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    05 July 2019, Volume 39 Issue 4
    Effects of Different Light Qualities on Growth and Photosynthetic Characteristics of Mulberry Seedlings
    HU Ju-Wei, DAI Xin, SONG Tao, SUN Guang-Yu
    2019, 39(4):  481-489.  doi:10.7525/j.issn.1673-5102.2019.04.001
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    Light quality can influence the photosynthetic characteristics, morphology and physiological processes of seedlings. We studied the effects of different light qualities(white light, W; red light, R; blue light, B; a mixture of red and blue light, RB) using light emitting diodes(LEDs) on growth, morphology and photosynthesis of mulberry(Morus alba L. cv. Longsang No. 1) seedlings. The results showed that compared with seedlings grown under W, the dry matter accumulation of seedlings grown under R, B and RB was inhibited. The stem length and leaf area of seedlings grown under R were significantly larger than that of seedlings grown under W, RB and B. However, the leaf mass per area(LMA), chlorophyll a/b ratio, soluble protein content, sucrose and starch content, and total leaf nitrogen(N) content of seedlings grown under W, RB and B were significantly higher than that of seedlings grown under R. The net photosynthetic rate(Pn), stomatal conductance(Gs), and the actual photochemical efficiency of PSⅡ(ΦPSⅡ) of seedlings grown under RB treatment were similar to that of seedlings grown under W treatment, and the Pn and ΦPSⅡ of seedlings grown under R and B were lower than those of seedlings grown under W and RB. Antioxidant enzyme activity of seedlings grown under R, RB and B was higher than that of seedlings grown under W, yet the malondialdehyde(MDA) content of seedlings grown under R, RB and B was lower than that of seedlings grown under W. The number of leaf stomata on mulberry leaves grown under B was greater than all other treatments, and R-grown seedlings had the fewest stomata. Light quality also changed the leaf anatomical structure of mulberry seedlings. The leaf thickness, palisade tissue length and spongy tissue length of seedlings grown under R treatment significantly decreased. Hence, this study indicates that a certain ratio of mixed red and blue LEDs light could change the growth, photosynthetic characteristics, physiological characteristics, and leaf anatomy of mulberry seedlins, just like those of W-grown seedlings. Additionally, the mixed red and blue LEDs light reduced the adverse effects of monochromatic red and blue LEDs light on plant growth and development.
    Morphological and Physiological Responses of Populus ussuriensis Kom. to Drought Stress
    JIN Jiao-Jiao, SONG Zi-Wen, MA Xiao-Yu, SUN Guo-Yu, LI Kai-Long
    2019, 39(4):  490-496.  doi:10.7525/j.issn.1673-5102.2019.04.002
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    In order to explore the adaptability of Populus ussuriensis Kom. to drought stress, polyethylene glycol(PEG-6000) was used to simulate drought, the growth traits, biomass production and partitioning, the physiological and biochemical response characteristics were determined and analyzed under different stress levels. The seedling height, ground diameter and biomass accumulation of P. ussuriensis decreased significantly with the increase of drought stress intensity and the extension of stress time. The height growth under mild, moderate and severe stress decreased by 17.71%, 20.29% and 45.96%, the growth of ground diameter decreased by 4.78%, 13.62% and 17.29%, and the root ratio increased by 11.93%, 32.61% and 64.10%, respectively. The relative water content of the leaves showed a downward trend and the relative conductivity showed an upward trend. The relative water content under mild, moderate and severe stress was 23.60%, 38.56%, and 66.78% lower than the contrast, respectively, the relative conductivity was 106.97%, 161.84% and 241.75% higher than the contrast, indicating that drought stress caused obvious damage to the leaves. The contents of proline, soluble protein and soluble sugar all showed a trend of increasing first and then decreasing, which was higher than the contrast in the early and middle stages of stress, the change range of mild stress was smaller than that of moderate and severe stress. In the early and middle stages of stress, it was higher than the contrast and the seedlings had certain drought resistance. At the end of the stress period, there was a trend lower than the contrast, the seedling cells and tissues were irreversibly damaged. The net photosynthetic rate of leaves showed a double peak pattern, the stressed seedlings were significantly lower than the contrast and the time of reaching the peak was different. The first peak of the seedlings stressed was 2 h earlier than the contrast. P. ussuriensis can tolerate short-term mild drought stress environment. Moderate and severe drought stress have significant effects on the growth and physiological processes of P. ussuriensis seedlings.
