside components (CSTGCS) against ischemia and hypoxia injury. The chemical constituents of CSTGCS were analyzed qualitatively by UPLC-TOF/MS/MS. Main chemical constituents were docked with key receptor proteins of myocardial ischemia to preliminarily screen anti-ischemia active ingredients, and screening for main active ingredients with Libdockscore. Then a H9c2 cell hypoxia injury model was established, and creatine kinase (CK), lactate dehydrogenase (LDH), superoxide dismutase (SOD), malondialdehyde (MDA) were determined to screen the representative combinations in CSTGCS. In addition, apoptosis index, apoptotic protein expression and mitochondria-associated mRNA levels were determined to verify the inhibition of the representative components (RCS) on the apoptosis of hypoxic cells. Eventually, the representative components of CSTGCS were determined. The results showed that paeoniflorin, albiflorin, benzoyl paeoniflorin and oxypaeoniflorin were considered to be the main active components because of their high matching with target proteins (4TWT, 3O4O, 4KZN, 1M9J) in space and energy. There was no statistical difference in regulating CK, LDH, SOD, MDA levels and maintaining mitochondrial function as well as inhibiting cell apoptosis between CSTGCS group and RCS group (paeoniflorin + albiflorin + benzoyl paeoniflorin combination). Therefore, paeoniflorin, albiflorin and benzoyl paeoniflorin were selected as the most representative ingredients of CSTGCS against ischemia and hypoxia injury, providing a basis for the overall properties of the components and formulation of CSTGCS.

12.
Bol. latinoam. Caribe plantas med. aromát ; 16(2): 110-120, mar. 2017. tab, ilus
Article in English | LILACS | ID: biblio-881391

ABSTRACT

The peel components and juice quality of three mandarin hybrids were investigated in this study. Peel components were extracted using the cold-press method and analyzed using GC-FID and GC-MS. Total soluble solids, total acid, pH value, ascorbic acid as well as density were determined in juice obtained from mandarin hybrids. Twenty-six, thirty-five and nineteen peel components were identified in Fortune, Robinson and Osceola respectively including: aldehydes, alcohols, esters, monoterpenes, sesquiterpenes and other components. The major components were limonene, γ-terpinene, (E)-ß-ocimene, ß-myrcene, sabinene, linalool and α-Pinene. Among the three scions examined, Fortune showed the highest content of aldehydes and Robinson showed the highest content of TSS. This study shows that scion has a profound influence on aldehyd and TSS that are important to quality improvement.


Los componentes de la cáscara y la calidad del jugo de tres híbridos de mandarina fueron investigados en este estudio. Los componentes de la cáscara fueron extraídos por el método de prensado en frío y se analizaron mediante GC-FID y GC-MS. el total de sólidos solubles, acidez total, pH, ácido ascórbico, así como la densidad se determinaron en el jugo obtenido a partir de híbridos de mandarina. Veintiséis, treinta y cinco y diecinueve componentes de la cáscara de Fortuna, Robinson y Osceola respectivamente fueron identificados, incluyendo: aldehídos, alcoholes, ésteres, monoterpenos, sesquiterpenos y otros componentes. Los componentes principales eran limoneno, γ-terpineno, (E) -ß-ocimeno, ß-mirceno, sabineno, linalool y α-pineno. Entre los tres vástagos examinados, Fortune mostró el mayor contenido de aldehídos y Robinson mostró el mayor contenido de SST. Este estudio muestra que el vástago tiene una profunda influencia en aldehído y SST que son importantes para mejorar la calidad.


