Cell Ultrastructure of Kiwifruit （Actinidia chinensis） Shoot Tips During Cryopreservation

The changes in the cell ultrastructure of in vitro cultured shoot tips from dwarf genotype of kiwifruit （Actinidia chinensis Ganmi 5） during cryopreservation were investigated. Shoot tips were preserved in liquid nitrogen using vitrification, and the cell ultrastructure was examined using transmission electron microscopy （TEM）. The regular ultrastructure of the cell wall, cell membrane and nucleus of shoot tips could be damaged during the freezing and thawing associated with preservation using liquid nitrogen. The cell plasmolysis was increased and freezing tolerance was improved after precultufing and dehydrating in a preservation and vitrification solution （PVS2） （30% glycerol （Gly）＋ 15% ethylene glycol （EG）＋ 15% dimethylsulfoxide （DMSO） ＋ 0.4 mol L^-1 sucrose）. The structure of some cells with low degree of injury and reversible damage was similar to that of the control and they could undergo normal cell division and differentiation. Besides, they could recover automatically and regenerate after their reculture.     

Effects of Wounding and Exogenous Jasmonic Acid on the Peroxidation of Membrane Lipid in Pea Seedlings Leaves

The changes of malondialdehyde （MDA）, H2O2, and O2^7 content, or the activities of superoxide dismutase （SOD）, catalase （CAT）, ascrobate peroxidase （APX）, peroxidase （POD）, phenylalanine ammonia lyase （PAL）, and polyphenol oxidase （PPO） in pea seedlings （Pisum sativum L.） under wounding and treatment of exogenous jasmonic acid （JA） were investigated. The results showed that the activities of both phenylalanine ammonia lyase （PAL） and polyphenol oxidase （PPO） were significantly increased by wounding and application of JA. The metabolism of reaction oxidative species （ROS） was enhanced, especially O2^7 and H2O2 appeared to rapidly increase. The activities of antioxidant enzymes such as SOD, CAT, APX and POD were also increased. Treatment of JA of 1 or 10 μmol L^-1 could effectively induce plant defense response, and thus decrease the peroxidation of cell membrane lipid. However, high concentration of JA （100 μmol L^-1） resulted in unbalance of metabolism of ROS and promoted the peroxidation of cell membrane lipid. We thus suggested that JA, under the suitable concentration, could induce defense response of pea seedlings to wounding.     

Analysis on Membrane Stability of Crown in Winter Wheat in Frigid Region in China

Four varieties of winter wheat with different return green rates were used to analyze the plasma membrane stability and the factors that affect winter wheat in the frigid region during winter. The removal of reactive oxygen species, the degree of plasma membrane impairment, water composition and content, and the changes in cell viability in the crowns, which contain the growing point, were studied during the period from cold acclimation to the deep freezing. The results showed that electrical conductivity which reflects the degree of plasma membrane damaged under low temperature was significantly correlated with the free water and the total water content. The malondialdehyde (MDA) content, which reflects the degree of membrane peroxidation, was very significantly correlated with superoxide dismutase (SOD), peroxidase, and ascorbic acid. During the deep freezing period, the SOD activity and glutathione (GSH) content of the winter wheat varieties were relative to their cold resistance. During this period, the MDA stability, SOD, GSH, and the total water and the free water content might be used to identify the cold resistance of winter wheat varieties.     

Protective Effect of Wheat Peptides Against Small Intestinal Damage Induced by Non-Steroidal Anti-Inflammatory Drugs in Rats

Non-steroidal anti-inflammatory drugs（NSAIDs） were able to produce tissue damage and oxidative stress in animal models of small intestinal damage. In this study, the putative protective effect of wheat peptides was evaluated in a NSAID-induced small intestinal damage model in rats, different doses of wheat peptides or distilled water were administered daily by intragastric administration for 30 d until small intestinal damage was caused. Before sacrificing, NSAIDs（aspirin and indomethacin） or physiological saline were infused into the digestive tract twice. Wheat peptides administration reduced edema and small intestinal damage, and significantly decreased the level of tumor necrosis factor（TNF）-α in mucous membrane of small intestine. Oxidative stress was significantly increased after NSAID infusion and was reduced by wheat peptides. Wheat peptides increased glutathione peroxidase（GSH-Px） activity in mucous membrane of small intestine. μ-Opioid receptor mRNA expression decreased more significantly in wheat peptides treated rats than in the model control group. Overall, the results suggest that non-steroidal anti-inflammatory drugs induced small intestinal damage in rats and wheat peptides administration may be an effective tool for protecting small intestinal tissue against NSAID-induced small intestinal damage and oxidative stress.     