    Physiological Mechanisms in Astragalus membranaceus Seedlings Responding to Cadmium Stress
    MA Yue-Hua, GUO Xiao-Rui, YANG Nan, ZHANG Ye, TANG Zhong-Hua, WANG Hong-Zheng
    2019, 39(4):  497-504.  doi:10.7525/j.issn.1673-5102.2019.04.003
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    The experiment was conducted to study the effects of different concentration Cd treatments on the growth, the adsorption and translocation of mineral elements, and the secondary metabolism of Astragalus membranaceus(Fisch.) Bunge(A. membranaceus), a traditional Chinese herb by employing a hydroponic system. The treatments of 25-200 μmol·L-1 Cd significantly inhibited the biomass growth and induced the increasing of malondialdehyde(MDA) content in whole plant and the decreasing of photosynthetic pigment contents in the new leaves, obviously exhibiting adverse effects. The Cd adsorbed by the plant dominantly accumulated in the roots. The greatest proportion of the shoots Cd was accumulated in its underpart, and orderly followed by the middle and the upper parts. It suggested that the seedlings took the advantage of Cd compartmentalization to minimize the adverse impact on the growing tissues of the shoots. Cd exposure hindered the adsorptions and translocations of a variety of mineral nutrition elements. However, the contents of Ca and Mg markedly enhanced by the Cd treatments in the upper and under shoots, respectively. We presumed that this might play positive roles in reducing the harms of Cd in the growing and old leaves. Furthermore, the contents of four isoflavonoids were elevated by the Cd treatments in roots of the seedlings. On the contrary, the contents of astragaloside Ⅰ-Ⅳ were depressed. It was implied that the isoflavonoids rather than astragalosides helped the seedlings of A. membranaceus to counteract Cd stress in their roots.
    Resistance Capability of Aquilaria sinensis under Combinations of Inorganic Salts,Hormones and Fungus Induction
    SONG Xiao-Chen, HUANG Gui-Hua, WANG Xi-Yang, SONG Jie, ZHANG Qing-Qing, LIANG Kun-Nan, ZHOU Zai-Zhi
    2019, 39(4):  505-513.  doi:10.7525/j.issn.1673-5102.2019.04.004
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    We explored the effect of different combinations of inorganic salts hormones and fungus induction on resistance capability of Aquilaria sinensis and the relationship between resistance and discoloration length of wood core in early stage of agarwood. Two induction experiments bydifferent combination of 3 kinds of inorganic salts, 3 hormones and 3 fungi were carried out for ten-years-old trees of A. sinensis by uniform experimental design. The results showed that:(1)POD and SOD activity and MDA content firstly increased and then decreased; CAT activity reached the highest value at 1 day after induction, and then decreased; (2)The resistance capability under combinations of inorganic salts and fungus induction was greater than that of hormones and fungus. The capability in treatment 4(1.0% CaCl2+0.5% FeSO4+2.0% NaCl+Trichoderma atroviride:Fusarium solani:Lasiodiplodia theobromae(1:1:1)) from combinations of inorganic salts and fungus and treatment 2(0.01% Methyl Jasmonate+0.1% Ethephon+0.2% Sodium salicylate+Trichoderma atroviride:Fusarium solani:Lasiodiplodia theobromae(1:1:1)) from combinations of hormones and fungus was higher in the experiments respectively; (3)There was a significant positive correlation between the resistance capability of A. sinensis and the length of wood core discoloration; (4)Theoretically, 0.93% CaCl2+0.53% FeSO4+2.5% NaCl+Trichoderma atroviride:Fusarium solani:Lasiodiplodia theobromae(1:1:1) and 0.005% Methyl Jasmonate+0.006% Ethephon+0.2% Sodium salicylate+Trichoderma atroviride:Fusarium solani:Lasiodiplodia theobromae(1:1:1) was combination with the highest resistance ability respectively.
