微生物快速检验技术进展

传统的微生物分离、 培养与鉴定需时较长,难以适应诊断、治疗的需要。近年己发展了多种快速诊断方法,可在常规微生物实验室应用。如自血、尿、粪便、脑脊液中直接检查微生物的抗原或抗体,应用细菌的专有酶和酶的产色或荧光底物进行快速鉴定,应用各种载体的快速凝集试验,自标本中直接检查细菌的毒素以及快速细菌药敏试验等。

过氧化氢生物传感器的研究新进展

过氧化氢在各个领域被广泛应用,然而过氧化氢在致癌、加速衰老或诱发心血管疾病等方面有一定的毒性作用,因此,建立一种快速、灵敏、高效、稳定的方法来测定过氧化氢具有重要的意义。由此文章综述了在有酶和无酶对比条件下过氧化氢生物传感器的发展历程及作用原理。

电化学方法检测胆固醇的研究进展

随着人类生活质量的提高,胆固醇的检测成为一个重要的研究课题。电化学方法具有选择性强、检测速度快等优点。综述了近年来利用电化学方法在有酶和无酶的条件下测定胆固醇含量的电化学原理和最新进展,并对用电化学方法检测胆固醇进行了展望。

Effects of Metal Ions and Organic Solvents on Alkaline Phosphatase from Rice-field Eel

[Objective]The mechanism of alkaline phosphatase（ALP） was studied to promote rice-field eel aquaculture industry. [ Method] The effects of effectors such as multiple metal ions and organic solvents on ALP in viscera of rice-field eel. [ Result] Na^＋ and K ^＋ didn＇t generate big influences on enzyme activity;Mg^2＋ and Ca^2＋ could promote ALP while Li^＋,Cu^2＋ and Zn^2＋ could restrain ALP enzyme activity. Both HPO4^2- and WO4^2- generated by en- zyme catalyzing disodium phenyl phosphate possessed strong inhibitory effects on emzymc, and 9.5 mmol/L HPO4^2 - would make enzyme activity decline by 13% while 9.5 mmol/L WO4^3- would make enzyme decline by 34%. The inhibition types of them were both competitive inhibition on enzyme activity. The organic solvents such as methanol, ethanol,ethylene glycol,isopropannl all generated influences on ALP and the order according to their inhibitory effects was isopropanol 〉 ethanol 〉 methanol 〉 ethylene glycol. [ Conclusion] The inflncnces of various effeetors on ALP aetivity of rice-field eel were studied from dynamics perspective to provide theoretical basis for further clarifying ALP mechanism.

Effects of Chitosan and Organosilicon on Antioxidant Enzyme Activity of Tomato Seedlings Diseased with Botrytis cinerea

[Objective] The aim was to explore effects of chitosan and organosilicon on tomato resistance against Botrytis cinerea. [Method] With leaf spraying method adopted, four groups were set in the test, including control group, groups treated with Botrytis cinerea, with chitosan and Botrytis cinerea, and with chitosan, organosilicon, and Botrytis cinerea, in order to study on effects of chitosan and organosilicon on antioxidant enzyme activities of tomato. [Result] Antioxidant enzyme activities of tomato seedlings were improved in groups with chitosan, organosilicon, and Botrytis cinerea and with chitosan and Botrytis cinerea compared with the group treated with Botrytis cinerea. On the 6th d, activities of PAL, SOD, POD, PPO and CAT in groups with chitosan and Botrytis cinerea, and with chitosan, organosilicon, and Botrytis cinerea increased by 27.36%, 52.07%, 43.55%, 82.02% and 71.82%, and 18.91%, 30.22%, 57.14%, 38.09% and 53.64%, respectively. [Conclusion] Antioxidant enzyme activities of tomato seedlings could be improved by chitosan and organosilicon.

