Abstract:AuMan5A，a glycoside hydrolase family 5 β-mannanase of Aspergillus usamii，only consists of a catalytic domain. To explore the effect of fusion position of carbohydrate-binding module on the enzymatic properties of AuMan5A，two fusion genes，Auman5A-cbm27 and cbm27-Auman5A，were constructed as designed theoretically by separately linking family 27 CBM of Thermotoga maritime β-mannanase into C- and N-termini of AuMan5A. After Auman5A and two fusion genes were expressed in Pichia pastoris GS115，enzymatic properties of the expressed products，AuMan5A，AuMan5A-CBM27 and CBM27-AuMan5A，were analyzed and compared. The results showed that the temperature optima（Topt） of three β-mannanases were 70，70 and 60 ℃，respectively. AuMan5A-CBM27 was thermostable at 68 ℃， while AuMan5A and CBM27-AuMan5A were at 60 and 58 ℃. Besides，three β-mannanases displayed the same pH optimum of 4.0. AuMan5A was stable at a pH range of 2.5~7.5，while AuMan5A-CBM27 or CBM27-AuMan5A was at pH 2.0~9.0. Compared with that of AuMan5A，Km values of AuMan5A-CBM27 and CBM27-AuMan5A towards locust bean gum decreased by 45.0% and 26.3% respectively. It was demonstrated in this work that the fusion of CBM27 and its position into AuMan5A played significant roles in its enzymatic properties.

Abstract:Lactic acid bacteria（LAB） can be used for prevention and auxiliary treatment of hyperuricemia and its complications. By using such way，the detrimental effect of people’s living quality caused by maintaining strict dietary restriction（the conventional treatment of hyperuricemia） could be reduced or eliminated. In this study the purine nucleoside-decomposing ability of LABs were isolated from Chinese sauerkraut bought from local supermarket by HPLC method through the determination of their insosine and guanosine degradation abilities. Among which strain 5-1 showed the best probiotic potential with degradation rate of insosine and guanosine are 89.37% and 93.20% respectively. Based on physiological，biochemical tests and 16S rDNA classification，strain 5-1 was identified as Lactobacillus curvatus. After 4 h treatment under the circumstance of pH 3 or 0.3% bile salt，strain 5-1 could keep a survival level of 107 cfu/mL. Furthermore the great tolerance capacity was shown in the artificial gastrointestinal fluid with food suspension and excellent purine nucleoside-decomposing ability in artificial intestinal juice with food suspension．Therefore，Lb. curvatus 5-1 has a great potential for applications in the development of microbial agents that help to prevent and auxiliarily treat hyperuricemia.

Abstract:Using the biological primary databases at the National Center for Biotechnology Information（NCBI），We construct a secondary database of human cancer-related nucleotides p53. The database design mainly includes four aspects：cancer information，p53 sequence information，sample information and reference information. We store the data from NCBI in XML file，then by parsing the files to secondary database and initially realize the data of searching，linking ，statistical analysis and other functions. p53 cancer gene sequences are compared by quasi alignment one dimensional cellular neural network method，and the sensitivity and reliability of the global alignment of the two sequences are enhanced by improving the similarity evaluation formula. We applied the improved sequence alignment algorithm in non small cell lung cancer and breast cancer p53 gene sequence alignment , the result shows that there are great differences in mutant p53 Exon 5 of two cancer sequences which can be used to discriminate these two cancers. In addition, by transforming the primary database into the secondary database in the form of XML, the dynamic exchange of network data and local data is redized, which provides an excellent platform for the study of cancer and p53 gene.

