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    20 November 2020, Volume 40 Issue 6
    TwoNew Species of Delphinium from Xinjiang
    Wen-Tsai WANG, Zong-Zong YANG
    2020, 40(6):  801-804.  doi:10.7525/j.issn.1673-5102.2020.06.001
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    Two species of the genus Delphinium(Ranunculaceae), D.adenopodum and D.gongliuense, are described as new from Xinjiang Weiwuer Autonomous Region. The diagnostic differences between the two species and their allies are given respectively.

    Pollen Morphology and Systematic Analysis of Fifteen Sanicula Species from China
    Chen YANG, Zhi-Xiang CHEN, Xue-Ying YAO, He-Shan WANG, Qi-Zhi WANG
    2020, 40(6):  805-812.  doi:10.7525/j.issn.1673-5102.2020.06.002
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    The pollen morphology of 15species of Sanicula was observed by light microscopy(LM) and scanning electron microscopy(SEM). The pollen size of 15 species of Sanicula was(28.51-54.29) μm-(12.23-28.98) μm, and the ratio of polar axis to equatorial axis(Polar axis/Equatorial axis) was 1.48-2.46 with a three-hole ditch. Views of pollen grains include near-rectangular, equatorial contraction, long ellipse etc. Among them, near-rectangular and equatorial contraction are the majority. Polar views are round or triangular. Under the scanning electron microscope, the outer wall surface is reticular. Our studies support that in the Saniculoideae(Apiaceae), the pollen of the genus Sanicula has moderately evolved and relatively evolved morphological features. The pollen morphology can provide palynological evidence in the classifying individual species.

    Floral Organogenesis of Hypericum przewalskii(Hypericaceae) and Its Systematic Implications
    Xiang ZHAO, Xue SU, Hai-Yan WU, Hui ZHANG, Kun SUN
    2020, 40(6):  813-819.  doi:10.7525/j.issn.1673-5102.2020.06.003
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    The floral organogenesis of Hypericum przewalskii(Hypericaceae) was observed under scanning electron microscope. Two bract primordia occur first, and the flower primordium develops in the package of the bract primordia. Followed the initiation of bract primordia, five sepals are initiated by 2/5 circumference spiral. With the sepal primordia initiated, the angles between the sepal interstices swell up into large protuberances which become differentiated as common stamen-petal primordia. Five common stamen-petal primordia initiated simultaneously. Petal primordia are separated from the lower half of the common stamen-petal primordia by a median slit. The remaining upper half of the common stamen-petal primordia grows into a hemispherical structure, thus the five stamen primordia initiate opposite to the petal primordia. Then the stamen primordia sequentially differentiated from the inside to the outside to produce the secondary stamen primordia. With the development of the secondary stamen primordia and increasement of primordia, fascicled stamens were formed. When the secondary stamen primordia initiated, the five carpal primordia initiated in a simultaneous type. The development way of the fascicled stamens in H.przewalskii indicated that fascicled stamens in Hypericum may originate from haplostemonous structure. The significant difference in patterns of petal and stamen primordia between Hypericaceae and Clusiaceae supports the viewpoint of APG Ⅲ classification that Hypericaceae are separated from Clusiaceae.

    Fruit Shape and Micromorphological Characteristics of Pericarp Surface of 18 Lamiaceae Plant in Jinggangshan Area
    Xian-Lan DENG, Xia-Xia CHEN, Zheng-Guang ZHANG
    2020, 40(6):  820-829.  doi:10.7525/j.issn.1673-5102.2020.06.004
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    The fruit shapes and micromorphological characteristics of fruit surface of 18 species of Lamiaceae in Jinggangshan area were examined by scanning electron microscopy(SEM). Six types of the fruit shape were recognized, including oblong, circular, oval, triangular, globose, and ellipsoid. The cell shapes of exocarp surface were tetragon-hexagon, irregular-shaped and invisible, and the protrusion of epidermis were observed as smooth, depressed and raised. The waxy ornamentation of exocarp surface was more complex and diverse, which can be classified into four types: smooth and no ornamentation, negative-reticular ornamentation, reticular ornamentation with warty or lumpy projections, reticular ornamentation with secondary stripes. The accessory structures mainly included epidermal hairs and filamentous structures, and with more or less squamaccous or granulated secretions on fruit epidermal surface. In conclusion, fruit shapes and micromorphological characteristics of pericarp surface of the 18 species exhibited rich diversity, which could provide important taxonomic evidence for the study of inter-genera and interspecies relations of Lamiaceae.