Effects of Exogenous Organic Acids on Dehydrogenase Activity in Dark Brown Forest Soils under Nutrient Deficiency

Objective] This study aimed to investigate the effects of exogenous or-ganic acids on dehydrogenase activity in dark brown forest soils under nutrient defi-ciency. [Method] Different proportions of A1 and B horizon dark brown forest soils （A1∶B=1∶2） were utilized to establish soil conditions with nutrient deficiency for cultivation of Larix olgensis seedlings. The effects of oxalic acid, citric acid and succinic acid on dehydrogenase activity in dark brown forest soils under nutrient deficiency were studied systematical y by adding different concentrations of organic acid solutions. [Result] Under nutrient deficiency, dehydrogenase activity in dark brown forest soils was reduced significantly, and the reduction increased with the extension of stress duration. Most organic acid treatments improved dehydrogenase activity in nutrient-deficient dark brown forest soils, and the effects varied with different treatment du-ration and types and concentrations of organic acids. Furthermore, 10.0, 5.0 and 10.0 mmol/L organic acid treatments exhibited the most significant effects on day 10, 20 and 30, respectively. The increment of dehydrogenase activity in different durations showed a downward trend of 30 d〉20 d〉10 d; the improvement effects of three organic acids on dehydrogenase activity showed a downward trend of succinic acid〉 citric acid〉oxalic acid. [Conclusion] Exogenous organic acids improved signifi-cantly dehydrogenase activity in dark brown forest soils under nutrient deficiency and also improved the microbial activity and soil fertility to a certain extent.

Effects of Diazepam on Rat Cholinesterase andMembrane Stability of Red Cell in vitro

The results confirmed that diazepam inhibits the cholinesterase in rat serum,red cell,corpus striatum and diaphragm in vitro,that the higher the diazepam concentration,the stronger the cholinesterase inhibition,and that diazepam is a reversible inhibitor to acetylcholinesterase and diazepam has a stabilizing action on red cell membranes.The role of these effects of diazepam in the treatment of organic phosphate poisoning is discussed

Inhibition of Porcine Small Intestinal Sucrase by Validamine

As an important medicinal intermediate with broad uses, validamine, an aminocyclitol, isolated from the enzymolysis broth of validamycins, has gained more and more attention. The absolute configuration of validamine is similar to that of a-D-glucose, and it demonstrates powerful inhibition activity on glycosidase. In this paper, the inhibitory effect of validamine on porcine small intestinal sucrase was investigated. Validamine was found to be a potent, competitive inhibitor to porcine small intestinal sucrase in vitro with an IC50 value of 6.85 X 10(-4) mol center dot L-1. Validamine exhibited a dose-dependent inhibition effect on porcine small intestinal sucrase, whereby the inhibition interaction of validamine and porcine small intestinal sucrase was a fast binding process. The inhibition of validamine on porcine small intestinal sucrase was pH-dependent.

Involvement of ERK1/2 and p38 MAPK in up-regulation of 14-3-3 protein induced by hydrogen peroxide preconditioning in PC12 cells

Objective To investigate the protective effects of hydrogen peroxide preconditioning （HPP） on the pheochromocytoma （PC12） cells treated with 1-methyl-4-phenylpyridinium （MPP^＋） and to explore the potential mechanisms. Methods The viability and apoptosis of PC 12 cells were determinded by 3-（4,5-dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide （MTT） assay and 4′,6′-diamidino-2-phenylindole （DAPI） staining, respectively. The expressions of 14-3-3 protein and phospholylated p38 mitogen-activated protein kinase （MAPK） were determined by Western blot. Enzyme-linked immunosorbent assay （ELISA） was used to measure the activity of extracellular signal-regulated protein kinase 1/2 （ERK1/2）. Results The cell viability decreased and the number of apoptotic cells increased dramatically in MPP^＋ group compared with that in Control group. HPP induced a significant increase in cell viability and a marked decrease in population of apoptotic cells of the MPP^＋- treated PC 12 cells, accompanied with up-regulation of 14-3-3 protein and increase of ERK 1/2 and p38 MAPK activities. The 14-3-3 protein expression was positively correlated with the phosphorylation of ERK1/2. Furthermore, inhibition of the ERK1/2 with PD98059 abolished the 14-3-3 protein up-regulation in PC 12 cells induced by HPP. Conclusion HPP protects PC 12 cells against MPP＋ toxicity by up-regulating 14-3-3 protein expression through the ERK1/2 and p38 MAPK signaling pathways.

