棉秆韧皮纤维化学-生物酶联合脱胶工艺研究

参照GB 5889—1986《苎麻化学成分定量分析方法》,测定经正交试验设计的化学-生物酶联合脱胶后棉秆韧皮纤维各组分的质量分数,寻找最优生物酶脱胶工艺;再利用红外光谱仪分析最优工艺条件下,生物酶处理前后棉秆韧皮纤维结构的变化,并对经过化学预处理与经过化学-生物酶处理的棉秆韧皮纤维的力学性能进行测试。研究表明:生物酶浓度4 g/L、p H值11、温度65℃、处理时间18 h,是棉秆韧皮纤维生物酶脱胶的最优工艺组合;化学-生物酶脱胶工艺并未改变棉秆韧皮纤维的纤维素Ⅰ结构,但其承重却提高了近2倍。

The Study on the Localization of Acid Phosphatase Activity During Spermiogenesis in Chinese Mitten-Handed Crab,Eriocheir sinensis

Ultrastructural cytochemical techniques and electron microscopy were used for localization of acid phosphatase activity during spermiogenesis in Eriocheir sinemsis. The results showed that： Acid phosphatase was synthesized in the endoplasmic reticulum in the early spermatids. The acid phosphatase was found gradually in nucleus, the membrane of acrosomal vesicle, the cytoplasmic region and the acrosomal tubule. And then the reaction product particles became thicker during the spermiogenesis. In the mature sperm, acid phosphatase was localized in the percutor organ slightly, but it was massive and compact in the acrosomal tubule.

Enantio-selective preparation of (S)-1-phenylethanol by a novel marine GDSL lipase MT6 with reverse stereo-selectivity

We previously functionally characterized a novel marine microbial GDSL lipase MT6 and identified that the stereo-selectivity of MT6 was opposite to that of other common lipases in trans-esterification reactions.Herein,we have investigated the use of MT6 in stereo-selective biocatalysis through direct hydrolysis reactions.Notably,the stereo-selectivity of MT6 was also demonstrated to be opposite to that of other common lipases in hydrolysis reactions.Parameters,including temperature,organic co-solvents,pH,ionic strength,catalyst loading,substrate concentration,and reaction time,affecting the enzymatic resolution of racemic 1-phenylethyl acetate were further investigated,with the e.e.of the final（S）-l-Phenylethanol product and the conversion being 97%and 28.5%,respectively,after process optimization.The lengths of side chains of 1-phenylethyl esters greatly affected the stereo-selectivity and conversion during kinetic resolutions.MT6 is a novel marine microbial GDSL lipase exhibiting opposite stereo-selectivities than other common lipases in both trans-esterification reactions and hydrolysis reactions.

菠萝叶纤维精细化处理及其应用技术研究进展

介绍菠萝叶纤维的结构、化学成分、物理性能以及菠萝叶纤维精细化技术研究现状,分析了精细化加工过程中存在的问题,指出化学-生物酶联合法是菠萝叶纤维精细化未来研究的方向。简要介绍了国内外对菠萝叶纤维综合开发的现状。

BIOCHEMICAL GENETIC STUDIES ON CUTTLEFISH SEPIELLA MAINDRONI (CEPHALOPODA: SEPIIDAE)──ACTIVE LOCI SCREENING OF ISOZYME

Screening of 46 putative enzyme coding loci and 4 different kinds of tissues of Sepiella maindroni de Rochebrone, 1884 for enzymatic activities using starch gel electrophoretic technique proved that the 21 enzymes such as AAT, AK, ALP, AP, CK, DIA, ES, FBP, G3PDH, GPI, GRS, IDH, LDH, MDH, MEP, MPI, NP, PGDH, PGM, SOD and XO *, were active to Sepiella maindroni after being stained. The tissue exhibiting stable and clear bands was also determined. Among tissues tested, mantle muscle tissue was the best for electrophoretic survey of isozymes. Buccal bulb muscle, eye and liver were fairly good for some special enzymes, such as DIA, ES, MPI, NP, etc.