    Analysis on Community Structure and Dominant Population Point Pattern of Secondary Forest of Quercus mongolica
    Xiang LI, Wan-Sheng LIU, Wei ZHOU, Fu-Yuan CHEN, Li-Qiang MU
    2020, 40(6):  830-838.  doi:10.7525/j.issn.1673-5102.2020.06.005
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    To explore the mechanism of species coexistence, biodiversity conservation and maintenance in ecotone of Daxing’an and Xiaoxing’an mountains, the community species composition, structural characteristics, size structure and spatial point pattern of dominant population were analyzed based on the data of 25 hm2 forest plot of natural secondary forest of Quercus mongolica at Black-billed Capercaillie National Nature Reserve of national station in Heilongjiang province. The results showed that there were 10 species of vascular plants with DBH ≥1 cm in the plot, and 34 778 living trees with an average density of 1 392 trees·hm-2. There were 4 species with important values of more than 10%, which were Quercus mongolica Fisch. ex Ledeb.,Betula dahurica Pall.,Betula platyphylla Suk.,Larix gmelinii(Rupr.) Kuzen successively, accounted for 89.65% of the whole plot. The Shannon-Wiener diversity, Simpson diversity and Pielou evenness index were 1.577, 0.760 and 0.685, respectively. The results showed that in this community, there were fewer rare species, larger proportion of common species than the others. What’s more, the community structure was simple. The average DBH of all tree species in the sample plot was 10.77 cm. The total diameter distribution in the community was of “reversed J type”, which was consistent with the distribution characteristics of typical uneven-aged forests. The diameter distribution of B.dahurica and B.platyphylla was approximately “reversed J type”, belonging to the growth populations, while the population of Q.mongolica showed a bimodal structure. The population of L.gmelinii was a declining species with an approximate normal size distribution and a tendency to disappear with succession. Under the CSR model, the spatial distribution of the four dominant populations was aggregation distribution and the degree of aggregation decreases with the increase of the scale, finally presenting a random or uniform distribution trend. Under the NS model, except L.gmelinii, the remaining 3 planted strains showed aggregation at 0-5 m scales, possibly influenced by the diffusion limitation of seeds. Comparing aggregation intensity of the four populations, it was found that B.dahuricaB.platyphyllaQ.mongolicaL.gmelinii. Therefore, we might reasonably infer that the aggregation intensity of different populations was not only related to the scale of the study, but also to the population abundance.

    Effects of Rare Earth Element Lanthanum on Adventitious Buds Induction, Plantlets Growth and Synthesis of Secondary Metabolites of Dendrobium officinale
    Yong-Mei MIAO, Yuan Tong, Da Fang, Yu-Hao Wang
    2020, 40(6):  839-845.  doi:10.7525/j.issn.1673-5102.2020.06.006
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    The experiment was conducted to study the effects of rare earth element lanthanum on adventitious buds induction, plantlets growth and secondary metabolites synthesis of Dendrobium officinale by adding 30-150 μmol·L-1 La(NO33 to the media. The effects of 70-90 μmol·L-1 La3+ on induction and growth of adventitious buds were the best. The treatment of 130 μmol·L-1 La3+ could significantly increase chlorophyll content of seedlings, and 110 μmol·L-1 La3+ could significantly promote plant growth with 15.22 times of fresh weight, and 12.05∶100 of dry weight-fresh weight ratio, which was 64.39% higher than that of control. A certain concentration of La3 + could significantly improve the content of four components in D.officinale. The polysaccharide content reached the peak value of 98.84 mg·g-1 under 110 μmol·L-1 La3+ treatment. The contents of flavonoids, phenolic acid and bibenzyl reached maximum value of 4.31, 7.56 and 21.01 mg·g-1, respectively, when 90 μmol·L-1 La3+ added to media. Both indexes of bibenzyl and flavonoids were highly correlated with other five indexes, while phenolic acid and chlorophyll content were little correlated with other indexes. This study laid the foundation for further exploring the mechanism of rare earth on promoting growth and improving quality of D.officinale, and provided a theoretical basis for scientific and rational utilization of rare earth elements in plant tissue culture.