Abstract:In order to study the resource of the endophyte from Gentiana straminea Maxim in Sichuan，the cultivable endophyte in different organs of Gentiana straminea Maxim（roots，stems，leaves and flowers） were isolated and identified by the dilution separation methods and the sequence analysis of 16 S rDNA and ITS-rDNA. Then the content of gentiopicrin in each organ was determined by high performance liquid chromatography（HPLC） and its correlation with the abundance of cultivable endophytes was analyzed. The results showed that 13 strains of cultivable endophytes were separated from Gentiana straminea Maxim as a whole，which belong to 3 phyla，6 classes，8 orders，9 families and 10 genera，and among them，ten strains were bacteria and three were fungi. Meanwhile，there were differences of the quantity and microbial population of the cultivable endophytic bacteria and fungi in different organ or position of Gentiana straminea Maxim. The maximum quantity of endophytic bacteria and fungi were（8.77±0.71）×105 cfu/g and（2.00±1.05）×104 cfu/g respectively and both in the roots of Gentiana straminea Maxim.. Statistical analysis showed that correlations between the content of gentiopicrin and the quantity of cultivable endophytic bacteria and fungi，respectively，were extremely significantly positive（P<0.01）. More over，the quantity of each predominant strain，bacteria QJ2（Duganella sp.），bacteria QJ6（Rahnella sp.），bacteria QJ10（Pseudomonas sp.） fungi QJ11（Aspergillus sp.） and fungi QJ13（Aspergillus sp.），was extremely，significantly and positively related with the content of gentiopicrin，respectively（P<0.01）. The cultivable endophytes in Gentiana straminea Maxim were characterized by abundant genetic diversity and there were differences of the endophytes community structure in different organ of Gentiana straminea Maxim. All the results will serve as the foundation of development and utilization of endophytes in Gentiana straminea Maxim.

Abstract:To study the processing technology of fermented brown rice steam sponge cake，using indica brown rice as raw material. The results of single factor experiment showed that the early period fermented by lactic acid bacteria mainly affected the flavor，the fermented brown rice steam sponge cake taste best when the brown rice flour fermented for 4 h at 28 ℃ by lactic acid bacteria. while the yeast addition volume and the mixed fermentation temperature and time affected texture quality significantly. Orthogonal experiment results showed that the best three fermentation conditions according to the lower hardness and adhesiveness and higher springiness respectively were A2B3C3，A3B3C3 and A2B1C3. The results of sensory analysis on the basis of orthogonal experiment showed the optimum fermentation is adding 1.2% of the lactic acid bacteria fermented in 28 ℃for 2 h，and next adding 2.5% of yeast fermented in 32 ℃for 2 h.

Abstract:Pretreatment of lignocelluloses by ionic liquid was paid more and more attention recently due to its high efficiency and environment friendly features. In this study，the effect of four different ionic liquids（[Emim]Ac、[Amim]Cl、[Bmim]HSO4、[Bmim]BF4） pretreament on the enzymatic efficiency was investigated. Besides，the change of morphology，chemical composition and structure of corn stalk was also investigated to explore the relative mechanism. The results indicated that the connection bond between lignocellulose and hemicellulose was destoryed and enzymatic hydrolysis was raised after treated by ionic liquids of [Emim]Ac、[Amim]Cl and[Bmim]BF4 at 130 ℃ for 1.5 h. The ionic liquid of[Emim]Ac showed an selective effect in removal of lignin，reaching to 54.47%. Besides，the compact network structure of corn stalk was destroyed and a coarser surface was formed. Meanwhile，the crystal structure of cellulose was transformed from I to II，and the enzymatic hydrolysis was up to 91.20%，equaling to 4.77 times of untreated.

Abstract:In order to reveal the relationship between the tryptophan residue and the activity of Man1602，the expression of Man1602 was increased，the authors studied the reaction of tryptophan-specific chemical modifier with mannanase Man1602，at the same time，according to the sequence alignment results of Man1602，the relationship between tryptophan residues and enzyme activity was deduced，the fixed position of tryptophan was fixed，and the activity of the mutant was detected. The results showed that tryptophan is the active essential amino acid of Man1602，in which the mutation of W196 and W199 sites leads to almost complete loss of enzyme activity. It can be inferred that W196 and W199 are the active centers of amino acids. W198 also has a high conservation，but its mutation did not cause the complete inactivation of the enzyme so it was concluded that W198 was an important residue to sustain the hydrophobic constant of the enzyme active site.