Cellular localization and biochemical characterization of a novel calciumdependent protein kinase from tobacco

By screening tobacco cDNA library with MCK1 as a probe, we isolated a cDNA clone NtCPK5 （accession number AY971376）, which encodes a typical calcium-dependent protein kinase. Sequence analyses indicated that NtCPK5 is related to both CPKs and CRKs superfamilies and has all of the three conserved domains of CPKs. The biochemical activity of NtCPK5 was calcium-dependent. NtCPK5 had Vmax and Km of 526nmol/min/mg and 210μg/ml respectively with calf thymus histone （fraction Ⅲ, abbreviated to histone Ⅲs） as substrate. For substrate syntide-2, NtCPK5 showed a higher Vmax of 2008 nmol/min/mg and a lower Km of 30μM. The K0.5 of calcium activation was 0.04μM or 0.06μM for histone Ⅲs or syntide-2 respectively. The putative myristoylation and palmitoylation consensus sequence of NtCPK5 suggests that it could be a membrane-anchoring protein. Indeed, our transient expression experiments with wild type and mutant forms of NtCPK5/GFP fusion proteins showed that NtCPK5 was localized to the plasma membrane of onion epidermal cells and that the localization required the N-terminal acylation sites of NtCPK5/GFP. Taking together, our data have demonstrated the biochemical characteristics of a novel protein NtCPK5 and its subcellular localization as a membrane-anchoring protein.

Effect of salt stress on activity of superoxide dismutase （SOD） in Ulmus pumila L.

The injury tolerance of cell plasma membrane and the correlative enzymes activities of plasma-membrane protection system in the Ulmuspumila leaves treated by nine concentrations （0.3%, 0.6%, 0,9%, 1,2%, 1.5%, 1.8%, 2,1%, 2.4%, 3.0%） of Na2CO3 and NaHCO3 mixtures were studied in a greenhouse of Northeast Forestry University, Harbin, China. The rate of electrolyte leakage （REL） and SOD （Superoxide dismutase） activity in leaves of different samples were determined. Results showed that the REL in leaves of U. pumila presented a slowly increasing trend at the salt concentrations less than 1.5%, which indicated that cell plasma membrane of U. pumila leaves had rather strong resistance to the injury of salt ion, and had a significant increase at the salt concentrations more than 1.5%. The SOD activities in leaves of U, pumila presented an increased trend at salt concentrations less than 1.5%, the growth of seedlings did not decline, and tress and leaves had no symptom of injury, while the salt concentrations exceeded 1.5%, SOD activities sharply decreased and REL increased promptly.

Studies on Partial Molar Volumes of Some Amino Acids and Their Groups in Aqueous Solutions from 293.15K to 333.15K

Densities of aqueous solutions of eight amino acids, glycine, L-alanine, L-valine, L-isoleucine, L-serine, L-threonine, L-arginine and L-phenylalanine, are measured as a function of amino acid concentration from 293.15K to 333.15K. These data are used to calculate the apparent molar volume Vφ and infinite dilution apparent molar volume Vφo (partial molar volume). Data of five amino acids are used to correlate partial molar volume Vφo usinggroup contribution method to estimate the contributions of the zwitterionic end groups (NH3+,COO-) and CH2 group, OH group, CNHNHNH2 group and C6H5(phenyl) group of amino acids. The results show that Vφo values for all kinds of groups of amino acids studied increase with increase of temperature except those for CH2 group, which are almost constant within the studied temperature range. Data of other amino acids, L-valine, L-isoleucine and L-threonine, are chosen for comparison with the predicted partial molar volume Vφo using the group additivity parameters obtained. The results confirm that this group additivity method has excellent predictive utility.

Research Progress in Plant Invertase

Invertase is a key enzyme in sucrose catabolism and crucial for plant assimilate distribution. With the development of molecularbiology, a lot of invertsae genes were cloned recently, and significant progress have been made in regulators on the expression of invertase genes.Thus, this article summarized theresearch progress of invertase in biological characteristics, molecular characteristics and expression regulation.

Effects of Hg and Cu on the activities of soil acid phosphatase

Comparative study on the activity and kinectic properties of acid phosphatase （ACPase） of three soils amended with Hg and Cu at constant temperature and humidity was carried out. The results indicated that the inhibition on ACPase of the three sample soils by Hg and Cu varied with the content of soil organic matter and pH, where, Soil 1 was the most seriously contaminated due to its lowest content of organic matter and the lowest pH among three samples, Soil 2 took the second place, and Soil 3 was the least contaminated. Except Soil 3, the activity of soil ACPase tended to increase along with the contact time under the same type and the same concentration of heavy metal. In particular the Vmax values of ACPase in all three samples decreased with increasing Hg and Cu concentration, whereas the Km values were affected weakly. According to the change of Vmax and Km values, Cu and Hg had the same inhibition effect on soil ACPase. Both of them may he a type of compound of non-competitive and anti-competitive inhibition. Statistic analyses indicated that activities of soil ACPase and Vmax values could serve as bioindicator to partially denote the heavy metal Hg and Cu contamination degree.

