Abstract:Resources shortage, energy deficiency and environmental pollution have been hindering the sustainable development of the society. As “the third wave” in the course of the development of biological technology, industrial biotechnology has become an effective way to solve the problems of resources, energy and environment with which mankind is confronted. It is also one of the most promising technologies indispensable to the sustainable development of the industry. In this special issue of “Industrial Biotechnology”, selected papers from China Summit Forum on Industrial Biotechnology Development · 2008 were published, hoping to promote the academic exchanges and development of China’s industrial biotechnology. The papers focused on the researches in the fields of “Biorefinery process and biobased products”, “Microbial genomics and bioinformatics”, “Metabolic engineering and development of antibiotics”, “Industrial enzyme technology”, “Microbial cell factories and biorefinery”, “Biocatalysis and biotransformation”, “Bioprocess technology”, and “Industrial strain breeding and improvement”.

Abstract:As a clean energy source and industrial material, hydrogen is very valuable. Electrolysis of water and chemical methods are well-known for producing hydrogen, however, all of these methods need additional energy supply. Besides highly energy cost, the chemical methods will lead to serious environment pollution. Compared with traditional methods, biological production of hydrogen has showed significant advantages. Bio-hydrogen can be produced by anaerobic and photosynthetic microorganisms during treatment of organic waste. It provides a low cost method for producing hydrogen gas, and a way of utilizing waste at the same time. This paper summarized the procedures of treatment of solid waste and the production of bio-hydrogen.

Abstract:Indigo and indigo-like pigments are widely used in the industry of textile, food and medicine. Now people pays more and more attention to developing an alternative method of indigo production which is “environment-friendy”, especially microbial biosynthesis of indigo. Many microorganisms involved in the biosynthesis of indigo have been isolated and characterized, and monooxygenase and dioxygenase have been identified to catalyze indigo biosynthesis. Some genes encoding for these enzymes have been cloned and used to construct “engineering bacteria”. With this kind of bacteria, more efficient fermentation systems for indigo production have been exploited. In the meantime, biotransformation of the indigo produced by microorganisms has been under investigation. These progresses will bring us a greener method of indigo and indigo-like pigments production.

Abstract:Vitamin B12 is an important nutrient widely used in feed, food and medicine field. China is the primary producing area and the VB12 production is 27 t in 2007, 77% of total production in the world. VB12 is the most complex small molecule difficult to chemosynthesize. It ismanufactured by bacteria and archaea via two alternative routes, aerobic or anaerobic pathway. The main strains used in industry fermentation are Propionibacterium freudenrechii and Pseudomonas denitrificans. The basic characteristics, biosynthesis and fermentation of vitamin B12 are reviewed. The vitamin B12 application and market are also summarized.

Abstract:Anaerobic fermentation bio-hydrogen production has captured extensive attention, hydrogen-production microorganisms has become the research focus as core role. Based on the review of current status and main achievements of hydrogen-producing microorganisms research both domestic and abroad, the fermentative type, the hydrogen-production capability, the bacterium type, breeding, and the gene modification were presented. The main associated issues were analyzed and the research prospects were put forward.

Abstract:The concept of Industrial Biosystems Engineering (IBsE) was suggested as a new engineering branch to be developed for meeting the needs for science, technology and professionals by the upcoming bioeconomy. With emphasis on systems, IBsE builds upon the interfaces between systems biology, bioprocessing, and systems engineering. This paper discussed the background, the suggested definition, the theoretical framework and methodologies of this new discipline as well as its challenges and future development.

Abstract:The energy consumption in a Chinese ethanol manufacturer with cassava as the feedstock, has been reduced to a zero-closed level. If the R & D on technical integration of high ethanol concentration fermentation, methane fermentation technique, steam and electricity co-generation system, new distillation technology, and the wastewater reutilization, is carried out continuously, the proposed “zero energy consumption and wastewater” technique could be realized in fuel ethanol production process.