Design and Experiments of the High Voltage Pulsed Electric Fields Sterilization System

This experiment designed the pulsed electric fields （PEF） of high intensity of 100 kV· cm^-1 sterilization system. Fluorescent pseudomonas as target cell was operated 180 s in the PEF. By observing the difference of the bacteria before and after the disposal by TEM, it is found that the cell walls of the treated bacteria were broken. Irreversible perforations were formed on the cell membrane. The cell inclusions and cell fragments were leaked. The cell died as a result. The results showed that the PEF sterilization system designed can be used for liquid food sterilization experiments.     

Effects of Pretreatment by the Flavanol Ampelopsin on Porcine Kidney Epithelial Cell Injury Induced by Hydrogen Peroxide

This study investigated the protective effects of ampelopsin on H2O2-induced oxidative injury in porcine kidney epithelial cell line, PK-15; cells were pretreated with medium containing 0, 15, 30, and 60 ug mL-1 ampelopsin, respectively, for 1 h prior to the addition of 100 umol L-1 H2O2. After 2 h, the cell viability, intracellular superoxide dismutase （SOD） and glutathione peroxidase （GSH-Px） activity, cellular malondialdehyde （MDA） content, and DNA strand breakage were determined. Results showed that ampelopsin pretreatment did not modify the SOD and GSH-Px inactivation but significantly decreased MDA production; except for the medium dose （30 ug mL-1）, all the tested concentrations of ampelopsin did not significantly inhibit cell viability reduction and gave no influence on DNA damage. The results suggested that all the tested doses of ampelopsin pretreatment before H2O2 addition can protect PK-15 cell from H2O2-induced lipid peroxidation, which might not be mediated by the SOD and GSH-Px responsees. The appropriate dose of ampelopsin pretreatment prevents PK-15 cell from DNA damage and cell viability reduction.     

Relationship Between Anthocyanin Biosynthesis and Related Enzymes Activity in Pyrus pyrifolia Mantianhong and Its Bud Sports Aoguan

The aim of this article is to study the relationship between biosynthesis of anthocyanin and activities of phenylalanine ammonia lyase （PAL）, chalcone ismoerase （CHI） enzymes in Pyrus pyrifolia. Changes in the level of anthocyanin and the activities of enzymes of anthocyanin biosynthesis including PAL, CHI were studied in the pericarp of Pyrus pyrifolia Aoguan and Mantianhong during the period of pigment formation. Bagging treatment was also carried out to manipulate the synthesis of anthocyanin and the activities of related enzymes during the period of pigment formation. The results demonstrated that the level of anthocyanin of Aoguan was higher than that of Mantianhong. However, the content of anthocyanins has the similar changing trend in Aoguan and Mantianhong, highest anthocyanin concentrations of two varieties appeared in immature fruit and faded toward harvest. Meanwhile, similar changing trends of activities of PAL and CHI were also observed in both varieties. Aoguan has a lower activity of PAL than Mantianhong, whereas activity of CHI in Aoguan was higher than that in Mantianhong. Activity of PAL decreased during the period of pigment formation and was apparently not limiting to color development, whereas CHI activity increased at the same period and was closely related to the synthesis of anthocyanin. The results of bagging treatment showed that bagging treatment inhibited the activity of CHI, as well as the synthesis of anthocyanin, whereas debagging enhanced both the activity of CHI and synthesis of anthocyanin. The activity of CHI in debagging Aoguan pericarp was higher than the untreated Aoguan. However, effect of bagging treatment toward PAL activity was not obvious. Anthocyanin of bagging treated Aoguan decreased toward harvest. The content of anthocyanin of Pyruspyrifolia increased at the beginning of fruit coloration period and decreased toward fruit harvest. Activity of PAL was apparently not limiting to color development, whereas CHI activity was closely related to the synthesis of ant     

Physiological Changes and Cold Tolerance of Three Camphor Species During Natural Winter Temperature Fluctuations

The dynamic changes in the malondialdehyde（MDA）, superoxide dismutase（SOD）, soluble sugar, proline, and soluble protein contents, as well as the relative electrolyte conductivity and the corresponding cold resistance, of Cinnamomum bodinieri Level., C. camphora L., and C. caudiferum Kisterm were investigated during the winter months of October 2009 to April 2010. During the short period of temperature decline that lasted until mid-December, the changes in the relative electrolyte conductivity and MDA content with temperature were insignificant. In January, SOD activity continued to increase and then peaked as a result of rapid increases in soluble sugar, proline, soluble protein, as well as the inhibition of the relative electrolyte conductivity and decrease in MDA content. These physiological changes protected the camphor trees from cold damage during winter. From February to March, SOD activity and the soluble protein and proline contents increased with the increase in temperature. However, the relative electrolyte conductivity and MDA content decreased, indicating that the cell membrane damaged by low temperature was gradually being repaired. The cold dip in April led to slight increases in the relative electrolyte conductivity and MDA content. Using a fuzzy mathematics method, the cold resistance adaptability of the camphor trees was divided into three periods namely, the enhancement setting stage, the vigorous stage, and the reducing stage. The cold tolerance abilities were ranked as the following order： C. bodinieri Level〉C. camphora L.〉C. caudiferum Kisterm.     