IN VITRO ANALYSIS OFτPHOSPHORYLATION SITES AND ITS BIOLOGICAL ACTIVITY

To explore the association between the abnormal phosphorylation sites found in Alzheimer disease (AD) 蚲 and the inhibition of its biological activit y. Methods. Ultracentrifugation, chromatography, manual Edman degradation and autos equence techniques were used to prepare and phosphorylate human recombinant 蚲, isolate and purify 32P 蚲 peptides and determine phosphorylation sites. Results. Phosphorylation of 蚲 by casein kinase 1 (CK 1), cyclic AMP dependent protein kinase (PKA) and glycogen synthetase kinase 3 (GSK 3) separately inhi bited its biological activity and the inhibition of this activity by GSK 3 was significantly increased if 蚲 was prephosphorylated by CK 1 or PKA. The most po tent inhibition was seen by a combined phosphorylation of 蚲 with PKA and GSK 3 . The treatment of 蚲 by PKA and GSK 3 combination induced phosphorylation of 蚲 at Ser 195, Ser 198, Ser 199, Ser 202, Thr 205, Thr 231, Ser 235, Ser 262, Ser 356, Ser 404, whereas Thr 181, Ser 184, Ser 262, Ser 356 and Se r 400 were phosphorylated by GSK 3 alone under the same condition. Conclusion. Phosphorylation of 蚲 by PKA plus GSK 3 at Thr 205 might play a ke y role in 蚲 pathology in AD.

Long interspersed elements in three species of Gracilaria (Gracilariaceae, Rhodophyta)

In order to find out whether long interspersed elements （LINEs） existed in macro-algae gehomes or not, we tested the LINE homologues in representative families （species）： Gracilaria （G. eucheumoides Harv., G. tenuistipitata Chang et Xia, and G. textorii （Sur） De-Toni）, Laminaria （L. longissima Miyabe and L. japonica Aresch.）, and Ulva （U. lactuca L. and U. pertusa Kjellm.） during 2004 to 2005. Polymerase Chain Reaction （PCR） was carried out with degenerate oligonucleotide primers designed from LINEs of rice homologues and Cin4 of maize. Cloning and nucleotide sequencing of the PCR products revealed that 4 clones that derived from 3 species of Gracilaria have LINE homologues. The nucleotide sequences of the 4 LINE homologues diverged greatly, but the amino acid sequences deduced from them were relatively conserved. The endonuclease regions of the LINE homologues greatly diverged from that of other plants, but they had closer phylogenetic relationship to Zepp elements in Chlorella sp., which indicated that sequence divergence by vertical transmission has been a major influence on the evolution of algal LINEs.

Genetic identification of two species of Pleuronichthys by DNA barcoding

DNA barcoding is a new method for biological taxonomy, offering the ability to identify species from fragments in any life-history stage. Pleuronichthys cornutus and P. japonicus are two morphologically similar species. Pleuronichthys japonicus has never been found previously in China. However, in this study, we identified both species using DNA barcoding （cytochrome c oxidase subunit I （COI））, the mtDNA control region and cytochrome b. The results reveal that： l） intraspecific variation in the DNA barcode is much less than interspecific variation; 2） the two morphologically similar species were placed into separate clades distinguishable by high bootstrap values; 3） COI barcodes are more powerful for identifying the two species than the other two mtDNA fragments.

The Process of Immobilization of ZnO Nanorods Surface with Galactose Oxidase

ZnO nanorods, with the c-axis orientation used for transparent conductors, solar cells, sensors especially the functionalized ZnO nanorods with some kinds of enzymes have been used for biosensor. In this work, we describe the process immobilization of galactose oxidase on ZnO nanorods surface with glutaraldehyde as a cross-linker molecule to make the working electrode in electrochemical biosensor. ZnO nanorods were grown on FTO （Fluorine-doped tin oxide） substrate by solution method at low temperature. The crystalline phase and orientation of ZnO nanorods were identified using X-ray diffraction. The efficiency of the immobilization was calculated by Braford method showed that about 36% enzyme content was immobilized on ZnO nanorods surface. The working electrode based on the immobilized ZnO nanorods was tested in galactose solution by CV （cyclic voltammetry） method indicated the value of current intensity is about 0.14 μA. These results clearly demonstrate the potential of galactose sensor based on ZnO nanorod.

