Abstract:Biomass pigments are better than synthetic dyestuffs which are applied currently，especially the aspects concerning sustainability of source，environmental conservation during manufacture as well as ecology dyed textiles. Biomass pigments are synthesized from organism，and microorganisms pigments are more potential than dyestuffs extracted from animals and plants in the aspect of industrialized application. The paper reviews the preparation of microbial synthesis with respect to some kinds of significant biomass pigments，analyzes the precursors of biosynthesis of dyestuffs，presents microorganism species that could synthesize pigments，elaborates the biosynthesis pathways of pigments as well as relevant enzymes，analyzes synthetic efficiency of biomass pigments as well as application condition currently，and looks into the perspective biomass dyestuffs via bioconversion.

Abstract:Homology modeling of tyrosinase was performed using the SWISS-MODEL based on the known crystal structure of tyrosinase from S. castaneoglobisporus（PDB No. 2AHL）. The POPMuSiC algorithm was applied to predict the folding free energy change （ΔΔG） of amino acid substitution. Site-directed mutagenesis was used to construct mutants at Arg95Tyr and Gly123Trp. The mutant and wild-type enzymes were expressed in Escherichia coli（DE3）. As compared to the wild-type tyrosinase，all the mutant enzymes showed improved thermal properties. The mutant with combined substitution（Arg95Tyr/Gly123Trp） showed the most pronounced shifts in temperature optima，which was about 5 ℃ upward，and the half-life for thermal inactivation at 60 ℃ was increased by 1.5 times.The structure-based rational design strategies in this study may also provide further insight into the thermostability of other industrial enzymes and suggest further potential industrial applications.

Abstract:The main focus of this study is to evaluate the effects of dietary supplement of EPA and DHA in vivo. The design of this study including the synthetic diets for mice，which were based on the balance of total energy，comprised the ω-3，ω-6 and control groups，and oral supplementation of fish oil for human；and to analyze the absorption efficiency of EPA and DHA in mice and in human after specified dietary supplementation. Our study showed that before and after one week dietary supplementation，the increasing ratio of EPA and DHA in blood of ω-3 group were 9.1 fold and 1.99 fold respectively. Compared to ω-6 group，the increase ratio of EPA and DHA in ω-3 group were 66 and 3 fold in liver，8.1 and 4.9 fold in epididymal fat pad respectively. Compared to Ctl group，the increasing ratio of EPA and DHA in ω-3 group were 35 and 3.5 fold in liver，8.1 and 5.8 fold in epididymal fat pad respectively. Human study also showed that oral administration of fish oil led to an increase of 4.4 and 1.4 fold for EPA and DHA respectively. Relative to DHA，the change of EPA level in body was more subjected to dietary fatty acid supplementation. Thus，EPA levels in mice and human are more subject to the dietary supplement compared to DHA level，probably due to the insufficient EPA intake or high turnover ratio in vivo，supplementation of fish oil to increase EPA levelin vivo is effective and probably necessary.

Abstract:Superoxide dismutase （SOD） can be widely used in the field of food，cosmetic and medicine for its high capacity to scavenge free radicals. This paper reported a method of gene engineering attempting to solve the issue of SOD production. The gene of Cu，Zn-SOD with the yeast bias codon was firstly chemically synthesized and integrated into the genome of Pichia pastoris and the expression condition was optimized through orthogonal experiment. It was found that the maximum expression level of SOD was nearly three times greater than some reserach reported and the optimal temperature，methanol concentration and inoculation were 28 ℃，1.0% and 1.0% respectively. These results suggest that the problem of SOD production can be solved by improving the expression level of P.pastoris through the optimization of gene codon and fermentation conditions.