Study on Active Oxygen Quantum Yield, Insecticidal Activities and Stability of Diphenylthiophene

Active oxygen quantum yield, insecticidal activities and stability of diphenylthiophene were studied for the first time. The results showed that maximum diphenylthiophene absorbency variety was 0.438 after irradiation for 280 min and that of α-terthienyl (α-T) was 0.480 after irradiation for 200 min with UV. LC50 values ofdiphenylthiophene and α-T against 3rd instar larvae of Aedes albopictus were 9.18 × 10-3 and 9.69 × 10-4 μg mL-1 when treated for 24 h, respectively. LC50 values of the two chemicals against the 3rd instar larvae of Plutella xylostella were 267.87 and 222.22 μg mL-1 when treated for 24 h, respectively. The half lives of diphenylthiophene and α-T in methanol were 113.62 and 10.65 h. Difference between quantum yield of diphenylthiophene and α-T was not significant and they all possessed high toxicity to Aedes albopictus and Plutella xylostella, but diphenylthiophene was more stable than α-T. It could be concluded that diphenylthiophene has overcome the deficiency of photoactivated insecticides which degraded quickly in the environment and could not be applied on the field. Diphenylthiophene could kill the insects and accelerate the degradation rate of triazophos. The benefits of diphenylthiophene can be further exploited and applied on the field.     

Oxidative Damage to Spodoptera litura Cell Induced by α-Terthienyl

In this study, the oxidative damage of α-terthienyl (α-T) to the Spodoptera litura (SL) cell and its mechanism were investigated. MTT was used to compare the toxicity of α-T and rotenone to the SL cell. The output of malondialdehyde and relative content of glutathione were determined with 2-thiobarbituric acid (TBA) and 5, 5'-dithio-bis (2-nitrobenzoic acid) (DTNB), respectively. Transmission electron microscope (TEM) was employed to observe the influence of α-T on the membrane and organelle of the SL cell. The result showed that the IC50 value of α-T to the SL cell was 0.21 μg mL-1, whereas the corresponding dose of rotenone was 12.25 μg mL-1. The output of MDA had the same changing tendency with the concentration of α-T, whereas the content of GSH had the negative correlation with it. According to TEM, the cell membrane and karyotheca swelled and couldn't retain integrity, the intracellular substances leaked out, unidentified granules appeared in the SL cell. The mitochondria expanded, and the membrane and subcellular organelle were damaged severely.In this study, it was found that after oxidative damage induced by α-T, the output of MDA increased notably, whereas the relative content of GSH decreased. This indicated that the antioxidant ability of cell weakened. The result of TEM implied that the SL cell suffered from oxidative damage under the appointed dose.     

α-三联噻吩致斜纹夜蛾SL细胞氧化损伤的研究

 【目的】研究光敏剂α-三联噻吩（α-terthienyl，α-T）对斜纹夜蛾（Spodoptera litura，SL）细胞的毒杀活性及其机理。【方法】MTT法测定α-T对离体培养的SL细胞的细胞毒力，并与鱼藤酮进行比较。用硫代巴比妥酸（TBA）法测定细胞内丙二醛（MDA）的生成量，5,5′-二硫代双-（2-硝基）苯甲酸（DTNB）法测定细胞内还原型谷胱甘肽（GSH）的相对含量，通过透射电子显微镜观察α-T对SL生物膜及细胞器的影响。【结果】光活化后的α-T对SL细胞48h的IC50值为0.21μg•ml-1，而鱼藤酮为12.25μg•ml-1。同时发现α-T光照处理后细胞内MDA生成量与药剂浓度呈正相关，表明细胞受到α-T的氧化损伤，而GSH相对含量与α-T浓度呈负相关。透射电镜显示，α-T光照处理后SL细胞膜和核膜缺损、肿胀，内容物外泻，胞内出现不明深色颗粒，线粒体膨大，膜和亚细胞器结构受到严重破坏。【结论】SL细胞受到α-T的氧化损伤后，MDA生成量显著增加，GSH含量下降，细胞的抗氧化能力减弱，透射电镜结果显示SL细胞在α-T的试验剂量下受到了严重的氧化损伤。     