    Effect of Forest-fire Rehabilitation Time on Plant Diversity in Daxing'an Mountains, Northeastern China
    YANG Yang, ZHANG Xi-Ting, XIAO Lu, YANG Yan-Bo, WANG Ke, DU Hong-Ju, ZHANG Jian-Yu, WANG Wen-Jie
    2019, 39(4):  514-520.  doi:10.7525/j.issn.1673-5102.2019.04.005
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    Forest fire is an important factor controlling forest ecosystem successions in the Daxing'an Mountains, Northeastern China, while long-term temporal dynamics in plant diversity and species abundance are still very limited to date, for modeling forest ecological services variations. We chose 48 pairs plots in different fire-rehabilitation years(1-5, 5-10, 10-20, 20-30, 30-40, and 40-50 years) in Daxing'an Mountains(fired site and neighbor unburned site as control), and diversity indices(Shannon-wiener index, Simpson index and species richness) and species abundance in tree, shrub and herb layers were calculated based on the field-surveyed data. Firstly,both fire-control differences in different parameters showed that tree diversity and richness declined at begin(lowest at 10-year rehabilitation), then increased afterward to the control-level after 30-40 fire-rehabilitation years. The shrub diversity index was similar to the change trend of trees, while most of the trends were statistically significant(P<0.05). The shrub Shannon index and richness indexlinearly increased with rehabilitationyears, andreached the control level after 30-year rehabilitation. The Simpson index of the herb decreased linearly with increasing fire age, but there was no linear change in uniformity and richness. Secondly, In terms of relative abundance of dominant species:In the arbor layer, Betula platyphylla in the fire siteduring 5-30 years rehabilitationtook >30% in proportion, and thereafter, the proportion was<15%; Larix gmelinii took over 50% in abundance in the 30-40 years rehabilitation stage. In the shrub layer, the largest proportion was Vaccinium vitis-idaea in less than 30-year rehabilitation stages, and the Corylus heterophylla be come the largest proportion. In the herbaceous layer,Deyeuxia angustifolia took the largest proportion in less than 30-year rehabilitation stages. In all, tree species recovered in a lower rate, and shrub and herb species recovered in a much faster rate. Plant diversity and dominant species alternation are basis for studying the forest ecological services(such as carbon sinks), and our data support future scientific management of the boreal forests in China, especially in the implementation of the Natural Forest Protection Program(a national policy of China).
    Expression Pattern of PagC3H3 Gene in Populus alba×P.glandulosa
    FAN Er-Qin, LIU Cai-Xia, FU Peng-Yue, YANG Chuan-Ping, QU Guan-Zheng
    2019, 39(4):  521-528.  doi:10.7525/j.issn.1673-5102.2019.04.006
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    As the main component of wood, lignin is usually polymerized from three monomers. During its biosynthesis, a total of ten enzyme families are involved in the conversion of phenylalanine to monomeric lignin, of which C3H is the key control enzymes in the hydroxylation process of p-coumaroyl CoA to caffeoyl CoA and in the formation of G/S monomer. Exploring the expression pattern of PagC3H3 gene is of great significance for further understanding of the function of this gene. In this study, the tissue-specific expression of PagC3H3 gene was analyzed by quantitative PCR; the promoter sequence of PagC3H3 with a length of 2 035 bp was cloned, and it was predicted to contain multiple cis-acting elements. At the same time, the promoter sequence of PagC3H3 was constructed to construct a plant expression vector pBI121-PagC3H3pro::GUS for transient transformation of Arabidopsis thaliana. The expression of PagC3H3 gene was higher in the roots, middle stem segments and basal stem segments of Populus alba×P. grandulos; transient transformation of A. thaliana showed that GUS activity was stronger in hypocotyls and roots, suggesting that PagC3H3 gene plays a role in lignin synthesis.
    Cloning and Expression Analysis of FmWRKY44 Gene in Fraxinus mandshurica
    LI Xin-Yu, HE Li-Ming, ZHAN Ya-Guang
    2019, 39(4):  529-538.  doi:10.7525/j.issn.1673-5102.2019.04.007
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    To clone the FmWRKY44 gene from Fraxinus mandshurica and explore its role in abiotic stress and hormone stress, specific primers were designed using the sequences of the variceal drought-transcriptome, the complete ORF sequence of FmWRKY44 gene was cloned, and bioinformatics analysis was carried out on the sequence and its coded products. The expression pattern of FmWRKY44 was analyzed using qRT-PCR. A WKRY gene of F. mandshurica was cloned, encoding 1 383 bp, encoding 460 amino acid. It was found to be a stable hydrophilic protein by the analysis of its encoded protein. The subcellular localization was mainly predicted in the nucleus, and the conserved domain and homology were analyzed. It belonged to the WRKY class I family and was named FmWRKY44. By qRT-PCR analysis, FmWRKY44 was highly expressed in the seeds. Different degrees of response to low temperature, high temperature, salt and drought four abiotic stresses. FmWRKY44 and NAA, ABA, GA3, JA phytohormone responses were also found. FmWRKY44 gene of F. mandshurica positively responds to low temperature and high temperature stress.