Abstract:In order to overexpress soluble human insulin-like growth factor-1（IGF-1） in Escherichia coli and obtain a large number of bioactive proteins.A fusion gene containing trx and igf-1 was constructed via fusion of PCR technique and inserted into pET30a plasmid. Then the recombinant plasmid was transformed to E.coli strain C43（DE3） and a series of conditions optimization was carried out. After purification，target protein was identified by Western blot and determined for bioactivity. Trx-IGF-1 fusion protein was high expressed in soluble form in C43（DE3） after induction with 1mmol/L IPTG at 30 ℃ for 5 h. The recombinant protein reached a purity above 90% by using Ni2+ affinity chromatography and showed a strong antigenic ability by Western blot. The biological activity assay revealed that the fusion protein Trx-IGF-1 could significantly promote the proliferation and cell cycle progression of NIH3T3 cells. The carrier protein Trx can be used to realize the high-level soluble expression of bioactive IGF-1 in E.coli，which provide an evidence for the soluble expression of foreign proteins in E. coli.

Abstract:The gene encoding D-mannose isomerase（MIase） from Escherichia coli（E. coli） BL21 was cloned into expression vector pET-28a（+），and then the recombinant plasmid was transformed into the strain E. coli BL21（DE3）. Efficient MIase intracellular expression by the recombinant E. coli BL21 was achieved with an activity of 4.2 U/mL（D-mannose forming） after IPTG induction for 6 h. The enzyme was identified as a metal-independent protein. The enzyme activity reached the maximum with a conversion rate of 25% at 40 ℃ and pH 7.5. Using D-fructose as the substrate，a Km value of 123.32 mmol/L，Vmax value of 113.64 μmol/（min·mg） and catalytic efficiency kcat/Km value of 0.691 s·mmol/L were estimated，respectively.

Abstract:We constructed a recombinant Escherichia coli which could synthesize Neu5Ac with exogenous GlcNAc as sole substrate in previous work. Various nitrogen and carbon sources were used to investigate their effects on the production of Neu5Ac，and the key limiting factors in fluencing the whole process were revealed. It was shown that the increase of NeuB activity could not improve the Neu5Ac yield and the NeuB activity was not the limiting factor. The intracellular PEP was supposed to be the limiting factor for Neu5Ac production. The intracellular PEP was saved and shifted to the synthesis of Neu5Ac by adjusting the nitrogen source and restraining the cell growth. In addition，by using glycerol as sole carbon source which was PTS-independent，Neu5Ac synthesis in the glycerol/nitrogen-free medium reached （4.42±0.08） g/L，10.3 times higher than the initial synthesis conditions.

Abstract:Characterization of the microflora during malting is an essential step towards process management and optimization. The aim of this study was to characterize the bacterial communities using two culture-independent methods，including Terminal Restriction Fragment Length Polymorphism（T-RFLP） and 454 pyrosequencing，targeting the 16S rRNA gene. In order to analyze bacterial communities and dynamics fully and precisely，the same batch of barley originated from two harvest years using two malting methods was studied in entire bacterial community and lactic acid bacteria（LAB）. The results showed that the bacterial community structure of barley from different years was different and changed during the malting process，but there was no significance difference between the two malting methods. Zooming in the LAB community using 454 pyrosequencing revealed a total of 139 operational taxonomic units（OTUs）. LAB diversity appeared relatively limited since 88% of the sequences were covered by the same five OTUs（representing members of Weissella，Lactobacillus and Leuconostoc） present in all samples investigated. Fluctuations in the relative abundances of the dominant LAB were observed with the process conditions. In addition，both the year of harvest and malting environment influenced the LAB community structure.

Abstract:A strain of s-Trioxane degrading bacterium was isolated from the sediment of Polyoxymethylene wastewater treatment plant，and identified to be Bacillus methylotrophicus by 16S rDNA sequence analysis. The optimum culture conditions of the strain were at about pH 7 and 30 ℃，and the threaholds of maximal tolerable concentrations for NaCl and TOX were 3% and 1 200 mg/L，respectively. In order to detect its degradation characteristics，the strain was mixed together with formaldehyde degrading bacteria，Candida maltosa and Pseudomonas putida，to form multiple strains to treat POM wastewater at the laboratory scale. The treatment results according the order of final effluent quality were：single strain + formaldehyde degradation group > formaldehyde degradation group > single strain group > control group. In single strain + formaldehyde degradation group，the degradation rates of COD，formaldehyde and TOX were 92.80%，96.52 and 95.88%，respectively，which had much better consequences than other groups. There is a synergistic effect of the degrading bacteria combination，which could be used as multiple strains for bioaugmentation treatment of POM sewage.