    In vitro Germination,Callus Induction and Primary Haustorium Organogenesis in the Parasitic Plant Cynomorium songaricum
    Xin YUE, Gui-Lin CHEN
    2020, 40(6):  846-854.  doi:10.7525/j.issn.1673-5102.2020.06.007
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    Cynomorium songaricum Rupr. is an obligate parasitic plant widely used as traditional Chinese medicine and Mongolian medicine. Here, we firstly describe protocols for in vitro germination, callus induction and haustorium organogenesis in C.songaricum Rupr. In this study, adequate concentrations of gibberellic acid(GA3),in combination with other plant growth regulators in the medium, promoted embryo development and germination of C.songaricum seeds. The highest callus induction rate from seeds(13.7%) after a 40 d incubation was obtained with B5 medium adding 2,4-dichlorophenoxyacetic acid(2,4-D; 1.0 mg·L-1), kinetin(KT;0.5 mg·L-1) and GA3(1.0 mg·L-1. This resulted in callusformation in 13.7%±3.1% of seeds. Addition of 2,4-D(0.5 mg·L-1) and KT(0.25 mg·L-1) yielded highest haustorium organogenesis from calluses. Some primary haustorium branched to form adventitious roots of 3-4 cm in length. Subsequently, the tip of each adventitious root formed a nascent primary haustorium, which was then branched out into adventitious roots. The role of auxins(2,4-D in this study) in the formation of primary haustorium and adventitious roots from seed callus in C.songaricum was also discussed.

    Differences in Population Structure and Dynamic Characteristics of Davidia involucrata Baill. Between High and Low Latitude Regions
    Yun-Fei XU, Qin-Song LIU, Wen-Juan XU, Jun-Feng TANG, Ting-Fa DONG, Bao-Zhen YANG, Xiao XU
    2020, 40(6):  855-866.  doi:10.7525/j.issn.1673-5102.2020.06.008
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    Davidia involucrata Baill. is a rare and endangered tree species which is endemic to China. Though population characteristics of D.involucrata have been reported by many studies, the effect of latitude on the structure and dynamic characteristics of D.involucrata has been largely neglected. In this study, we compared the variance of the age structure dynamic index, static life table, viability and quantity alteration of the populations between high latitude region(Pingwu) and low latitude region(Northeastern Yunnan). Our results showed that the population structure in Pingwu and Northeastern Yunnan were of growth type, the survival curve of the two D.involucrata populations was of a Deevey-Ⅱ type, and the population in Northeastern Yunnan displayed a higher growth trend and stronger anti-interference ability. Moreover, the population dynamics analysis revealed that younger individuals grew better in Pingwu and the middle-aged individuals competed fiercely. By contrast, younger individuals in Northeastern Yunnan population had high mortality, middle-aged individuals grew stable. Thus, based on the abovementioned findings, we conclude that latitude differences could affect population structure and dynamic characteristics of D.involucrata. The habitats of D.involucrata population at low latitude were suitable for middle-aged individuals to survive but not for younger individuals; younger individuals of D.involucrata populations at high latitude grew better, and higher mortality rate was observed in middle-aged individuals.

    Effects of Invasive Plant Datura stramonium on the Functional Traits of Native Plants and the Stoichiometric Characteristics of Soil Carbon, Nitrogen and Phosphorus
    Ke-Xin MA, Mei ZHANG, Xin FANG, Li-Li TANG, Jian-Hua HAN, Li-Fang YANG, Fu-Chen SHI
    2020, 40(6):  867-875.  doi:10.7525/j.issn.1673-5102.2020.06.009
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    The experiment was conducted to explore effects of invasive plants under the different pressures of invasion on the functional traits of native plants and the stoichiometric characteristics of soil carbon, nitrogen and phosphorus. We used invasive plant Datura stramonium and coexisting native plants as the research object, researches species, number and height of plants in sample plots and the species diversity of native plant communities with no invasion, low invasion and heavy invasion(according to the proportion of invasive plants coverage), and analyzed the specific leaf area, carbon content, nitrogen content, carbon to nitrogen ratio, leaf construction cost and carbon, nitrogen and phosphorus stoichiometric characteristics in different soil layers. With the increase of invasion pressure, the species richness and the number of native plants decreased gradually. The plant height and leaf nitrogen content of D.stramonium were significantly higher than that of native plants under different invasion pressures, and gradually increased with the increase of invasion pressure. The carbon to nitrogen ratio was significantly lower than that of native plants in sample plots with no invasion. The specific leaf area, carbon content and leaf construction cost were not significantly different from those of native plants in sample plots with invasion. With the increase of invasion pressure, soil total nitrogen content, total carbon content, nitrogen to phosphorus ratio and carbon to phosphorus ratio increased significantly, while total phosphorus content and carbon to nitrogen ratio decreased significantly. Soil carbon and nitrogen stoichiometric characteristics show a certain surface clustering effect. These results show that D.stramonium has a high resource capture ability, and has changed the characteristics of soil, so as to enhance its own competitiveness to improve the invasion ability, which may be one of the reasons for the successful invasion of D.stramonium.