Abstract:In order to enhance the caproicacid productionof Clostridium kluyveri and shorten thefermentation period，this strain was co-colturedwith Saccharomyces cerevisiae and themechanism of enhancing caproic acid production in the mixed culture system was furtherly analyzed. Studies have revealed that：caproicacid yieldin the fermentation tank with pure colture of Clostridium kluyveri reached 6.35 g/L. However，caproic acid production with co-cultured system reached 7.09 g/L，which was 12.5% higher than that of the pure culture system and thetime reaching peak of caproic acid production was shortenedfor approximately 48 h. Relative data indicated that when co-culture wit Saccharomyces cerevisiae，thecaproic acid production of Clostridium kluyveriwas improved byincreasing the conversion rate of acetic acid to butyric acid. Analysis of dissolved oxygen values and microbial growth parameters indicated that the main reason for this change was that the growth of Saccharomyces cerevisiae improved the anaerobiclevel of co-culture system，which promoted the formation and transformation of butyric acid，hence the caproic acid production was enhanced.

Abstract:The biosynthesis of GPI is carried out in the endoplasmic reticulum（ER） through multiple steps and many genes are involved in the reactions. PIGK is one of the subunits of the catalytic domain of the GPI transmidase complex，responsible for the transfer of GPI moiety on protein. In this study，we generated a PIGK-knockout（KO） HEK 293 cell line，which showed no surface expression of GPI-anchored proteins（GPI-APs）. By mediation of piggyBac（PB） transposon system，we rescued the PIGK gene in PIGK-KO cells，which showed the recovery of the surface expression of GPI-APs. We successfully established a HEK 293 cell line，HEK293pB-PIGK，named G36，which can be converted from GPI-AP positive to negative cells by the introduction of PB transposase or FLP recombinase. Our system is useful to modulate surface proteins quickly and can be applied for both basic and applied researches.

Abstract:To prepare the self-assembly hollow nanospheres of hyaluronic acid-graft-poly（ethylene glycol）（HA-g-PEG） and hydroxypropyl-beta-cyclodextrin（HPCD） containing uricase（U HPCD） and study their stability in vitro. UHPCD were prepared and the entrapment efficiency，size and zeta potential of UHPCD were detected. The optimum temperature，optimum pH，thermal stability，storage stability，pH stabilities and proteolytic stabilities of UOX and UHPHD were measured. The entrapment efficiency of UHPHD was（62.17±2.94）%. The average particle size was（299.60±13.05） nm，and the zeta potential was（-45.10±2.75） mV. The optimal temperatures for UHPHD and U0X were 40 ℃，the optimal pH values were 8.5. UHPHD significantly enhanced the stability of free UOX in vitro.

Abstract:A family 27 carbohydrate-binding module（CBM27） of Thermotoga maritima MSB8 was linked to the C-terminus of a glycoside hydrolase family 5 β-mannanaseof Aspergillus usamii YL-01-78（AuMan5A） by using a flexible peptide linker F（GGGGS）3 ora rigid peptide linker R（EAAAK）3. By overlapping PCR technique，twofusion enzyme genes，Auman5A-F-cbm27 and Auman5A-R-cbm27，was amplified. The gene Auman5A and two fusion genes were expressed in Pichia pastoris GS115，respectively. And then，the enzymatic properties of the recombinant β-mannanases，reAuMan5A，reAuMan5A-F-C and reAuMan5A-R-C，were analyzed. As a result，the Km ofreAuMan5A-R-C towards locust bean was 0.9 mg/mL，which was much smaller than those of reAuMan5A（1.7 mg/mL） and reAuMan5A-F-C（1.9 mg/mL）. The temperature optimum（Topt） of reAuMan5A-F-C was 65 ℃，which was 5 ℃ lower than that（70 ℃） of reAuMan5A or reAuMan5A-R-C. The half-lives at 70℃（t701/2） of reAuMan5A-F-C and reAuMan5A-R-C were 36 and 124 min，respectively，which were about 4- and 13.8 folds longer than that（t701/2=9 min） of reAuMan5A. In this study，the fusionenzymereAuMan5A-R-C was obtained with strong substrate affinity，high thermostability and wide range of pH stability. It will be a potential candidate for industrial processes，such as food，animal feed，energy，pharmaceuticals and so on.