Abstract:With a large amount of hazardous matter，such as virus，hospital wastewater experiences frequent variation in wastewater quality and quantity. Membrane bioreactor（MBR） technology has been extensively used in industrial wastewater treatment plants. However，membrane fouling always is one of key bottlenecks to obstacle the stable operation of MBR. Aimed at fouled membrane from hospital wastewater treatment plant，the optimal chemical cleaning strategy can be determined based on surface fouling element analysis using energy dispersive x-ray spectroscopy（EDX）. It is helpful for chemical cleaning of fouled membrane modules in wastewater treatment plant. The results showed that，compared with other seasons，the elements in/on fouled membrane surface in autumn season was most complicated，and organic elements were overlapped with inorganic. Under room temperature，organic and inorganic elements were effectively removed by 0.5% citric acid and 0.1% sodium hypochlorite，respectively. Concerning the cleaning order，the better method was firstly citric acid cleaning followed by sodium hypochlorite. The removal of inorganic elements by citric acid made fouling layer softer and was favorable for subsequent sodium hypochlorite cleaning. The element components of cleaned membrane surface were similar to those of new one. Also，the porosity rate，average pore size and contact angle of cleaned membrane with 0.858，0.104 ?滋m and 69.20±0.115° was approximate to those of new one.

Abstract:Protein recovery from waste activated sludge（WAS） is a kind of new way of sludge reutilization. The purpose of this manuscript is to investigate a combined method of enzymatic hydrolysis with thermal alkali hydrolysis for efficiently dissolving proteins from WAS. The results showed that thermal alkali hydrolysis strongly promoted the dissolution of WAS proteins after combined with enzymatic hydrolysis. Compared with single enzyme hydrolysis，the protein dissolution rates hydrolyzed by lysozyme and papain were increased by 46.84% and 45.56%，respectively. Combined with thermal alkali hydrolysis，the multi-enzymes with alkaline protease and papain got the highest dissolution rate of WAS proteins. With 30 g/L of WAS and 1% of multi-enzymes（alkaline protease∶papain=1∶12），the dissolution of WAS proteins was optimal after WAS was enzyme-hydrolyzed for 2.0 h under pH=7.0 and 55 ℃，and then experienced thermal alkali hydrolysis under pH=12.5 and 90 ℃ for 1.5 h. Accordingly，the soluble protein concentration and dissolution rate were 6 050 mg/L and 84.33%.

Abstract:In order to study the optimum conditions of B. brevis producing levansucrase and the optimum transformation conditions for the conversion of sucrose-lactose to lactosucrose by recombinant enzyme. A recombinant strain containing the encoding gene of Bacillus flexus levansucrase，B. brevis/pNCMO2-Lsc had been constructed in our laboratory previously. The optimum fermentation medium was determined by single factor test and orthogonal test：20 g/L of glucose，40 g/L of nitrogen source（industrial yeast powder：cottonseed powder=2∶1），0.5 mmol/L of CaCl2，The optimal temperature for the enzyme production was 30 ℃. Under these conditions，the production of levansucrases reached 62.1 U/mL，which was 3.69 times higher than the original enzyme activity. The crude enzyme was used for the preparation of lactosucrose. Under the condition of sucrose and lactose concentration of 200 g/L，the optimum conditions were as follows：reaction temperature 35 ℃，pH 6.0，enzyme loading 2 U/g substrate，reaction time 8 h. Reaching 39.1%.

Abstract:Sucrose isomerase from the recombinant strain was immobilized on chitosan cross-linked with glutaral-dehyde. The effects of chitosan concentration，glutaraldehyde content，enzyme concentration，crosslinking time were investigated using the immobilized enzyme activity recovery rate as an index.，and the effects of temperature，pH，enzyme dosage，reaction time and substrate concentration on the production of isomaltulose by immobilized sucrose isomerase were also investigated. The immobilization efficiency reached up to 70.3% under the optimal condition of immobilization achieved with 3.0% of the chitosan，0.75% glutaraldehyde content，50 U/g chitosan beads and 16 h. The optimum conditions of the synthesis of isomaltulosewere as follows：reaction temperature 30 ℃，initial pH 4.5，enzyme concentration 15 U/g sucrose，conversation time 12 h，initial sucrose concentration 600 g/L. When the reaction was performed under this specified conditions，the isomaltulose yield reached the maximum of 87.8%. The yield ratio was still up to 87.52% after 16 times of continuous conversion and it shows a good operational stability and a high productivity of isomaltulose.