    Molecular Phylogenetic Analysis of Cerasus Plants in China
    Tao FU, Zhi-Long WANG, Le-Jing LIN, Li LIN, Wen LI, Dong-Ming YUAN
    2020, 40(6):  876-885.  doi:10.7525/j.issn.1673-5102.2020.06.010
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    To provide some molecular basis for the species classification and lineage location of cherry blossoms mainly planted in China, with 88 cherry blossoms, by nuclear gene ITS and chloroplast intergenic sequence, trnH-psbA were sequenced, and after processed by software MEGA 6, phylogenetic tree was constructed. The phylogenetic development was confused in Cerasus serrulata group, which is related to the complex genetic background and the close phylogenetic relationship among the primitive parental, meanwhile, it could be clustered together in most of the cultivars C.subhirtella group or C.campanulata group, and could be separated from the cultivars of the Cerasus serrulata group, however, it was difficult to distinguish in many hybrids using ITS and trnH-psbA, which indicated that DNA barcoding technology was difficult to apply to the phylogenetic relationship of hybrids, in addition, some cherry cultivars could be identified by ITS or trnH-psbA sequence length, and some unidentified varieties were also revised.

    Identification of Key Enzyme Genes Involved in Anthocyanin Synthesis pathway in Clinopodium gracile by Transcriptome Analysis
    Li-Qiang ZHAO, Chun-Miao SHAN, Sheng-Xiang ZHANG, Yuan-Yuan SHI, Ke-Long MA, Jia-Wen WU
    2020, 40(6):  886-896.  doi:10.7525/j.issn.1673-5102.2020.06.011
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    In order to further understand the pathway of anthocyanin synthesis, the four tissues of C.gracile, namely, root, stem, leaf and flower, were sequenced using BGISEQ-500 platform. 128 856 unigenes were assembled and 40 unigenes encoding six key enzymes involved in the anthocyanin biosynthesis were identified. Homologous alignment and structure simulation of bifunctional dihydroflavonol 4-reductase(DFRs) were carried out, which showed that they have well-conserved sequences, spatial structures and NAD+ binding sites. The DFR structure model showed a tetrameric structure with Rossmann-like α-helixes/β-sheets/α-helixes sandwich fold.

    Leaf Color and Growth Variation of Transgenic Gold Leaf Poplar
    Yi-Di LI, Chen-Rui GU, Hui-Xin GANG, Gui-Feng LIU, Su CHEN, Jing JIANG
    2020, 40(6):  897-905.  doi:10.7525/j.issn.1673-5102.2020.06.012
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    The chlorophyll content of the PaGLK gene Populus alba×P.berolinensis inhibited expression line was significantly reduced, and the leaves showed yellow(named golden leaf silver poplar). Using PaGLK transgenic Populus alba×P.berolinensis, its leaf color parameters and chlorophyll content were measured in time series, and its growth characteristics were analyzed. The results showed that PaGLK transgenic Populus alba×P.berolinensis changed the leaf color, and the chlorophyll content of the expressing lines was significantly lower than that of WT(P<0.05), and the leaf brightness was significantly higher than that of WT(P<0.05). During the growth and development period, the leaves showed dark yellow green. The daily growth height(GD) of seedling height in the Y2 fast-growing stage of the expressing lines was higher than that of the control lines, and the plant height at the seedling stage was not affected. The over-expressing transgenic Populus alba×P.berolinensis plants were significantly lower than the control plants in that year(P<0.05), and the average daily growth height(GD) of the seedling height in the fast-growing period was also 22.19% of that of the control plants. PaGLK inhibited expression strains have potential application value in urban landscaping.