    Cloning and Expression Analysis of Catalase Gene(LoCAT1) from Larix olgensis
    BAI Xiao-Ming, DONG Shi-Wei, YANG Yu-Ning, SONG Yue, ZHANG Han-Guo, LI Shu-Juan
    2019, 39(4):  539-546.  doi:10.7525/j.issn.1673-5102.2019.04.008
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    In order to know about genes of Larix olgensis catalase and determine the expression characteristics in different organs of L. olgensis under different stresses, a CAT1 gene was cloned based on specific primers designed according to the CAT1 gene sequences from transcriptome database of L. olgensis, named LoCAT1. The complete length of open read frame(ORF) is 954 bp, encoding 317 amino acids. The results of phylogenetic tree analysis indicated that there was a closer relationship between LoCAT1 and Picea sitchensisand Ginkgo biloba. By real-time quantitative polymerase chain reaction (qPCR) analysis, LoCAT1 is highly expressed in leaf, while less expressed in stems. The expressions of LoCAT1 were obviously difference in roots and stems and leaves of L. olgensis under abiotic stress. The expression patterns of LoCAT1 were not exactly similarity among these treatments. The expression levels of LoCAT1 in roots and stems of L. olgensis after the treatment of NaCl were down-regulated, with the lowest expression at 12 h. While expression level in leaves was significantly inhibited at 24 h, and then up-regulated, with the highest expression at 96 h. The expression levels of LoCAT1 in roots and stems of L. olgensis after the treatment of PEG6000 were significantly inhibited in the early stage and then up-regulated. While LoCAT1 expression level in leaves was up-regulated. It was suggested that LoCAT1 may play an important role in response to stresses in plant.
    Cloning and Bioinformatics Analysis 4CL3/4CL5 Gene of Populus alba×P.glandulosa
    SUN Xiao-Sha, WANG Sui, ZHAO Xi-Yang, QU Guan-Zheng
    2019, 39(4):  547-556.  doi:10.7525/j.issn.1673-5102.2019.04.009
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    Lignin is an important component of wood, mainly for mechanical support, water transport and defense against pests and diseases. The 4CL enzyme is one of the key enzymes in controlling the pathway of lignin synthesis. The Pag4CL3/4CL5 gene was cloned from 84K poplar leaf cDNA by PCR. GenBank accession numbers are MK183033(Pag4CL3) and MK183034(Pag4CL5). The functions and characteristics of the two genes were analyzed using bioinformatics software. The similarity of the 4CL3/4CL5 proteins in 84K poplar and the 4CL3/4CL5 proteins in Populus trichocarpa were 97%,and they contained two conserved motifs of SSGTTGLPKGV and GEICIRG; subcellular localization showed the main localization of Pag4CL3/4CL5 proteins. In the endoplasmic reticulum, it may be a membrane protein and both are hydrophilic proteins. The expression of the Pag4CL3/4CL5 gene in different tissues was analyzed by qRT-PCR. The results showed that the Pag4CL3 gene was mainly expressed in leave and stem, and the expression level in root and top buds was low. The Pag4CL5 gene showed high expression level in leave and root, and low level in stem and apical buds. Pag4CL3 and Pag4CL5 may function together in the leaves, however, Pag4CL5 ismainly function in the root, and Pag4CL3 mainly function in the stems.