Subject(s)
Citrus/chemistry , Fruit and Vegetable Juices/analysis , Terpenes/analysis , Chromatography, Gas/methods , Quality Control
13.
China Pharmacist ; (12): 434-437, 2017.
Article in Chinese | WPRIM | ID: wpr-510097

ABSTRACT

Objective:To investigate the optimal extraction and dripping pills preparation of total terpene constituents in platycladi seed. Methods:The extraction processes of ultrasound, reflux, microwave and homogenization were investigated by single factor tests. The effects of extraction time, temperature, solid-liquid ratio and extraction times on the extraction of total terpene constituents in platy-cladi seed were investigated by orthogonal tests. Orthogonal tests were used to investigate the effects of the ratio of polyethylene glycol 6000 to poloxamer 188, the ratio of drug to matrix, the temperature of liquid and the dropping process on the quality of dropping pills. Results:The homogeneous extraction was used, and the optimum extraction conditions were as follows: the extraction time was 10 min, the temperature was 80℃, the solid-liquid ratio was 1 :10, and extraction time was once. The best preparation process of drop-ping pills was as follows:the ratio of polyethylene glycol 6000 to poloxamer 188 was 5 :1, the ratio of drug to matrix was 1 :3, the temperature of liquid was 80℃ and the dropping distance was 10 cm. Conclusion: The method of homogeneous extraction is simple, which can break the materials and extract the active ingredients simultaneously. The preparation process of dropping pills is reasonable and feasible, which is easy for the production and application.

14.
Herald of Medicine ; (12): 665-668, 2017.
Article in Chinese | WPRIM | ID: wpr-614288

ABSTRACT

Objective To explore the feasibility of using the quantitative reference extract of ginkgo leaf total lactones instead of single component reference for the quantitative assay of Ginkgo Folium.Methods HPLC-ELSD method was performed by using a Diamonsil C18 column (250 mm×4.6 mm,5 μm) with methanol-water as the mobile phase at the gradient elution mode.Flow rate was 1.0 mL·min-1.The parameters of ELSD detector were as follows,the drifit tube temperature was 105 ℃,and the flow rate of nitrogen(N2) was 3 L·min-1.Results The linear ranges of ginkgolide A,ginkgolide B,ginkgolide C,and bilobalide were 0.735-5.879 μg (r=0.999 6),0.404-6.060 μg (r=0.999 6),0.296-4.439 μg (r=0.999 6),and 1.001-6.006 μg (r=0.999 7),respectively.The recoveries and RSD of the four components were 95.6% (4.0%),97.3% (4.5%),99.3% (5.0%),and 100.4% (2.1%),respectively.Conclusion The quantitative reference extract of ginkgo leaf total lactones can be used as the substitute for the determination of terpene lactones.

15.
Article in Chinese | WPRIM | ID: wpr-275135

ABSTRACT

The embryos of model organism zebrafish were used to evaluate the acute toxicity of the extracts of Euphorbiae Pekinensis Radix and vinegar-processing Euphorbiae Pekinensis Radix, and the total terpene content of each extract was determined by using euphol as the reference standards. Twenty-four h normally developed zebrafish embryos were chosen, and 8 concentrations were adopted for each extract. Then the growth and death of zebrafish embryos were observed at 96 h after administration, and median lethal concentrations (LC50) of the different samples on zebrafish embryos were calculated. The results showed that all of the extracts (before and after vinegar processing) had acute toxicity on zebrafish embryos. The toxicity of vinegar-processing Euphorbiae Pekinensis Radix was significantly lower than that of crude Euphorbiae Pekinensis Radix. Among different extraction methods, ethanol extract was more poisonous than water extract; in different polarity fractions, the toxicity was in the following order: petroleum ether>dichloromethane>ethyl acetate>n-butyl alcohol and remaining part. Combined with the results of the determination of terpene components, it can be concluded that the terpenoids are the main toxic components of Euphorbiae Pekinensis Radix, positively correlated with toxicity degree. It indicates that the zebrafish embryo model is appropriate for the toxicity evaluation of Euphorbiae Pekinensis Radix and provides appropriate research methods and theoretical basis for the further study of the toxic components and the mechanism of reducing toxicity.