Abstract:Optically pure 2，3-butanediol is one of the important building blocks. Bioreduction of diacetyl using diacetyl reductase is an environmentally friendly method for chiral 2，3-butanediol production. The medium components and induction conditions of diacetyl reductase produced by recombinant Escherichia coli were optimized in flasks. The optimal medium composition for the production of diacetyl reductase was listed as follows：glycerol 5 g/L，yeast extract 10 g/L，peptone 5 g/L，（NH4）2SO4 1.5 g/L，sodium citrate 5 g/L，KH2PO4 1 g/L，MgSO4·7H2O 0.5 g/L and NaCl 2 g/L. The optimal induction conditions were listed as follows：37 ℃ and 0.1 mmol/L IPTG. After induced for 6 h under the optimized conditions，the enzyme activities reached to 13.84 kU/g dry cell weight and 33.65 kU/L，which were 2.04 and 4.23 folds of those in basic medium，1.16 and 1.71 folds of those in LB medium. In a 3 L fermenter，under the optimized conditions，the maximum enzyme activities achieved at 14.09 kU/g and 64.62 kU/L after cultured for 8 h，which were both higher than those in flasks. This study provides useful guidance on the highly efficient preparation of diacetyl reductase by recombinant Escherichia coli for the preparation of chiral 2，3-butanediol.

Abstract:Lots of antibody drug have been successfully expressed in the prokaryotic system. Lacking knowledge on the process mechanism of recombinant protein expression has forced researchers to use "trial and error" to improve soluble production，which is time and labor consuming with low success rate. Adding different amino acids to the C-terminal of a dAb often results in significant variation of soluble expression levels. The amino acid sequence of dAbs highly concords with their soluble expression levels，with a consistency being 70%. Through analyzing the soluble expression and sequence data of 65 dAbs using clustering and linear modeling，we show that certain amino acids panel could significantly affect the soluble expression of dAbs，with the specific amino acids composition in these panels being（S，R，N，D，Q），（G，R，C，N，S） and（R，S，G），respectively，in the supernatant，pellet and total amount. In addition，polar was found a vital factor affecting the soluble production of dAb. These results may be helpful in orthomutation.

Abstract:Dendrobium officinale is a precious medicinal plant containingmany bioactive compounds and its stem was the main subject for structure and bioactivity research. In this study，the types and contents of active compounds in D. officinale stem and leaf were compared. Although D. officinale stem and leaf had similar compounds，the content of polysaccharide and dendrobine in stem was 2.71 and 3.02 times as high as in leaves，while the content of fat-soluble composition and crude protein in leaves was significantly higher than that in stem. After monosaccharide composition analysis of purified stems and leaves polysaccharide，D. officinalestem polysaccharide（DOSP） was composed of mannose and glucose with mole ratio of 3.07：1. However，D. officinale leaf polysaccharide（DOLP） mainly contained mannose，glucose，galactose and arabinose with mole ratio 8.81∶1∶0.57∶0.37. In addition，the intrinsic viscosity and of DOSP was 56.26 dL/g and DOLP was 8.05 dL/g，while the corresponding molecular weight of DOSP and DOLP were 1 207 000 and 318 000，respectively. The results of free radical scavenging capacity shows that DOLP has higher DPPH free radical scavenging capacity than DOSP indicating a high potential for applied as theantioxidant. Finally the extracted condition of DOSP and DOLP was optimized by comparing various temperatures，extracted times and solid-liquid ratio to enhance the yield of this active compound.

Abstract:In this work，the taste of fried rabbit meat products was mproved by enzymatic pretreatment process and using amendment. The enzymatic pretreatment was carried out by using papain and trypsin. By using one-factor-at-a-time and orthogonal array design methods，the optimal enzymatic pretreatment conditions were obtained as follows：the dosage ratio of papain and trypsin 300∶150，temperature 45 ℃，time 60 min. The product quality was improved by using soy protein，modified starch and baking soda，the optimal modifier formula was determined as 0.6%baking soda，10%modified starch and 4%soy protein. The sequence of the significance of these factors is as follows：soy protein dosage>modified starch dosage>baking soda dosage.