Effects of spaceflight and simulated microgravity on cell sub-microstructure and antioxidant enzyme activity in tomato

Controlled ecological life support systems provide food, air, water, and other basic living resources for crew members on long-duration spaceflight missions. Plants are an important basic requirement of these systems and their biological characteristics in space have very high research value. Based on experiments of spaceflight in Shenzhou 8 spacecraft and simulating microgravity effects on three-dimensional （3-D） clinostat, the biological characteristics of tomato＇s leaf cell sub-microstructure and antioxidant enzyme activities were studied and compared in this work. Results showed that leaf cell sub-microstructure of the tomato samples experiencing spaceflight had more changes than effects, and both peroxidase （POD） and superoxide dismutase （SOD） that of the samples processed by simulated microgravity activities increase obviously in both the environments.

Assembly of layer-by-layer films of superoxide dismutase and gold nanorods:A third generation biosensor for superoxide anion

Based on the layer-by-layer self-assembly of positively charged cetyltrimethylammonium bromide （CTAB） wrapped gold na- norods （AuNRs） and negatively charged superoxide dismutase （SOD） from their aqueous solutions on cysteine modified gold electrode （Cys/Au）, a third generation electrochemical biosensor （（SOD/AuNRs）2/Cys/Au） for superoxide anion （02＂-） was developed. The two layers assembly of SOD/AuNRs can significantly enhance the direct electron transfer between SOD and the electrode. The functional enzymatic activities of the SOD offer an electrochemical approach to the determination of 02＂-. In the reductive regions, the proposed sensor exhibits excellent analytical performances, such as wide linear range （200 nM to 0.2 mM O2-）, low detection limit （100 nM O2-）, high sensitivity （22.11 nA cm-2 μM-1）, short response time （less than 5 s）, good stability and reproducibility, while no obvious interferences are caused by commonly met interfering species including hydrogen peroxide （H202）, uric acid （UA） and ascorbic acid （AA）.

Gold nanolabels and enzymatic recycling dual amplification-based electrochemical immunosensor for the highly sensitive detection of carcinoembryonic antigen

A sensitive electrochemical immunoassay system for the detection of a protein tumor biomarker through a dual amplified strategy was reported. Firstly, this protocol involves in the electropolymerization of o-aminobenzoic acid (o-ABA) on a glass carbon electrode (GCE). Subsequently, capture anti-CEA (Abl) is covalently linked to poly(o-ABA) (PAB) film, via N-(3-dimethylamminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC), and N-hydroxysulfosuccinimid sodium salt (NHS) activation of the carboxyl groups and surface blocking with ethanolamine. Later, the target, carcinoembryonic antigen (CEA), is sandwiched between an electrode surface confined Ab1 and the alkaline phosphatase-labeled signal anti-CEA antibodies conjugated with gold nanoparticles (Ab2-ALP-AuNP bioconjugates). The dual biocatalytic signal amplification for CEA monitoring is achieved by coupling the numerous enzymes loaded on the AuNPs with redox-recycling of the enzymatic products in the presence of the secondary enzyme and the corresponding substrate. The novel dramatic signal amplification strategy, exhibits a good linearity at the studied concentration range from 0.005 to 50 ng mL-1 towards CEA with a detection limit of 2 pg mL-1 (S/N=3). There is a 5-100-fold improvement in detection limit compared to other similar studies. The developed dual signal amplified strategy shows good selectivity, regeneration, stability and acceptable reproducibility. Therefore, the signal amplification approach holds great potential applications in detection of ultra-trace protein biomarkers.

Advance in the Study of the Mechanisms Regulated by Sphingosine-1-Phosphate

Sphingosine-1-phosphate（S1P） is a bioactive lipid messenger in the cells that regulate gene expression and NF-κB signal pathway through unknown mechanisms.Recently,Cheng Luo,associate professor of DDDC in Shanghai Institute of Materia Medica,whose project was funded by the National Natural Science Foundation of China,joined in a research team led by Professor Sarah Spiegel of Virginia Commonwealth University.The team continuously made significant breakthroughs in understanding the regulation mechanism of Sphingosine-1- Phosphate.In September 2009,in a paper published on SCIENCE magazine（Science 2009, 325：1254-7）,they firstly demonstrated that S1P is a physiologically important regulator of histone deacetylases（HDACs）,HDACs are direct intracellular targets of S1P.Furthermore,they identified the mechanism that S1P regulates gene expression through regulating the activity of HDACs.In June 24th,2010,in another paper to be published on NATURE magazine（Nature 2010,June 24th,advance online publication,（doi：10.1038/ nature09128）） which reports the regulation of NF-κB signaling pathway by S1P.They demonstrate that S1P is the missing cofactor for TRAF2（tumour-necrosis factor receptor-associated factor 2） and indicate a new paradigm for the regulation of lysine-63- linked poly-ubiquitination.The study also highlight the key role of SphK1 and its product S1P in TNF-αsignalling and the canonical NF-κB activation pathway, and then play crucial role in inflammatory,antiapoptotic and immune processes.The identification of new mechanisms fay which S1P regulates gene expression and TNF and NF-κB signaling pathway will light up the road to develop novel inhibitors that might be useful for treatment of cancer and in- flammatory diseases.