16.
Bol. latinoam. Caribe plantas med. aromát ; 15(4): 199-205, jul. 2016. tab
Article in English | LILACS | ID: biblio-907537

ABSTRACT

Aloysia polystachya and Lippia turbinata are medicinal and aromatic plants. Nevertheless, there are few reports in literature concerning the biological properties of species that grow in northeastern Argentina. The antibacterial activity and the chemical composition of both essential oils were evaluated in this work. The extraction was performed by steam distillation and their volatile compounds were determined by gas chromatography/mass spectrometry. The antibacterial activity was evaluated by disc diffusion and broth microdilution assay. The main compounds were carvone (78.9 percent) and limonene (14.2 percent) in A. polystachya and carvone (80.77 percent), limonene (8.73 percent), beta- caryophyllene (2.13 percent) and 1,8-cineole (1.70 percent) in L. turbinata. Both essential oils were bactericide against Escherichia coli ATCC 35218 and clinical isolates of Enterobacter cloacae and Klebsiella pneumoniae. Essential oil of A. polystachya was also bactericide against Staphylococcus aureus ATCC 29212, S. aureus ATCC 25923 and clinical strain of S. aureus methicillin susceptible.


Aloysia polystachya y Lippia turbinata son plantas medicinales y aromáticas. Hay pocos informes en la literatura sobre las propiedades biológicas de especies que crecen en el nordeste de Argentina. La actividad antibacteriana y la composición química de ambas especies se evaluaron en este trabajo. La extracción se realizó por destilación con vapor y sus compuestos se determinaron por cromatografía gaseosa/espectrometría de masa. La actividad antibacteriana fue evaluada por difusión en discos y microdilución en caldo. Los principales compuestos fueron carvona (78.9 por ciento) y limoneno (14.2 por ciento) en A. polystachya y carvona (80.77 por ciento), limoneno (8.73 por ciento), beta-cariofileno (2.13 por ciento) y 1,8-cineol (1.70 por ciento) en L. turbinata. Ambos aceites esenciales fueron activos contra Escherichia coli ATCC 35218 y aislamientos clínicos de Enterobacter cloacae y Klebsiella pneumoniae. El aceite esencial de A. polystachya fue bactericida contra Staphylococcus aureus ATCC 29212, S. aureus ATCC 25923 y aislamientos clínicos de S. aureus sensible a meticilina.


Subject(s)
Anti-Bacterial Agents/pharmacology , Oils, Volatile/chemistry , Oils, Volatile/pharmacology , Plant Extracts/pharmacology , Verbenaceae/chemistry , Anti-Bacterial Agents/chemistry , Enterobacter cloacae , Escherichia coli , Staphylococcus aureus , Terpenes/analysis
17.
Chinese Pharmaceutical Journal ; (24): 1278-1281, 2016.
Article in Chinese | WPRIM | ID: wpr-859017

ABSTRACT

OBJECTIVE: To study the terpene constituents from Cunninghamia lanceolata (Lamb.) Hook. METHODS: The ethanol crude extract was fractionalized by using EtOAc, then a series of chromatography methods were used to isolate the terpene compounds and the structures were elucidated by their physicochemical properties and spectroscopic data. RESULTS: Nine terpenes were isolated and identified as 8(17),12E,14-labdatrien-18-al(1),4S,5R,9S,10R-8(17),12,14-labdantrien-18-ol acetate(2),4S,5R,9S,10R-8(17),12,14-labdantrien-18-ol(3),4S,57R,9S,10R-8(17), 12,14-labdantrien-18-oic acid(4),3β-hydroxy-8(17),13E-labdandien-15-oic acid(5), hinokiol(6),8,15-pimaradien-18-oic acid(7),15-nor-14-oxo-8(17),12-labdandien-18-oic acid(8), and 15-nor-14-oxo-8(17),12-labdan-dien-18-ol(9). CONCLUSION: Compounds1,2,5-7 and 9 are isolated from this plant for the first time.