Variation in Enzymes Activities of Rhizospheric Substrate and Influencing Factors during Nursing of Watermelon Seedlings

[Objective] This study aimed to investigate the effects of substrate en- zymes activities on nursing of watermelon seedlings. [Method] The composted mushroom residue was mixed with garden soil according to a certain proportion to prepare nursing substrate for watermelon seedlings. During the nursing, the activity variation in alkaline phosphatase, acid phosphatase, neutral phosphatase and urease was investigated. In addition, the correlations between pH value, total nitrogen con- tent, total phosphorus content and organic matter content in substrate and enzyme activity were studied. At different young seedling stages, the rhizospheric substrates with different formulas were sampled for determination of enzymes activities. [Result] The enzyme activity differed significantly among different substrates. The correlation analysis results showed that the higher the organic matter content and total nitrogen content in substrate are, the higher the urease activity is; the phosphatase activity was significantly related to the organic matter content, total nitrogen content and to- tal phosphorus content in substrate; the pH value of substrate was significantly relat- ed to rhizospheric alkaline phosphatase activity; the shoot dry weight was positively related to urease activity; there was a significant correlation between phosphatase activity and root dry weight. [Conclusion] Through determining enzymes activities in the rhizospheric substrate for nursing watermelon seedlings, the optimum substrate M3 was screened out. The activities of rhizospheric urease, alkaline phosphatase, acid phosphatase and neutral phosphatase in the substrate M3 were all higher than those in the substrate fertilized with manure.

Cloning, overexpression, and characterization of a novel organic solvent-tolerant lipase from Paenibacillus pasadenensis CS0611

We found a novel lipase gene in the Paenibacillus pasadenensis CS0611 strain.The lipase gene sequence was cloned into the pET-28a expression vector to construct a recombinant lipase protein containing 6×His tags at the C-and N-termini,respectively.High-level expression of the lipase in E.coli BL21(DE3)was obtained upon induction with IPTG at 20°C.The recombinant lipase activity was approximately 1631-fold higher than the wild type.His-tagged recombinant lipase was purified rapidly and efficiently by using Ni-charged affinity chromatography with 63.5%recovery and a purification factor of 10.78.The purified lipase was stable in a broad range of temperatures and pH values,with the optimal temperature and pH being 50°C and 7.0,respectively.Its activity was stimulated to different degrees in the presence of metal ions such as Ca2+,Mg2+,and some non-ionic surfactants.In addition,the purified lipase was activated by a series of water-miscible organic solvents such as some short carbon chain alcohols and was highly tolerant to some water-immiscible organic solvents.

Synthesis of Vitamin A Esters by Immobilized Candida sp. Lipase in Organic Media

Vitamin A ester was synthesized in organic solvents with immobilized lipase from Candida sp. The types of lipases, influences of solvent, the molar ratio of substrates, the reaction temperature and the water activity in the reaction were studied in detail in order to obtain the optimum conditions for Vitamin A palmitate synthesis. In a system of hexane, 100mg immobilized Candida sp. lipase was used in the presence of 1.2mmol vitamin A acetate and 3.6mmol palmitic acid. The yield of vitamin A palmitate reached 81% in 12h at 25℃. The immobilized Candida sp. lipase was prepared by adsorbing Cand/da sp. fermentation broth on pretreated textile and could be reused for at least six batches.

Pedogenic Carbonate and Soil Dehydrogenase Activity in Response to Soil Organic Matter in Artemisia ordosica Community

Little attention has been paid to the role of soil organic matter （OM） in the formation of pedogenic carbonate in desert soils. The relationships among soil OM, soil dehydrogenase activity （DHA）, and soil CaCO3 in a plant community dominated by Artemisia ordosica, located on the eastern boundary of Tcngger Desert in the Alxa League, Inner Mongolia, China, were studied to understand whether OM was directly involved in the formation of pedogenic carbonate. The results showed that DHA and CuCO3 positively correlated with OM content, and DHA, OM, and CaCO3 were correlated with each other in their spatial distribution, indicating that abundant OM content contributed to the formation of CaCO3. Therefore, the formation of pedogenic CaCO3 was a biotic process in the plant community dominated by A. ordosica.