Abstract:Aspergillus niger glucose oxidase（GOD） was expressed in Pichia pastoris SMD1168 with the alcohol oxidase gene promoter（AOX1）.A gene of glucose oxidase from Aspergillus niger PCTC was cloned.The gene was fused to the pPICZαA plasmid which had the alcohol oxidase gene promoter（AOX1） and expressed in Pichia pastoris SMD1168.After screening by zeocin gradient resistance plate ，shake culture，and the SDS-PAGE analysis of proteins in the culture，one strain was obtained，which gave the higest enzyme activity（1.12 U/mL） after one day induction by 1% methanol.Through the optimization of the shake flask experiments ，the highest enzyme activity of 32 U/mL was achieved in the flask by induction for 7 d under optimal conditions（30 ℃，200 r/min，pH 5 and 1.5% methanol）.

Abstract:KEGGonline database and six subcellular prediction databaseshave been studied for the process of auto-refinement. The weighted scoring mechanism was proposed to analyze the results of subcellular prediction databases，using image processing algorithm to determine high credibility specific reaction.As an illustration example，all of the automatic methods were implemented in the process of genome-scale metabolic network refinement of Spathasporapassalidarum NRRLY- 27907，whichconfirmed that these methods can improve the efficiency of model reconstruction.

Abstract:Artemisia argyi polysaccharide extracted by hot water and measurement the antibacterial activity and minimum inhibitory concentration（MIC） of polysaccharides by using the filter paper method，also studied the pH stability，thermal stability，UV stabilizers stability of Artemisia polysaccharide antibacterial activity. The results showed that：Artemisia argyi polysaccharides have a certain inhibitory effect on three common bacteria，and have strong effect on gram-positive bacteria Staphylococcus aureus and Bacillus subtilis，but the effect on gram-negative bacteria E. coli is weak.The MICs of the polysaccharide agaginst Bacillus subtilis，Staphylococcus aureus and Escherichia coli were 0.625、0.625、1.25 mg/mL respectively. The antimicrobial activities of the polysaccharide were stable under the conditions of heat and ultraviolet ray，but were unstable under different pH values，strong in acid and weak in base. Artemisia argyi polysaccharide has good antibacterial activity，it can be used for the development and utilization of natural food preservatives.

Abstract:Method of multi-enzyme was used to optimize the extraction process of polysaccharide from Ploygonatum sibiricum，phenol-sulfuric acid colorimetric method was applied to determine the content of polysaccharide，defined the polysaccharide extraction rate as the index to choice the kind and ratio of enzymes，the experimental factors were selected by single factor trials such as enzymatic hydrolysis temperature，pH，the rate of material to solvent，the amount of enzyme concentration，the important ones of which would be chosen to optimize by orthogonal test. Results showed that multi-enzyme extraction was better than extraction by single-enzyme or water. The rate of cellulose to papain was 3 to 7. The optimum enzymolysis conditions were as following：pH value of 5.0，temperature of 50 ℃，the rate of material to solvent was 1 to 20，the amount of enzyme concentration was 1.5%，which was 0.45% of cellulose and 1.05% of papain，2 hour-hot water extraction after 2 hour-enzymatic hydrolysis. In this condition，the maximum polysaccharide rate was 21.55%，which was 2.75 times as high as water，and had 12.6% higher than that of single factor trials.

Abstract:In the paper influence of particles size on protein sedimentation rate from cow and goat after heat treatment（95 ℃，15 min）were explored. Centrifugal sedimentation，atomic force microscope，laser particle size analyzer，and SDS-PAGE methods were used to determine protein precipitation rate，and protein surface morphology after heat treatment，casein composition，and protein particle size of fresh milk respectively. The results showed that protein sedimentation rate of goat milk was significantly higher than that of cow milk（P < 0.05），protein particles size of goat milk were significantly bigger than that of cow milk，the clumping together degree（35 858.15 nm2） of goat milk protein were bigger than that （18 215.18 nm2） of cow milk，fresh cow milk protein particle size mainly focused on 1 nm or less，that of fresh goat milk protein mainly distributed during the range of 1～1 000 nm，fresh cow milk and goat milk have two kinds of casein mainly，beta casein and alpha s1-casein，and their molecular weight are 34 000 and 26 000 respectively. Thermal stability of cow milk is better than that of goat milk. Protein particle size is the direct influence factor on the thermal stability of milk protein. Atomic force microscopy combined with laser particle size analyzer is an effective way to analyze the particle size of milk protein.