18.
Article in Chinese | WPRIM | ID: wpr-853710

ABSTRACT

Objective: To clone the full-length cDNA sequence which encodes one of the key enzymes of terpene biosynthesis, farnesyl pyrophosphate synthase (FPPS) and analyze the bioinformation. Methods: Based on the transcriptome data of Paeonia lactiflora (Pl), a pair of specific PCR primers were designed. The objective PCR band of PlFPPS was successfully amplified using RT-PCR technique and then it was cloned to pMD18-T vector. After clone and plasmid PCR characterization, the recombinants were sequenced and then a series of bioinformatic analysis was carried out including sequence homology search, construction of molecular phylogenetic tree, domain search, and 3D structure prediction, etc. Results: Sequencing results showed that the full-length PlFPPS was 1 315 bp, which contained 60 bp 5'-UTR, 1 050 bp CDS, and 205 bp 3'-UTR. It encoded 349 amino acids and the accession number of GenBank was KP708571.Blastn and Blastp analysis revealed that PlFPPS (KP708571) and its encoding protein (AKJ26301) had high homology with FPPS genes and proteins from several plants at the nucleotide and amino acid levels. Phylogenetic tree analysis indicated that the genetic relationship of PlFPPS with FPPS from other plants was relatively far. It predicted that the molecular weight and isoelectric point of PlFPPS were 40200 and 5.33, which was a hydrophilic and stable protein, located in cytoplasm without transmembrane domain and signal peptide. It contained seven conservative domains like substrate binding pocket, substrate-Mg2+ binding site, catalytic site, aspartate-rich region 1, and aspartate-rich region 2.The proportion of α-helix in its 3D structure was high and the α-helix was linked by β-loop. The central cavity was composed of 10 core α-helixes. Conclusion: The full-length cDNA sequence of FPPS is cloned firstly from P. lactiflora and the bioinformatic analysis is then carried out.

19.
Article in Chinese | WPRIM | ID: wpr-272716

ABSTRACT

The open reading frame of 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase (HDR) was cloned from Phlegmarirus carinatus by RT-PCR method and the sequence was analyzed by bioinformatics tools. After searching the transcriptome dataset of P. carinatus, one unique sequence encoding 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase was discovered. The primers were designed according to the cDNA sequence of 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase from the dataset. And then, the open reading frame (ORF) of 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase, named as PcHDR1 (GenBank Accession number:JQ957845), was cloned by RT-PCR strategy with the template of mixed RNA extracted from roots, stem and leaf of P. carinatus. The bioinformatic analysis of this gene and its corresponding protein was performed. The ORF of PcHDR1 consisted of 1 437 base pairs (bp), encoding one polypeptide with 478 amino acids. The sequence comparison showed that PcHDR1 is closest with GbHDR (Ginkgo biloba),and the sequence homology was up to 78%. Bioinformatics prediction and analysis indicated that PcHDR1 protein contained a conserved domain of LytB, without transmembrane region and signal peptides. This study cloned and analyzed 1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase from P. carinatus. The result will provide a foundation for exploring the function of PcHDR1 involved in terpene biosynthesis in P. carinatus plants.

20.
Article in Chinese | WPRIM | ID: wpr-854270

ABSTRACT

To compare the quality Ginkgo biloba L. leaves collected form various habitats, different tree ages, and different harvesting times and to confirm the source of G. biloba leave used in Xin Mai Capsule. To establish the HPLC fingerprint of G. biloba leaves to determine the fingerprint and content of total flavonol glucoside and terpene lactones in G. biloba leaves collected from various habitats, different tree ages, and different harvesting time, and then to analyze the results. The contents of total flavonol glucoside and terpene lactones and fingerprint similarity in G. biloba leaves from various regions, different tree ages, and collecting times were different. The fingerprint control method has provided the scientific basis for the reasonable collection and quality evaluation of G. biloba leaves. The G. biloba leaves in June to August, 2-3 year-old from Pizhou, Jiangsu province are better, which could be considered as the source of G. biloba leaves used in Xin Mai Capsule.

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