Function and regulation of Aurora/Ipllp kinase family in cell division

During mitosis, the parent cell distributes its genetic materials equally into two daughter cells through chromosome segregation, a complex movements orchestrated by mitotic kinases and its effector proteins. Faithful chromosome segregation and cytokinesis ensure that each daughter cell receives a full copy of genetic materials of parent cell. Defects in these processes can lead to aneuploidy or polyploidy. Aurora/Ipl1p family, a class of conserved serine/threonine kinases, plays key roles in chromosome segregation and cytokinesis. This article highlights the function and regulation of Aurora/Ipllp family in mitosis and provides potential links between aberrant regulation of Aurora/Ipllp kinases and pathogenesis of human cancer.

Lipase-catalyzed Kinetic Resolution of Racemic 1-Trimethylsilylethanol in Organic Solvent

The enantioselective esterification of racemic 1-trimethylsilylethanol with acids catalyzed by lipase in organic solvent was successfully performed. The influence of some factors on the reaction was investigated. Among the four lipases explored, Candida rugosa lipase (CRL) showed the highest activity and enantioselectivity. Octanoic acid was the best acyl donor among the eleven acids studied and n-hexane was the most suitable medium for the reaction. The optimum shaking rate and temperature were found to be 150r·min-1 and 20℃ to 30℃, respectively. The enantiomeric excess of the remaining (5)-(-)-1-trimethylsilylethanol was 93% when substrate conversion was 53% upon incubation of the reaction mixture at 30℃, 150r·min-1 for 12 h.

Aurantiamide Acetate from Stems of Zanthoxylum Dissitum Hemsley

The genus Zanthoxylum (Rutaceae) is comprised of about 50 species in China.Zanthoxylum dissitum Hems-ley, locally called 'Shanpipa', has been used to treat pains in the loin,limbs and arthritis in Chinese folk medicine. Our previous study reported the isolation andidentification of five quinoline alkaloids . In this continuing study on the chemcial constituentsof Zanthoxylum dissitum Hemsley, a phenylalanine derivative was obtained .

Role of p38 MAPK in lipopolysaccharide-induced iNOS expression by endothelial cells

Objective:To examine the role of p38 mitogen-activated protein kinase (MAPK) in NO production and Inos expression in human endothelial cells stimulated by lipopolysaccharide (LPS). Methods: The NO level in the supernatant of the cell culture media was measured with Griess method, expressions of Inos protein and Mrna in vitro cultured endothelial cell line ECV304 were detected with immunofluorescence analysis and reverse transcriptase-PCR respectively. Immunokinase assay was employed to measure P38mapk activity. Results: Compared with the basal level of Inos expression and NO production, the NO level and the expressions of Inos Mrna and protein in the cells were increased after LPS stimulation. P38mapk activity in ECV304 cells exhibited a marked increase at 15 min after LPS stimulation, lasting for about 45 min before gradually decline. The Inos protein and Mrna expressions induced by LPS stimulation was significantly inhibited by SB203580 [4-(4-fluorophenyl)-2-(4- methylsulfinylphenyl)-5-(4-pyridyl) imidazole], a highly specific inhibitor of p38 MAPK. Conclusion: p38 MAPK plays an important role in iNOS expression and NO production in ECV304 cells, and the inhibition of the signal transduction pathway can be effective to reduce the production of iNOS and other cytokines, and therefore constitutes a useful strategy for treating septic shock or inflammation.