    Variation Analysis and Selection of Pinus koraiensis Half-sib Families
    ZHANG Qin-Hui, WANG Hong-Wu, JIANG Guo-Yun, SHEN Guang, WANG Lian-Kui, LI Yan-Long, WANG Lei, WANG Li-Xiang, LI Yue-Ji, LI Rui, ZHAO Xi-Yang
    2019, 39(4):  557-567.  doi:10.7525/j.issn.1673-5102.2019.04.010
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    In order to evaluate and select superior germplasm of Pinus koraiensis, with the 28-year-old 53 half-sib P. koraiensis families in Sanchazi forestry bureau, Jilin Province as materials, we measured the growth traits(tree height, basal diameter, diameter at breast height, diameter at 3 m, tree height under the first branch and tree height under the sixth branch) and shape traits(branch angle, stem straightness degree and branch number per node). Variance analysis showed that except for some of the shape traits, most of the growth traits showed strongly significant differences among the sources of variation(P<0.01). The phenotypic coefficient of variation of all the traits ranged in 6.97%-37.39%. The genotypic coefficient of variation of all the traits ranged in 1.76%-26.75%; The family heredity of all the traits ranged from 0.031-0.827; The single heredity of all the traits ranged in 0.019-0.864, lower heritability of a few traits. Correlation analysis results showed that there were significantly positive correlations(r>0.073) of the most traits except for branch angle. The results of general combining ability analysis showed that the families with high general combining ability of different traits had great differences, which was difficult to carry out combined screening and further analysis was needed. The results of principal component analysis showed that the cumulative contribution of the three principal components was 72.31%, which indicated that the three principal components contained most information of the growth and shape traits of the tested families. Tree height, basal diameter, diameter at breast height, diameter at 3 m height and volume contributed greatly to principal component Ⅰ, and described the growth characters of P. koraiensis, so it can be used as an evaluation index for selecting excellent families. Selection of family and single plant by multiple-trait comprehensive evaluation, based on the real gain of family volume over 35%, 5 excellent families were selected, and the selection rate was 10%, average tree height, basal diameter, diameter at breast height, diameter at 3 m height and volume were 7.90 m, 25.02 cm, 19.21 cm, 16.23 cm and 0.107 4 kg·m-3, and the real gains were 3.94%, 14.71%, 17.26%, 19.34% and 39.48%, respectively. Selecting single plants within the excellent families, according to the standard of genetic gain over 100% of single plant volume, 6 excellent individual plants were selected, and the selection rate was 4%, the tree height, basal diameter, diameter at breast height, diameter at 3 m height and volume mean values of the single plants were 1.60 m, 8.54 cm, 7.04 cm, 6.69 cm and 0.097 kg·m-3 higher than the total average, and the genetic gains were 5.99%, 29.92%, 35.53%, 37.06% and 102.04%, respectively. The selected excellent families and individuals can provide the basis for validation of improved varieties and provide materials for the construction or reconstruction of the orchard.
    Genome-Wide Analysis of the WOX Family Reveals Their Involvement in Stem Growth of Populus trichocarpa
    WANG Shuang, ZHANG Yang, REN Meng-Xuan, LIU Ying-Ying, WEI Zhi-Gang
    2019, 39(4):  568-577.  doi:10.7525/j.issn.1673-5102.2019.04.011
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    The WUSCHEL-related homeobox(WOX) family is a plant-specific transcription factor family involved in multiple developmental processes including meristematic cell division and differentiation, primary and secondary biomass metabolism, and plant hormone signaling. At present, there is no relevant research on genome-wide analysis of the gene family involved in stem development of poplar. We identified the poplar WOX gene family, and found 18 WOX candidate genes in the poplar gene. These candidate genes were divided into three groups. Most of the WOX family members in the same group had similar gene structure and conservation motif. According to the transcriptome data of stems at different developmental stages, the specific expression of WOX family members in different developmental stages of stems was systematically analyzed, and the above results were verified by qRT-PCR. The poplar WOX gene family showed different expression patterns at different developmental stages of the stem, which laid a foundation for the functional research and utilization of the WOX family.