Abstract:Response surface methodology was used to optimize the fermentation medium based on the medium MRS to increase the viable bacteria count of B. adolescentis. Meanwhile，the growth curve and pH were compared before and after optimization. According to the results of response surface analysis，the enrichment medium components were determined as follows：glucose 15.85 g/L，fructo-oligosaccharide 15.85 g/L，peptone 14.45 g/L，beef extract 8.67 g/L，yeast extract power 5.78 g/L，ammonium citrate 2.75 g/L，K2HPO4·3H2O 2.75 g/L，sodium acetate 6.875 g/L，Tween-80 1.0 mL，MgSO4·7H2O 0.2 g/L，MnSO4·H2O 0.05 g/L，L-cystein-HCl 0.5 g/L. Cultivated in optimized enrichment medium，B.adolescentis reached stationary phase at about 28 h，which was shorter 12 h than before； the viable counts reached up to 8.9×109 CFU/mL，which was 1.73 times as many as before.

Abstract:The extraction and development of polysaccharide was researched to multipurpose use of cordyceps militaris culture medium with wheat. This article studied the optimum extraction process of coarse polysaccharide in cordyceps militaris culture medium through the orthogonal factor experiment on hot reflux water on the base of single factor experiment on hot reflux water and ultrasonic extraction，researched the antitumor activity of polysaccharide purified by cordyceps militaris culture medium by MTT method. The results showed that hydrothermal refluxing extraction is better than supersonic water extraction on the basis of the results of single factor experiment. The results of orthogonal test showed the optimal technological conditions for the extraction conditions of coarse polysaccharide of were as follow：extracting temperature was 100 ℃， material-water ratio was 1∶25，extracting times was three and time was 90 min. The primary and secondary relationships of four Factors influencing the efficiency of extraction coarse polysaccharide are：extraction temperature > extraction time > extraction times > solid-liquid ratio. The coarse polysaccharide yield increased with the increasing of concentration of alcohol. The MTT test found the polysaccharide purified by different processing has inhibitory effect on the liver cancer cells，5 mg/mL concentration of polysaccharide，effect time for 72 h，inhibition rate of polysaccharide gained by degreasing and deproteinization from culture medium of Cordyceps militaris for hepg2 cells can reach 83.02%，inhibition rate of polysaccharide gained by degreasing medium for hepg2 cells can reach 75.39%，inhibition rate of polysaccharide gained without degreasing and deproteinization for hepg2 cells can reach 30.61%，purified polysaccharide showed higher inhibition rate. The polysaccharide extracted by cordyceps militaris fruiting body with 100 ℃ hot reflux water ，5 mg/mL concentration of polysaccharide，training after 72 hours，the inhibition rate reached 92.974% on Hepg2 cell，there was no significant difference compared with SB，but was significant difference compared with polysaccharide gained by degreasing and deproteinization from culture medium of Cordyceps militaris.

Abstract:To study the effect of suitable dietary iron supplement on the quality of fillet，surimi and Cathepsin B/L（CAT B/L） of Jian carp（Cyprlnus carpio var Jian）. The fish fed for 90 days with the supplemented diets containing iron of 146.1 mg/kg（suitable group） was collected as experimental group，and those fed without supplemented diets containing iron of 53.9 mg/kg（lacking group） was the control. After sampling，the quality indexes of fillets and the activities of CAT B/L were detected. Then the protein expression level of CAT B/L was quantified by immunohistochemical analysis. Further，the fillet was processed into surimi，and its stability of each group was measured. The results indicated that the MBF and MFI in suitable group increased or declined slightly（P>0.05） when compared to lacking group. While the shear force of suitable group increased 31.80%（P<0.01）；the contents of soluble collagen，insoluble collagen and the total collagen increased 305.45%，180.98%，219.35% respectively（P<0.01）. The pH and water loss rate of suitable group was in the trend of decline（P>0.05）. Moreover，the CAT B/L activities of suitable group was less than the lacking group to the degree of extreme significance（P<0.01） and significance（P<0.05） respectively. Meanwhile，the immunohistochemical analysis indicated that in both groups，CAT B/L were expressed in the muscle tissue and the expression level of suitable group was less than the lacking group extremely significantly（P<0.01）. Additionally it was found that suitable dietary iron supplement could increase the initial CA，TSH（P<0.01），and decrease the initial DB（0.01