    Analysis of SHMT Gene Family and Production of PtrSHMT9 Mutant in Populus trichocarpa
    Bing LI, Yu-Xiang CHENG
    2020, 40(6):  906-912.  doi:10.7525/j.issn.1673-5102.2020.06.013
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    Serine hydroxy methyltransferases(SHMTs) played an important role in the one-carbon(C1) metabolism pathway in plant cells. We identified 9 SHMT genes from the genome of Populus trichocarpa. Populus eFP resource showed that 7 PtrSHMTs were expressed in multiple tissues, among which PtrSHMT9 was the most highly expressed gene in xylem. By analysis of RT-PCR and Aspwood, PtrSHMT9 had a high level in secondary wall thickening of stem wood tissue, suggested its involvement in poplar wood formation. Using Cas9/gRNA technique, PtrSHMT9-edited mutants were prepared and 4 different ptrshmt9 knockouts were obtained. Our results provided basic information and genetic material for exploring the function of SHMT and PtrSHMT9 in trees.

    Cloning,Structure and Function Analysis of MiMYB2 Gene from Macadamia integrifolia
    Wen-Lin WANG, Hai-Sheng CHEN, Shu-Fang ZHENG, Song-Le FAN, Li-Feng WANG, Qiu-Jin TAN, Zhen-Shi QIN, Xi-Yun HUANG, Peng HE, Xiu-hua TANG, Peng XU
    2020, 40(6):  913-922.  doi:10.7525/j.issn.1673-5102.2020.06.014
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    In order to study the mechanism of R2R3-MYB transcription factor gene family member in growth,development and yield formation in Macadamia integrifolia,the MiMYB2 gene was cloned from the leaves of M.integrifolia “GUIRE1” cultivar by PCR and its structure and function were analyzed by bioinformatics method. The cloned cDNA sequence of MiMYB2(NCBI accession number:MN254976) gene is 1 210 bp in length,has 1002 bp open reading frame(ORF),encoding 333 amino acids. MiMYB2,an unstable hydrophilic protein that has no transmembrane structure,no signal peptide and localizes in the nucleus,has two SANT conserved domains belonging to the R2R3-MYB family. BLAST analysis showed that the MiMYB2 share high amino acid sequence identity with Lotus NnMYB3-like protein. Phylogenetic analysis clustered MiMYB2 and AtMYB17,AtMYB106 and AtMYB16 into S9 subfamily. The transcriptome data were used to analyze the expression patterns of MiMYB2 gene in branch,flower and leave of M.integrifolia “GUIRE1” and “695” cultivar. Results indicated that MiMYB2 has the lowest expression in the flower of “GUIRE1”,and the expression level of “695” is the highest. It is speculated that MiMYB2 is closely related to M.integrifolia growth and development. This study would provide a theoretical reference for clarification the role MiMYB2 in growth,development and yield formation mechanism of M.integrifolia.

    Analysis of Limonium bicolor by SSRMolecular Markers in the Yellow River Delta Region
    Dong-Xue LÜ, Xue-Jie ZHANG, Luo-Yan ZHANG, Shou-Jin FAN
    2020, 40(6):  923-931.  doi:10.7525/j.issn.1673-5102.2020.06.015
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    Genetic diversity parameters, principal coordinates analysis(PCoA), and analysis of molecular variance(AMOVA) were conducted by using GENALEX6.502 and ARLEQUINv3.5 software. STRUCTUREv 2.3.4, STRUCTURE Harvester, CLUMP, and Distruct software were used to analyze genetic structure. We systematically sampled 202 individuals from 12 Limonium bicolor populations distributed throughout the Yellow River Delta(YRD) region in China. By exploring the micro-geographic genetic structure using the codominant microsatellite(SSR) markers, we aimed to answer questions as to what extent the population structure and dynamics of L.bicolor YRD. The demonstrated a medium level of genetic differentiation in L.bicolor. The genetic variation was mainly maintained within populations, with a weak but significant genetic differentiation being detected among populations(FST=0.067). Shannon’s information index was 1.037, and gene flow was 4.106, the observed heterozygosity(Ho) and expected heterozygosity(He) was 0.43 and 0.529. Genetic diversity and population genetic structure were analyzed in L.bicolor to provide useful information for management, protection and utilization. In brief, suggesting that more individuals from different populations should be collected to protect the genetic diversity of L.bicolor.