    Characteristics and Coupling Relations of Green Space in Major Areas of Harbin
    GUAN Yue-Yuan, ZHU Si-Yu, YI Zhao-Qin, WU Yan, MU Li-Qiang
    2019, 39(4):  578-589.  doi:10.7525/j.issn.1673-5102.2019.04.012
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    The plant diversity of 17 green spaces in Harbin was investigated by sample method, and the influence of native plant diversity on alien plants in urban green space ecosystem was studied. There are 75 alien plants, 42 alien cultivated plants and 33 alien non-cultivated plants in green space of Harbin. In the greenbelt plant community of Harbin, the average coverage, average importance and average frequency of exotic cultivated plants are higher, while the average coverage and average importance of exotic non-cultivated species are lower, but the average frequency of exotic non-cultivated species is higher. The number of native plant species was positively correlated with the number of exotic cultivated species and exotic non-cultivated species(P<0.01), and negatively correlated with the average frequency of exotic non-cultivated species(P<0.05). The Shannon-Will index and species richness of native plants in the arbor layer of green space were negatively correlated with the number of alien cultivated species and non-cultivated species(P<0.05). The homogeneity of native plants in the arbor layer was negatively correlated with the average frequency of non-cultivated species(P<0.01). The species richness of native plants in the shrub layer was negatively correlated with the average importance of exotic cultivated species(P<0.05); the evenness of native plants in herbaceous layer was positively correlated with the average relative coverage of non-cultivated species(P<0.05). With the increase of the total area of green space, the number of exotic cultivated species and exotic non-cultivated species increased, and the average frequency decreased significantly. With the increase of construction time, the average frequency of exotic cultivated species did not change linearly, while the average frequency of exotic non-cultivated species showed a decreasing trend. We should improve the diversity of native plants in Harbin green space, strictly control the introduction and cultivation management of alien plants in newly built green space with a small total area, in order to prevent the development of alien plants into invasive plants that endanger the local ecosystem.
    Spatial Heterogeneity of Urban Street Trees in Harbin,China
    LIU Xiao, ZHOU Wei, WANG Wen-Jie
    2019, 39(4):  590-597.  doi:10.7525/j.issn.1673-5102.2019.04.013
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    Spatial heterogeneity analysis of urban street trees is the basis for optimizing the surveying method and scientificmanagementandimproving the ecological service of urban forest. In order to provide basis for analyzing of reasonable sampling sites of urban forest, the internet street view picture-based remote measurements were used to determine canopy size[CS](average width),under branch height[UBH], tree height[TH] and diameter at breast height[DBH] in a total of 879 plots(26 140 trees), combining with the semivariogram analysis and fractal analysis, to characterize the spatial heterogeneity of urban treesin Harbin. By anisotropy analysis,the insignificant differences in the spatial heterogeneity of CS, UBH, TH, DBH in Harbin city were found at 0°, 45°, 90°,amd 135° directions. As shown by the Nugget/base ratio, random effects accounted for 26.56% of spatial heterogeneity of CS, while much less random effects were responsible for the spatial heterogeneity of UBH, TH, DBH(8.16%, 8.14%, and 12.80% of the total heterogeneity, respectively). Thus, the results suggested that the spatial variations of UBH, TH and DBH were likely to be mainly due to the urbanization-related structural factors(such as urban planning etc.) (>88%). The results indicated that spatial heterogeneityof urban tree varied with different tree size parameters, showing significant difference by the self-correlation length(semi-variogram range), i. e. the semi-variogram range values for CS, UBH, TH, DBH were 2 340, 1 320, 1 470 and 1 890 m, respectively. For precisely identifying thespatial heterogeneity of tree size, sampling interval should be within the semi-variogram range. Our data showed the minimum sampling numbers for measuring the targeted indicators are 156 plots for the CS, 440 plots for the UBH, 360 plots for the TH and 224 plots for the DBH. Fractal analysis showed that the fractal values(D value) of CS, UBH, TH, DBH were 1.957, 1.961, 1.961 and 1.971, respectively. The similar fractal values of the targeted indicators, which were close to 2, could suggest that that their spatial variations were mainly observed in smaller scales between neighboring plots(distance:666m). The results of this study provide basic data to support the further research and scientific management ofthe urban forest in Harbin City, as well as to determine reasonable sampling plans in the future study.
    Effects of Long-term Nitrogen Addition on Soil Enzymes in Larix gmelinii Plantation in Northeast China
    MA Peng-Yu, ZHANG Hong-Guang, ZAN Peng, GU Wei-Ping, WEN Lu-Ning, ZHANG Zi-Jia, WENG Hai-Long, SUN Tao, MAO Zi-Jun
    2019, 39(4):  598-603.  doi:10.7525/j.issn.1673-5102.2019.04.014
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    The increase of nitrogen deposition hadbecame a significant ecological factor effecting the material cycle in forest. We examined the effects of nitrogen deposition on soil enzyme activities in inorganic nitrogen addition and control plots in Larix gmelinii(Rupr.) Rupr. plantation, in northeast China. The experiment was divided into two treatment groups, one was control treatment(CK:0 kg·N·hm-2·a-1) and another was inorganic nitrogen treatment(IN:10 kg·N·hm-2·a-1). Each treatment included 3 replicate plots. Over 8 years of inorganic nitrogen addition, our results showed that N fertilization increased the activities of β-1, 4-glucosidase(BG), acid phosphatase(AP), and peroxidase(PER)(5.56%, 8.66% and 0.61%)while reduced the activities of cellobiohydrolase(CBH), leucine aminopeptidase(LAP) and phenol oxidases(POX)(10.71%, 3.85% and 12.30%). The long-term inorganic nitrogen addition treatment had a significant effect on soil enzyme activity and soil carbon, nitrogen and phosphorus cycles(0.80%, 12.66% and -2.25%). Therefore, it is important to include the soil enzyme activity when studying the response of biogeochemical cycle and soil carbon dynamics to climate changein forest ecosystems in future research, while nitrogen deposition increases year by year.