Abstract:One yeast strain, which was isolated from 256 natural samples, was found to be able to utilize D-xylose effectively. On the basis of assimilation physiological and molecular biological tests, the yeast strain was identified as a strain of Candida tropicalis. Furthermore, metabolic engineering breeding strategy was applied to change the metabolic flux in order to increase ethanol productivity. In this study, the C. tropicalis was used as the host strain and the plasmid pYX212-XYL2, which was formerly constructed for over expression of XYL2 gene encoding xylitol dehydrogenase (XDH) from Pichia stipitis, was used as the backbone of the recombinant vector. A hygro gene was inserted into downstream position of XYL2 gene, meanwhile, the result plasmid pXY212-XYL2-Hygro transformed into C. tropicalis by electroporation. Thus, a recombinant yeast C. tropicalis XYL2-7 was obtained through hygromycin B resistance screening and its specific XDH activity was 0.5 u/mg protein, which was 3 times more than that of the parent strain. Additionally, the recombinant yeast was applied in the fermentation of xylose. Compared with the parent yeast, it was concluded that the xylitol yield in the broth decreased by 3 times, however, the ethanol yield increased by 5 times. The feasibility of ethanol production from xylose by C. tropicalis was firstly studied in this paper. These research results are helpful to advance the bioconversion of renewable resources (e. g. straw, wheat bran, and husk) to fuel ethanol.

Abstract:The gelatinization process of the starch is replaced by unpolluted steam-pretreatment on the base of the Radix Puerariae rich in fiber and isoflavones. The production of ethanol and isoflavones by simultaneous saccharification and solid state fermentation (SSF) of steam-pretreatment Radix Puerariae is presented. The optimal technological conditions were obtained: Radix Puerariae being steam-pretreated at a saturated vapor pressure of 0.8 MPa for 3.5 min, glucoamylase(65 u/g), cellulase(1.5 u/g), 0.1%(NH4)2SO4, 0.1%KH2PO4 and activated yeasts being added in, and fermentation at 35~37oC for 60 h. Under these conditions, the yield of ethanol and isoflavones from 100 g Radix Pureriae (dry basis) were 27.47 g and 4.43 g, respectively, the starch utilization rate was 95%. In comparison with the traditional fermentation technology, the simultaneous saccharification and SSF of steam-pretreatment Radix Puerariae is clean and energy-saving. It provides new way of the production of ethanol from the non-food starch material, and worthwhile to be explored and implemented in industry.

Abstract:For the purpose of revealing the mechanism of the reduction of yeasts ethanol production rate after entrance of post-log phase, we used microarray to study expression profiles of the yeast Saccharomyces cerevisiae during the transition from mid-log growth phase to post-log growth. The results demonstrate that the global pattern of gene expression is very stable during the mid-log phase. However, a dramatic metabolic remodeling was found when the yeast entries post-log phase, during which many of amino acid synthesis and metabolism related genes are up-regulated, moreover, ion transport, energy generation and storage related genes are also up regulated during this phase, while a large number of genes involved in transposition and DNA recombination are repressed. Central metabolic pathways also engage in metabolic remodeling, within which the genes involved in succinate and α-ketoglutarate synthesis pathways are up regulated, accordance with those of amino acid synthesis and metabolism. These results demonstrate that the increasing demand for amino acids in post-log phase lead to a metabolic transition into TCA cycle and glyoxylate cycle, which subsequently reduce the ethanol production rate. This suggests a global insight into the process of yeast ethanol fermentation.

Abstract:Biohydrometallergy technology received more and more attention because of its simple process, low cost and kind to environment, especially in dealing with low-grade and complex minerals. However, it is difficult to optimize microorganism species and process parameters in bioleaching procedure because of the lack of suitable bacteria and quantitative analysis methods at micro-level for bioleaching system. This has resulted in the low efficiency and poor yield of the target metal in bioleaching. With the development of microarray and bacteria conservation technology, solutions to the above problems were being found. This article summarizes the latest findings on genetic elucidation and the community structure of microorganisms in sulfide minerals bioleaching system, in the aim of providing a better understanding on the significance of cross-field technology of biohydrometallergy and genomics.

Abstract:Six hundred and eight fungi strains were isolated from seventy-eight samples of mangrove plants and soil that collected from Qinglan harbor. Cyctotoxic activity was detected by observing the growth inhibition or killing of the tumor cells under microscope. The result showed that 81 strains (about 13.32% of the total strains isolated) displayed cytotoxic activity against B16 tumor cell. The most fungi strains were isolated from mangrove plant Sonneratia alba, and most of cytotoxic active fungi strains were isolated from mangrove plant Heritiera littoralis.