    Subcellular Localization of the Protein Coded by the UDP-Glucose Dehydrogenase Gene from Paper Mulberry and Functional of Its Promoter 5′-end Deletion Fragment
    Ren-Hua JI, Wen-Bo ZHANG, Xiao-Fei LIN, Ying-Liang BAO, Ri-Ge-Le TE, Hui-Ga BAO, Shu-Lan BAI
    2020, 40(6):  932-942.  doi:10.7525/j.issn.1673-5102.2020.06.016
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    We analyzed the function of cis-acting elements of UDP-Glucose Dehydrogenase Gene from Paper Mulberry(DDBJ, BpUGDH accession No.LC457701)promoter, the five different lengths of BpUGDH promoter 5'-terminal deletion fragment were ligated with the GUS gene by using 5'terminal deletion and homologous recombination techniques, and conducted through heterologous expression in Nicotiana benthamiana by agro-infiltration method. In order to locate the specific location of the protein encoded by BpUGDH gene in cells, subcellular localization of BpUGDH protein was carried out by using the fusion target gene of GFP reporter gene. The results show that the sequences within -244 bp can mediate the induction of GUS gene expression, and the region between -973, -465, -355, -281, -244 bp of the BpUGDH promoter were crucial for regulating the promoter activity. Furthermore, BpUGDH was located in chloroplast.

    Preparation and Bioavailability of Coenzyme Q10 Xylan Nanometer Suspension Agent
    Xiao-Xue ZHANG, Xiu-Hua ZHAO, Yan-Jie LIU, Ling-Ling WANG, Qian ZHANG
    2020, 40(6):  943-950.  doi:10.7525/j.issn.1673-5102.2020.06.017
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    A novel copolymer-supported coenzyme Q10 formed a nanometer suspension agent and increased the water solubility of CoQ10 and improved its oral bioavailability. The quercetin-Xylan(qt-xylan) copolymer coupling was used as the basis for synthesis. The single factor experiment design was adopted, and the particle size was used as the inspection condition of the single factor experiment. The factors affecting the particle size included high pressure homogenization pressure, high pressure homogenization times, copolymer concentration, mass ratio between copolymer and CoQ10, and a series of in vitro experiments were carried out. When the homogenization pressure was 60 MPa, the number of homogenization was 7, the copolymer concentration was 1 mg·mL-1, and the mass ratio of copolymer to CoQ10 was 1∶1, it was the best preparation process for the nanometer suspension agent, and the particle size was 166.7 nm. Under the optimum technological conditions in vitro dissolution experiment,thedissolution rate of CoQ10 nanometer suspended agent coated in artificial gastric juice(SGF) and artificial intestinal juice(SIF) was 1.89 and 1.48 times that of CoQ10 active drug, respectively. The bioavailability in vivo experiments, rat were loaded on gastric CoQ10 and nanoparticle suspension agent respectively. Then the blood drug concentration, different time points in drug absorption and metabolism of rats were detected respectively. The load of CoQ10 nanometer suspending agent in rats blood drug concentration was significantly higher than that of CoQ10 technical, and the bioavailability of CoQ10 reversed-phased was improved 2.64 times.

    Extraction of Camptothecin and Hydroxyl Camptothecin from Camptothecin Leaves by Micellar Mediation Combined with Ultrasound-Microwave-Assisted Method
    Jian-Hang YANG, Xiu-Hua ZHAO
    2020, 40(6):  951-960.  doi:10.7525/j.issn.1673-5102.2020.06.018
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    In order to explore a new method to extract camptothecin and hydroxy-camptothecin from camptothecin leaves and make full use of camptothecin resources, the active ingredients of camptothecin and hydroxy-camptothecin from camptothecin leaves were extracted from camptothecin leaves powder as raw material. In this paper, the influence of microwave processing time, solid-liquid ratio, microwave power and the concentration of surfactant on the total extraction rate were investigated respectively, and the optimum extraction conditions were obtained. The results showed that the optimal extraction rate was achieved when the microwave processing time was 10 min 30 sec, the solid-liquid ratio was 1∶60, the microwave power was 850 W, and the concentration of surfactant was 8.5 g·L-1. The total extraction rate was 0.056 4% under the optimal process.