Abstract:In order to increase the contents of the active ingredients，improve the functional activity and broaden the utilization pathways of wheat bran，wheat bran was fermented with three fungi （Aspergillus niger，Aspergillus oryzae，Trichoderma reesei） and free phenolic content，free phenolic composition and antioxidant activity were investigated by spectrophotometry and HPLC. The results showed that three fungi all could degrade wheat bran and increase the contents of total free phenol，phenolic acids composition and antioxidant activity. The effect of Aspergillus oryzae was the most obvious among the three fungi. In conclusion，fungal fermentation is an effective way to improve bioactive ingredients of wheat bran and Aspergillus oryzae is an excellent choice for fermentation of wheat bran.

Abstract:In order to study the conditions of chelating Znic with α-glucosidase inhibitory active peptides from Porphyra yezoensis enzymatic hydrolysis and evaluate the stability of gastrointestinal digestion The polypeptide，inhibiting the α-glucosidase activity highly，would be gained by the compound enzymolysis technics with using protease of ex- and in- under various treatment factors. Thereafter，the hydrolysate was separated by filtering membrane technique combined ultrafiltration with nanofiltration firstly，and then concentration through rotary evaporator，which the inhibition ratio of α-glucosidase（0.5 mg/mL） could be reached 68% at 1 mg/mL of concentrated polypeptide solution；The optimization factors of chelating reaction involved polypeptide and zinc solution were 1.5 h of reaction time，37 ℃ of temperature，6 mg/ml of polypeptide and pH 4.5 respectively，which the chelating degree of polypeptide and zinc had reached 25.6%， and the inhibition for α-glucosidase had increased to 79.8% after chelation；In addition，the model of gastrointestinal digestion in vitro would be built to evaluate the toleration of the chelate for gastrointestinal digestion by assessing their capacity to inhibit the α-glucosidase activity. The results indicated that the decrease of α-glucosidase activity only had 7% after gastrointestinal digestion under various reaction time and enzyme-substrate ratio；and then reaching 5% approximately after duodenal digestion. Therefore，the chelate of polypeptide and zinc solution was shown better stability of gastrointestinal digestion in this study.

Abstract:In order to study the flavor changes of kimchi in fermentation process，we firstly measured the lactic acid bacteria contents，the pH values and the total acidities of kimchi carrot and kimchi cabbage. Then we compared the difference in odour and taste by using e-nose and e-tongue，respectively. In addition，we compared flavor features of the two kinds of kimchi by PCA（principal component analysis） and LDA（linear discriminant analysis）. The result shows that the lactic acid bacteria content of Korean carrot kimchi arrived at the maximum value after fermentation of 6 days，and that of Korean cabbage kimchi achieved the maximum value on fourth day of fermentation. Both the lactic acid bacteria contents of the two kinds of kimchi declined rapidly after reaching the maximum value. The lactic acid bacteria could be detected after fermentation of 3 days. The pH value of Kimchi Carrot after 6 days fermentation and Kimchi Cabbage after 4 days fermentation was 3.40 and 3.48 respectively，and their total acidity was 1.26% and 0.51% respectively. Significant difference in flavor change with fermentation time was observed between Korean carrot kimchi and Korean cabbage kimchi. Odor characteristics of the two kinds of kimchi were very close to each other. The kimchi can still be classified into four categories according to their discrepancy of odor characteristics. In terms of taste characteristics，there was remarkable difference between the two，especially in the salty value，bitterness value and astringency value. These values of Korean cabbage kimchi were larger than those of Korean carrot kimchi. Results indicated that kimchi differed obviously in favor characters such as odor and taste features，even under the same process and fermentation conditions. And the flavor characters changed with fermentation time，type of raw material and other factors.