Abstract:Here we report a systematic method for constructing a large scale kinetic metabolic model and its initial application to the modeling of central metabolism of Methylobacterium extorquens AM1, a methylotrophic and environmental important bacterium. Its central metabolic network includes formaldehyde metabolism, serine cycle, citric acid cycle, pentose phosphate pathway, gluconeogensis, PHB synthesis and acetyl-CoA conversion pathway, respiration and energy metabolism. Through a systematic and consistent procedure of finding a set of parameters in the physiological range we overcome an outstanding difficulty in large scale kinetic modeling: the requirement for a massive number of enzymatic reaction parameters. We are able to construct the kinetic model based on general biological considerations and incomplete experimental kinetic parameters. Our method consists of the following major steps: 1) using a generic enzymatic rate equation to reduce the number of enzymatic parameters to a minimum set while still preserving their characteristics; 2) using a set of steady state fluxes and metabolite concentrations in the physiological range as the expected output steady state fluxes and metabolite concentrations for the kinetic model to restrict the parametric space of enzymatic reactions; 3) choosing enzyme constants K’s and K’eqs optimized for reactions under physiological concentrations, if their experimental values are unknown; 4) for models which do not cover the entire metabolic network of the organisms, designing a dynamical exchange for the coupling between the metabolism represented in the model and the rest not included.

Abstract:In the current work, the fusion gene including somatostatin (SS) and the hepatitis B surface antigen gene was cloned into a balanced lethal system plasmid (pYA3493), and then transformed into asd? attenuated Salmonella choleraesuis C500 strain, the positive transformant without antibiotic resistance gene was confirmed by restriction analysis and DNA sequencing, designated as pYA-SS. The expression and immunogenicity of fusion protein were detected by SDS-PAGE and Western blot analysis. These results show that the recombinant prokaryotic expression plasmid pYA-SS could express the SS fusion protein with good immunogenicity in C500 strain. In above all, this study could provide reliable materials to develop novel, good and safe vaccine in enhancing the growth of animals.

Abstract:Glutinous rice wine is a traditional food in south of China and it can coagulate milk. It has been proved that its function of coagulating milk is because of the presence of milk-clotting enzyme produced by microorganisms in glutinous rice wine. The aim of this work is to isolate milk-clotting enzyme producing strain from glutinous rice wine and study the fermentation condition. We screened out four bacteria and fungus by gradient dilution. It was proved that mold played the most important role in the production of milk-clotting enzyme. This is further confirmed by casein plate method. The optimization of fermentation conditions revealed that two times concentrated potato medium supplemented with 5% glucose without additional nitrogen was better for production of the enzyme. The enzyme activity was increased 144% under the conditions established.

Abstract:D-amino acid oxidase (DAAO) is one of important industrial enzymes. To increase the solubility and activity of the TvDAAO from Trignoposis variabilis expressed in recombinant Escherichia coli (E. coli), a maltose binding protein (MBP) and Vitreoscilla hemoglobin (VHb) was introduced to fuse with N-terminal of the TvDAAO, respectively. Fusion protein of MBP-TvDAAO was constitutively expressed in JM105/pMKC-DAAO and inductively expressed in JM105/pMKL-DAAO. With respect to the control strain of BL21 (DE3)/pET-DAAO without MBP fusion, the constitutive fusion expression obtained 28% of soluble protein with 3.7 folds of solubility improvement. As for the inductive fusion expression, corresponding results changed to 17% and 1.8 folds, respectively. However, the DAAO activity significantly decreased in the MBP-fusing expression. Fusion protein of VHb-TvDAAO was constructed and inductively expressed in BL21 (DE3)/pET-VDAAO. Its DAAO activity highly reached 3.24 u/mL in flask culture, about 90% increase in contrast to the control without VHb.

Abstract:Global transcription machinery engineering (gTME) was employed to engineer xylose metabolism. Mutation of the transcription factor gene Spt15 was introduced by error-prone PCR, followed by screening on media using xylose as the sole carbon source. One recombinant strain growing well on such media was chosen for further research. This strain showed modest growth rates in the media containing 50 g/L xylose or glucose at the condition of 30oC, 200 r/min, 96 h, 94.0% and 98.9% of xylose and glucose were consumed, with the ethanol yield were 32.4% and 31.6%, respectively. The control strain had the ethanol yield of 44.3% under the glucose concentration of 50 g/L. When the carbon source was 50 g/L glucose/xylose (1:1), the utilization ratio of xylose and glucose was 91.7% and 85.9%, with the ethanol yield was 26%. Xylose was eventually exhausted. Concentration of the by-product xylitol was very low.

Abstract:Chloramphenicol-resistant gene was cloned and analyzed by constructing genomic DNA library of Serratia marcescens KMR-3. It showed that cloned chloramphenicol-resistant gene encoded a protein product of 397 amino acids. The protein belonged to PRK10473 protein, and it showed 92% similarity to drug resistance transporter, Bcr/CflA subfamily of Serratia proteamaculans 568. Regulation elements including promoter, terminator, Shine–Dalgarno (SD) sequence and transcription start site also were identified.