Antifeedant Activity and Active Ingredients Against Plutella xylostella from Momordica charantia Leaves

With the bioguided fractionation of the ethanol extracts from the leaves of Momordica charantia, we obtained two most active compounds against the feeding of the diamondback moth, Plutella xylostella larvae. The antifeedant activity of momordicine Ⅰ and momordicine Ⅱ against the second and the third instar larvae of Plutella xylostella were tested using leaf discs of cabbage in the laboratory. The results showed that momordicin Ⅰ and momordicin Ⅱ had significant antifeedant activity on the larvae of P. xylostella, and momordicin Ⅱ was more active than momordicin Ⅰ. The concentrations for 50% antifeedant effects （AFC50） of momordicin Ⅱ against the second and the third instar larvae of P. xylostella were 76.69 and 116.24μg mL^-1, whereas that of momordicin I was 144.08 and 168.42μg mL^-1, respectively. In addition, momordicin Ⅰ and momordicin Ⅱ had significant inhibitive effect on the rate of weight gain and survival of P. xylostella larvae.     

Sensitivities of Phytophthora infestans to Metalaxyl, Cymoxanil, and Dimethomorph

The isolates of Phytophthora infestans on tomato in the Guangxi Zhuang Autonomous Region, China, were determined for the sensitivities to metalaxyl, cymoxanil and dimethomorph to give the basic information for integrating disease management. Sensitivities were tested by measuring the radial growth on agar medium amended with fungicide, compared with the floating-leaf-disk method. 239 isolates were collected from eight tomato growing areas during 2000-2006. The testing results indicated that the frequencies of sensitive, intermediate, and resistant isolates to metalaxyl were 42.26, 35.98, and 23.53%, respectively. Variations in sensitivities amongst isolates from different areas or different years were very high for metalaxyl. All isolates from Tianlin and Wuxuan were sensitive to metalaxyl, but the metalaxyl-resistant isolates predominated in Tianyang, with the frequency of 51.35%. The EC50 values of certain isolates from Tianyang were higher than 500 μg·mL^-1 and their resistance levels were over 100 000 folds. Cymoxanil has been used for nearly 10 years in Guangxi, and dimethomorph has been used for 5-6 years. However, there was no decrease in sensitivity of P. infestans populations and the sensitivities of the pathogen were nearly normally distributed. Hence, their mean ECs0 value [cymoxanil （0.1647 ±0.0255） μg·mL^-1, dimethomorph （0.0970 ± 0.0052） μg·mL^-1 could be used as the baseline sensitivities for monitoring the field resistance development. The comparison with the floating-leaf-disk method indicates that both the techniques provided equivalent results. These studies suggested that metalaxyl can be continuously applied in Tianlin, Wuxuan, and Nanning due to the resistant isolates that have not been found, while for those areas with resistant isolate, the use of metalaxyl should be reduced or alternated, and cymoxanil or dimethomorph was recommended for controlling late blight disease of tomato.     

AFB1 Bio-Degradation by a New Strain - Stenotrophomonas. sp

The paper was to find the bacteria to degrade aflatoxin B 1 （AFB 1） and realize the application of biological degradation on AFB 1. Using cumarin as the carbon source and energy on the first screening, then the ten strains which were first screened out were taken to degrade AFB 1 100 pg kg^-1. Strain NMO-3 was screened out of ten strains, the degradation ratio of AFB 1 reached 85.7%, which was more prominent than the others （P 〈 0.01）. With the analysis of colony morphology, physiological and biochemistry experiments, and 16S rDNA gene sequence, the strain NMO-3 was finally identified as Stenotrophomonas sp. Using cumarin as the carbon source and energy could screen out the AFB 1 degradation strains. Acute toxicity tests show that the viable number of NMO-3 lower than 3.12 × 10^10 cfu mL-1 is safety. The crude enzyme was obtained by 65% ammonium sulfate fractionation, and it could degrade AFB1. It is the first report for the strain＇s detoxi- AFB1.     