INVOLVEMENT OF p38 MITOGEN-ACTIVATED PROTEIN KINASE IN E.coli-INDUCED U937 APOPTOSIS

Objective To investigate whether the effect of E. coli on U937 cell fines apoptosis is mediated via p38 mitogen-activated protein kinase （MAPK） activation. Methods The U937 cell lines were treated with E. coli at different time or together with SB203580, an inhibitor for p38. Cell apoptosis was analyzed by flow cytometry, p38 activities were detected by Western blotting. Results E. coli induced apoptosis in cultured U937 cell lines in a time-dependent manner. The phosphorylation of p38 was induced after 10 minutes infection, reached the peak after 20 minutes, and started to decline after 30 minutes. In contrast, the level of total p38 protein was not changed in whole experimental period. Inhibition of p38 with SB203580 significantly inhibited E. coli induced apoptosis in U937 cells. Conclusion The activation of the p38 MAPK in U937 cell lines by E. coli is a major pathway to mediate the apoptosis.

MEK AND p38 MAPK-DEPENDANT PATHWAYS ARE INVOLOVED IN THE POSITIVE EFFECT OF INTERLEUKIN-6 ON HUMAN GROWTH HORMONE GENE EXPRESSION IN RAT MtT/S SOMATOTROPH CELLS

Objective To investigate the effect of interleukin-6(IL-6)on the human growth hormone(hGH)gene expression in a rat somatotropic pituitary cell line MtT/S.Methods The plasmids containing various lengths of hGH gene 5'-promoter fragments were constructed.Stably transfected MtT/S cells were created by cotransfecting the above plasmids and pcDNA3.1(+)with DMRIE-C transfection reagent.After the administration of these cells with IL-6 and/or various inhibitors of signaling transduction pathways,the luciferase activities in MtT/S cells lysis were assayed to demonstrate the effects of IL-6 on hGH gene promoter activity and possibly involved mechanism.Results The 103 U/mL IL-6 stimulated GH secretion and synthesis,and promoted the 5'-promoter activity of GH gene in stably transfected MtT/SGL cells with the action of 1.69 times above the control.Among inhibitors of signaling transduction pathways,mitogen-activated protein kinase kinase(MAPKK/MEK)inhibitor PD98059(40 μmol/L)and p38 mitogen-activated protein kinase(MAPK)inhibitor SB203580(5 μmol/L)completely blocked the stimulatory effect of IL-6.Western blot analysis further confirmed the activation of phosphorylated MEK and p38 MAPK in MtT/SGL cells.Neither over-expression of Pit-1 nor inhibition of Pit-1 expression affected IL-6 induction of hGH promoter activity.A series of deletion constructs of hGH promoter were created to identify the DNA sequence that mediated the effect of IL-6.The results showed that the stimulatory effect of IL-6 was abolished following deletion of the-196 to-132 bp fragment.Conclusions IL-6 promotes GH secretion and synthesis by rat MtT/S somatotroph cells.The stimulatory effect of IL-6 on hGH gene promoter appears to require the activation of MEK and p38 MAPK,and a fragment of promoter sequence that spans the-196 to-132 bp of the gene,but may be unlinked with Pit-1 protein.

Pickering interfacial biocatalysis with enhanced diffusion processes for CO_(2) mineralization

Utilization of carbon dioxide(CO_(2))has become a crucial and anticipated solution to address environmental and ecological issues.Enzymes such as carbonic anhydrase(CA)can efficiently convert CO_(2) into various platform chemicals under ambient conditions,which offers a promising way for CO_(2) utilization.Herein,we constructed a Pickering interfacial biocatalytic system(PIBS)stabilized by CA‐embedded MOFs(ZIF‐8 and ZIF‐L)for CO_(2) mineralization.Through structure engineering of MOFs and incorporation of Pickering emulsion,the internal and external diffusion processes of CO_(2) during the enzymatic mineralization were greatly intensified.When CO_(2) was ventilated at a flow rate of 50 mL min^(–1) for 1 h,the pH value of PIBS dropped from~8.00 to~6.50,while the average pH value of free system only dropped to~7.15,indicating that the initial reaction rate of CO_(2) mineralization of PIBS is nearly twice that of the free system.After the 8^(th) cycle reaction,PIBS can still produce more than 9.8 mg of CaCO_(3) in 5 min,realizing efficient and continuous mineralization of CO_(2).