Abstract:Compared with cultured fish, the fish oil of ocean wild fish contains much more Eicosapntemacnioc acid (EPA), Docosahexenoic acid (DHA), fat-soluble vitamin. To improve the utility value of oacean wild fish, small hairtail was used as raw material to investigate the technology of extracting fish oil with enzyme. The variables to affect the efficiency of extraction, extracting and centrifugation were selected as temperature, reaction time and pH value. Optimal technology conditions were determined by the response surface method: The liquid/solid ratio is 6, pH 7.3, enzyme amount of 1000 u/g raw material, agitation speed of 200 r/min, enzymolysis under 45oC for 90 min. The optimum extraction conditions were as follows: 100 mL extractant (every 20 g surimi), pH4.0, extracted under 40oC for 25 min. The optimal centrifuge conditions were: centrifuge speed of 3000 r/min (1865 g), centrifuged for 10 min. The oil extraction efficiency was 79.9%. This study developed the traditional technology of fish oil extraction, and improved the protection of the active components.

Abstract:The extracellular polysaccharides (EPS) of N. flagelliforme were purified by DEAE anion exchange chromatography and Sephadex G100 gel filtration chromatography. And two main components named NFPS1 and NFPS2 were obtained respectively. The physico-chemical characteristics of NFPS2 were analyzed and compared with NFPS0, which was obtained from field colony of N. flagelliforme. These results showed that both of NFPS2 and NFPS0 were composed of four monosaccharides: glucose, xylose, galactose and mannose. The apparent molecular weight of NFPS2 and NFPS0 was estimated to be 2.79×105, 2.26×105 respectively. They are non-sulfated polysaccharides, free of protein and nuclear acid. The thermal analysis indicated that there was a decomposition peak at 245oC in thermogravimetric (TG) curves. However, the microstructure analysis showed that they had different porous structures.

Abstract:Curdlan is a water insoluble exopolysaccharide produced by Alcaligenes faecalis under nitrogen-limiting conditions. After excretion, the polysaccharide is attached the cell wall. Thus enhancement of biomass production during the cell growth phase is important to curdlan production. A strategy of increasing nitrogen source to improve biomass production was adopted for curdlan production by Alcaligenes faecalis (ATCC 31749). In the batch fermentation of curdlan, a relatively higher NH4Cl level of 3.6 g/L with continuous glucose feeding increased the cell density leading to improvement of curdlan production. However, excessive NH4Cl would inhibit curdlan production and biomass production was not improved significantly. In addition, feeding of ammonia water at the initial phase replaced NaOH solution to control pH at 7.0. Subsequently, feeding of NaOH solution was resumed to control pH at 5.6 for curdlan production after ammonia was consumed. As a result, biomass production and curdlan yield were both enhanced remarkably. Feeding of ammonia water during the first 24 h led to biomass production of 18.8 g/L. However, higher cell density did not lead to increase in curdlan production. The maximum curdlan production (72 g/L) was obtained by feeding ammonia water for the first 14 h, during which the cell density was about 11.9 g/L.

Abstract:The extreme microorganisms Ferroplasma spp., play an important role in bioleaching of sulphide ores at low pH value and temperatures around 50oC. Without cell wall, Ferroplasma spp. is sensitive to pulp density, shearing force and heavy metal ions. Thus it is difficult to obtain their high cell density cultures, which limits the large-scale industrial application. In this paper, the optimum culture conditions of Ferroplasma thermophilum were studied by shaking culture. The results showed that the optimum culture conditions are as follows: 50oC, initial pH 0.5, 50 mL working volume in 250 mL shaking-flask, inorganic nitrogen source (NH4)2SO4. The optimum combination of FeSO4·7H2O, yeast extract and peptone was determined by orthogonal experiments, including FeSO4· 7H2O 40 g/L, yeast extract 0.3 g/L, peptone 0.2 g/L. Under the optimum culture conditions, the cell density was up to 6.3×107 cell/mL, and the oxidation of 40 g/L ferrous sulfate heptahydrate was finished in less than 72 hours. The results might provide information for scale-up of archaeon culture as well as its industrial application.

Abstract:Glutathione (GSH) plays an important role in the responses of microorganisms to the environmental stimulation and stress. The effect of H2O2 stress under different fermentation time and H2O2 concentration as well as continuous stress on GSH fermentation of Candida utilis were investigated in this paper. It was found that low concentration of H2O2 accelerated GSH production. When treated by low concentration of H2O2 (36 mmol/L), the final concentration of GSH reached 922 mg/L and the intracellular GSH content reached 1.64%, which increased by 7% and 35% than the controls, respectively.