Insecticidal Constructure and Bioactivities of Compounds from Ficus sarmentosa var. henryi

Insecticidal activities of the petroleum ether-, chloroform-, ethyl acetate-, and water-soluble fractions of the methanolic extract of Ficus sarmentosa var. henryi were assayed against Musca domestica adults. The chloroform- and ethyl acetate-soluble fractions were the most active with 92.6 and 88.9% mortalities (24 h after treatment) respectively. Therefore, the two fractions were combined and four compounds, isolated from the fractions by activity-guided fractionation, were elucidated as 7-hydroxycoumarin, apigenin, eriodictyol, and quercetin by spectroscopic method and displayed excellent insecticidal activities against adults of M. domestica and 4th instar larva of Aedes albopictus. Among those, 7-hydroxycoumarin showed the strongest insecticidal activities with lethal concentrations (LC₅₀) values of 72.13 μg g⁻¹ sugar and 4.87 μg mL⁻¹ (48 h after treatment) against the test insects respectively. The cytoxicities of these compounds on BTI-Tn-5Bl-4 cell were also investigated for the insecticidal mechanism and found that quercetin represented superior inhibitory activity with MTT assay and reactive oxygen species (ROS) against BTI-Tn-5Bl-4 cell, but slightly weaker than that of the positive control (azadirachtin) and significantly greater than the negative control (DMSO only). Meanwhile, eriodictyol demonstrated the strongest effect on the mitochondrial membrane potentials (MMP). In conclusion, based on their comparative toxicities to commercial insecticides and their cytotoxic effects, some of the compounds from the F. sarmentosa have potential as botanical insecticides.     

3种化感作物水浸液对远志种苗生长的影响

用室内生物测定的方法测试3种化感植物（向日葵、高粱、玉米）叶片的不同质量浓度水浸液（0、 0.02 、0.025、0.033、0.05、0.1 g·mL^-1）对远志种子萌发和幼苗生长的影响。结果表明,3种水浸液在试验质量浓度范围内均对远志种子的发芽有低促高抑的化感效应,且质量浓度越高抑制效果越明显;3种供体材料的化感效应存在明显的供体植物种间差异,其抑制作用强弱表现为高粱>向日葵>玉米,对远志的最高安全质量浓度依次是0.025、0.033和0.1 g·mL^-1。其中玉米水浸液在质量浓度≤0.1 g·mL^-1时对远志的发芽率和发芽指数都是安全的,而且在最低质量浓度（0.02 g·mL^-1）的综合效应值为+0.004,表现为促进作用。因此,建议将玉米废弃秸秆作为首选材料进一步测试其废弃植株粉碎物对远志田杂草是否有抑制效果,以期为开发安全防治远志田杂草的植物源“除草剂”提供基础。     

Study on Agrobacterium-Mediated Transformation of Pepper with Barnase and Cre Gene

This study was designed to control plant fertility by cell lethal gene Barnase expressing at specific developmental stage and in specific tissue of male organ under the control of Cre/lox system, for heterosis breeding of chili pepper （Capsicum annuum L.）. Chili pepper inbred lines （A, D, E, and I） were transformed with Cre gene and Barnase gene situated between loxp, separately, by means of Agrobacterium co-culture. In this study, we had established a high transformation system by extensive study of affecting factors including genotype, selection of marker, and lethal dose. Cotyledon with petiole from 9-11-day-old seeding was pre-cultured on media MR[MB（MS mineral＋vitamine B5）＋BA（6-Benzyladenine） 5.0 mg·L^-1 ＋IAA（indoleacetic acid） 1.0 mg·L^-1＋GA3（gibberellic acid） 1.0mg·L^-1＋sucrose 3%＋agar 6.5g·L^-1] for 2d. The explants were infected by Agrobacterium tumefaciens when their OD600（optical density at 600 nm）reached 0.6-0.9. After co-cultured for 4-5 d on media MC [MB＋BA5.0 mg·L^-1＋IAA 1.0 mg·L^-1 ＋GA3 1.0 mg·L^-1＋sucrose 3% ＋agar 6.5 g·L^-1＋AS （acetosyringone） 200μmol·L^-1, these cotyledons with petiole were cultured on selective differentiation medium in the media MT[MB medium supplemented with BA [5.0 mg·L^-1＋ IAA 1.0 mg·L^-1＋ GA3 1.0 mg·L^-1＋ AgNO3 5.0 mg·L^-1＋ CW （coconut water） 5% ＋ Km （kanamycin） 65 mg·L^-1＋ Cb （carbenicillin） 500 mg·L^-1＋ 3% sucrose ＋ agar 6.5 g·L^-1].The Kmr （kanamycin resistant） bud rosettes were elongated on selective elongation medium and rooted on rooting medium. PCR and Southern blotting analysis of Kmr plantlet indicated that the foreign genes had been integrated into the genome of pepper. The transgenic plants with Cre gene developed well, blossomed out, and set fruit normally. The transgenic plants with Barnase gene grew well with normal appearance of flower, but they showed different fertility from complete sterility, partial sterility to complete fertility, and similar results were obtained from in vitro pollen germination experiments.