    Dissolution, Bioavailability and Tissue Distribution of Artemisinin Microspheres Loaded With Porous Starch
    ZHAO Xue, YANG Feng-Jian, GE Yun-Long, WANG Ling-Ling, LI Wen-Gang, DENG Yi-Ping, ZHAO Xiu-Hua
    2019, 39(4):  604-612.  doi:10.7525/j.issn.1673-5102.2019.04.015
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    To investigate the dissolution effect of artemisinin microspheres(ART-PS) loaded with porous starch in vitro compared with artemisinin, bioavailability and tissue distribution in rats, in the in vitro dissolution experiment, the dissolution effect of artemisinin was compared with that of artemisinin in water, artificial gastric juice and artificial intestinal fluid. In the in vivo bioavailability experiment, 18 rats were given artemisinin raw drug and porous starch loaded artemisinin microspheres respectively, and the blood drug concentrations at different time points were detected to investigate the differences in drug absorption and metabolism in rats. In the tissue distribution, the drug contents in heart, liver, spleen, lung, kidney and brain of 98 rats were detected at different points in time by gavage of porous starch loaded artemisinin microspheres and artemisinin active drugs. The dissolution rates of artemisinin microspheres loaded with porous starch in vitro were 4.04, 3.59 and 3.82 times higher than those of artemisinin in water, artificial gastric juice and artificial intestinal fluid, respectively. The blood content of artemisinin microspheres loaded with porous starch in rats was significantly higher than that of artemisinin, and the bioavailability was 2.90 times higher than that of artemisinin. In the results of tissue distribution, artemisinin microspheres loaded with porous starch and artemisinin active drugs were mainly distributed in the heart and liver, and the corresponding content of artemisinin active starch active microspheres in different tissues at different times was higher than that of active drugs. Compared with artemisinin, artemisinin microspheres loaded with porous starch have better dissolution effect in vitro and better absorption and release effect in vivo. The drug content in various tissues and organs is significantly higher than that of the original drug, providing an important theoretical basis for solving the limitation of artemisinin in practical application due to its insoluble in water.
    Cryo-sectioning Technique for Microstructure Studies on Dendrobium officinale
    JIANG Su-Hua, LIANG Fang, ZHANG Yan, ZHOU Yi-Ran, GUO Meng-Yuan, YUAN Xiu-Yun, CUI Bo
    2019, 39(4):  613-618.  doi:10.7525/j.issn.1673-5102.2019.04.016
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    In order to observe the microscopic structure of Dendrobium officinale quickly and efficiently. This study used the light microscope and the improved sucrose protection-liquid nitrogen quick-freezing freezing section method for orchid tissues and organs, Which improved sucrose protection-liquid nitrogen frozen method, observed root, stem and leaf of microscopic structure on D. officinale. The technology and methods treated D. officinale with sucrose phosphate buffer protection fluid,and then Vacuum processing. By liquid nitrogen frozen, embedding, sectioning, exhibition slice observation, staining and photo produced D. officinale roots, stems and leaves a complete microscopic section. The results show that the optimal condition for root was 8% sucrose mass volume fraction, condensation temperature -25℃, slice thickness 20 μm; the optimal condition for stem was 16% sucrose mass volume fraction, frozen temperature -20℃ and slice thickness 15 μm; the optimal condition for leaf was 4% sucrose mass volume fraction, frozen temperature -20℃ and slice thickness 10 μm. The study on the Orchidaceae microstructure observation and histochemical study will have broad prospect of application.