Histological,enzymohistochemical and biomechanical observation of skeletal muscle injury in rabbits

Objective： To explore the pathophysiological and biomechanical features of skeletal muscular injury for providing a rational basis for its treatment, prevention and rehabilitation. Methods ： In 70 adult rabbits, the left tibialis anterior （TA） muscle was stretched to injury, while the right TA muscle served as control. Histological, enzymohistochemical and biomechanical changes were observed on days 0, 1, 2, 3, and 7 after injury. Cytochrome oxidase （ CCO ）, acid phosphatase （ ACP ）, ATPase, succinate dehydrogenase （ SDH ）, malate dehydrogenase （ MDH ）, NADH-diaphorase （ NADHD ）, giutamatedehydrogenase （ GDH ）, α-giycerophosphate dehydrogenase （ α-GPD ） and lactate dehydrogenase （LDH） were measured. The examined biomechanical parameters included maximal contractile force, ultimate load, length, energy absorption, tangent stiffness, and rupture site. Results： Partial or complete rupture of TA muscle occurred near the muscle-tendon junction. There was an intense inflammatory reaction on day 1 and 2 after injury. Endomysium fibrosis and myotube formation were observed on day 3, and developed further on day 7. The activity of cell oxidases （ CCO, ATPase, MDH, α-GPD, SDH,NADHD and GDH） showed a significant drop from day 0 to 2, and resumed with different levels on day 3. The increment of enzymatic activities continued on day 7 and the levels of NADHD and α-GPD reached to the levels of control muscle. Maximal contractile force was 70.17 % ± 3.82% of controls immediately after injury, 54.82% ±3.09% at 1 day, 66.41% ±4.36% at 2 days, 78.39%± 4.90 % at 3 days and 93.64 % ±5.02 % at 7 days. Ultimate load was 85.78 %±7.54 % of controls at the moment of injury, 61.44 % ± 5.91% at 1 day, 49. 17 %± 4.26 % at 2 days, 64.43% ±5.02% at 3 days, and 76.71% ±6.46% at 7 days. Conclusions ： Endomysium fibrosis and scar formation at the injured site are responsible for frequent recurrence of skeletal muscle injury. Recovery of tensile load slower than that of maximal contractile force may be another cause. Whether the injured muscle returns to normal exercise is mainly determined by the teusility on which the muscle-tendon can bear rather than the maximal contractile force.

Shifts in chemical and microbiological properties belowground of invader Ageratina adenophora along an altitudinal gradient

Tropical mountain ecosystems are usually colonized by numerous invasive plant species and represent an ideal‘natural laboratory’to study the effects of altitude on plant invasion.The aim of this study was to investigate the soil chemical and microbiological properties along an altitudinal gradient on a mountain colonized by the invader Ageratina adenophora.Rhizosphere soil of A.adenophora was collected over an altitudinal gradient(1400–2400 m)in Ailao Shan,China.We determined soil organic carbon(C),nutrient contents,enzyme activities,bacterial community composition as well as C and nitrogen(N)contents of the plant roots.Ecoenzymatic stoichiometric indices were calculated to estimate the relative C,N or P limitations of the microbial community.There was a significant effect of altitude on soil organic C in the rhizosphere,and a turning point in these measured variables was detected at an altitude of 2000 m.At low elevations,the rapid growth of invasive plants depleted the deficient phosphorus(P)in tropical soils,leading to microbial P limitation;at high elevations,microbes invested more energy to obtain C from resistant litter,leading to microbial C limitation.Bacterial beta diversity and soil pH contributed most to the altitudinal differences in ecoenzymatic stoichiometry,and Proteobacteria and Acidobacteria were the dominant bacterial phyla that determined the nutrient uptake status of microorganisms.These results demonstrate how microbial nutrient acquisition belowground of A.adenophora along an altitudinal gradient,which could contribute to further knowledge about the effects of altitude on biological invasion.