    Extraction of Perilla Seed Oil by Aqueous Enzymatic Method
    YUAN De-Cheng, WANG Fei, CUI Xin-Shuang, LI Xiao-Xue, YANG Feng-Jian
    2019, 39(4):  619-626.  doi:10.7525/j.issn.1673-5102.2019.04.017
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    Central composite design(CCD)-response surface(RSM) was used to optimize the aqueous enzymatic extraction process of Perilla frutescens seed oil. On the basis of single factor experiment, we adopted the central composite design method to study the effects of enzymatic species, enzymatic hydrolysis temperature, pH, liquid-solid ratio, enzyme dosage and time interaction on the extraction rate of Perilla oil. The fitting equation was significant, the optimum conditions for the extraction of Perilla oil were as follows:pH is 9.5 in alkaline protease, the solid-liquid ratio(g) was 9.97:1, the amount of enzyme added was 2.75%, the temperature was 56.1℃, and the time was 5.25 h. Under this condition, the extraction rate of Perilla oil could reach 37.65%, which was very close to the theoretical value of 38.3%. The established model was true and reliable, and the optimum extraction process of Perilla oil was determined. The Perilla seed oil contains palmitic acid, stearic acid, oleic acid, linoleic acid and alpha-linolenic acid by gas chromatography, the relative content of alpha-linolenic acid in Perilla oil extracted by aqueous enzymatic method was 67.9%, and the physical and chemical indexes of relative solvent method and cold pressing method were the best.
    Formulation and Process Optimization of Paclitaxel-loaded PLGA Nanoparticles
    WANG Si-Ying, ZHAO Xiu-Hua, ZHAO Dong-Mei
    2019, 39(4):  627-633.  doi:10.7525/j.issn.1673-5102.2019.04.018
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    Paclitaxel-loaded PLGA nanoparticles(PTX-PLGA NPs) was prepared by emulsion solvent evaporation method with the biodegradable materials-poly(lactic-co-glycolic acid)(PLGA) used as carrier materials. The best formulation and preparation conditions were optimized by orthogonal test, investigating the influence of the ratio of paclitaxel and PLGA, the amount of surfactant, the amount of dichloromethane(DCM) and shearing rate on the preparation of nanoparticles. The optimal preparation conditions for PTX-PLGA NPs were as follows:the ratio of paclitaxel and PLGA of 4:1, the amount of surfactant of 0.1%, the amount of dichloromethane of 4 mL and shearing rate of 16 000 r·min-1. With optimization of the formulation and preparation conditions, the PTX-PLGA NPs are regularly spherical in shape with the mean size of (207.1±20.5) nm, and the ζ-potential of the NPs is about (-23.8±2.5) mV. The encapsulation efficiency was estimated to be 14.45%±0.04%, respectively. The size distribution of PTX-PLGA NPs frozen dry powder was sustained stable character in 12 h. The results indicate that PTX-PLGA NPs were successfully prepared by modified solvent evaporation method, which could provide the experimental basis for the development of a new anti-tumor sustained-release formulation.
    Purification and Lipid-lowering Activity Evaluation of Pinoleic Acid in Korean Pine Seed Oil
    ZHANG Jing, KOU Ping, QING Qi-Ping, YU Liang, ZHAO Xue-Lian, WANG Xi-Qing, YANG Yu-Chun, FU Yu-Jie
    2019, 39(4):  634-640.  doi:10.7525/j.issn.1673-5102.2019.04.019
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    With Korean pine seed oil we enriched and purifiedpinoleic acid by urea inclusion. Through the influence of the ratio of fatty acid to urea, the ratio of urea to ethanol, we optimized the inclusion temperature and the inclusion time on the content of pinoleic acid and recovery of pinoleic acid, the process parameters of urea bagging. Finally, the optimal purification condition of pinoleic acid was determined as follows:the ratio of fatty acid to urea was 1:6.5(w/w); the ratio of urea to 90% ethanol was 1:4(w/v); Inclusion temperature was -10℃; Inclusion time was 12 h. Under the above conditions, a purity of 58.3% pinoleic acid could be obtained, and the recovery rate was 51.8%. After purification by secondary urea inclusion, a purity of 72.5% of pinoleic acid could be obtained, and the recovery rate was 64.7%. To verify the lipid-lowering activity of pinoleic acid, oleic acid was used to induce lipogenesis in HepG2 cells. The results showed 40 μmol·L-1 pinoleic acid could significantly reduce the level of triglycerides and total cholesterol in HepG2 cells.