Abstract:Actinobacillus succinogenes is a promising candidate for the production of bio-based succinic acid. Previously, we isolated a succinic acid-producing strain Actinobacillus succinogenes CGMCC 1593 from bovine rumen. In this paper, the influence of the environmental factors such as gas phase, pH, ORP, on succinic acid production by A. succinogenes CGMCC 1593 was studied. The results showed that CO2 was the optimum gas phase for anaerobic fermentation of A. succinogenes CGMCC 1593 as well as one of the substrate for the succinic acid synthesis. Using MgCO3 as a pH regulator, the pH was maintained within 7.1–6.2 during the anaerobic fermentation for the cell growth and acid production of A. succinogenes CGMCC 1593. Our results showed that low initial ORP was disadvantageous for the growth of A. succinogenes CGMCC 1593 and an ORP of ?270 mV was demonstrated to be beneficial to the succinic acid production. By adding Na2S?9H2O to decrease ORP to ?270 mV at the end of exponential growth phase in batch culture of A. succinogenes CGMCC 1593, the succinic acid concentration reached 37 g/L and the yield of succinic acid was 129% at 48 h. This work might provide valuable information for further optimization of succinic acid fermentation by A. succinogenes CGMCC 1593.

Abstract:China is the largest country of cultivating sweet potato in the world. The resources of sweet potato vines (SPV) are huge. To exploit the rich resources, SPV of XuShu 18 was selected as raw materials since it is cultivated in China widely. Polysaccharide from SPV was extracted by pilot instrument and precipitated by ethanol. The decolouring technology was studied. By primary purification, we obtained the polysaccharide from SPV (PSPV) and studied its physical and chemical characters. Using DEAE-cellulose column and NaCl solution as eluting reagent, three kinds of pure polysaccharide, named PSPVⅠ, PSPVⅡ and PSPVⅢ respectively, were obtained. Then the molecular weight distribution and sugar compositions were studied using High Performance Gel Filtration Chromatography and GC. The followings are the results. PSPVⅠ was separated from PSPV which was extracted from SPV harvested in the middle of June. Its M.W. is 6.278×104 D and is mainly composed of Xylose, Mannose and Glucose. PSPVⅡ and PSPVⅢ were separated from PSPV which was extracted from SPV harvested in the middle of October. Their M.W. are 3.801×104 D and 1.418×104 D respectively. PSPVⅡ is mainly composed of Mannose and Galactan. PSPVⅢ is mainly composed of Glucose, Xylose and Rhamnose. The results provide theoretical basis for the utilization of SPV.

Abstract:Our previous work has indicated that enzymatic resolution of glycidyl butyrate are strongly affected by many factors, including concentration of substrate, amount of lipase, the temperature, pH, shaking speed and reaction time. In this study, Plackett-Burman design was undertaken to evaluate the effects of the six factors. By regression analysis, concentration of substrate, amount of lipase and the temperature were found to be important for enzymatic resolution of glycidyl butyrate. In the second phase of the optimization process, a response surfacemethodology(RSM) was used to optimize the above critical factors, and to find out the optimal concentration levels and the relationships between these factors. By solving the quadratic regression model equation using appropriate statistic methods, the optimal parameter of the variables were determined as: 0.499 mol/L glycidyl butyrate, 30.23 mg/g lipase and 29.68oC. In the optimum condition, the value of enantiomeric excess(ee%) was 93.28%. Compared to 84.65% which was the maximum ee% under the non-optimized condition, this study has a significant advancement. The experimental data under various conditions have validated the theoretical values.

Abstract:In order to improve the yield of bacterial cellulose (BC), the fermentation medium of BC-producing strain J2 (Gluconobacter) was optimized, and BC ultra-micro-structure was observed. Initially, Plackett-Burman design was employed to evaluate eight variables which were relevant to BC production. Three statistically significant parameters including yeast extract, ZnSO4, ethanol were selected and other 5 variables were not significant (P>0.05). The optimized levels of three variables were defined by Box-Behnken design and response surface methodology (RSM). BC ultra-micro-structure was observed by scanning electron microscope (SEM) with cotton cellulose as comparison. The results indicated that the BC yield under the optimum fermentation medium was 11.52 g/100 mL, which was as 1.35 times as that under the original fermentation medium. The SEM photos manifested that bacterial cellulose ribbon, with a diameter less than 0.1 mm, was less than cotton cellulose ribbon. The bacteria inside the cellulose net were eliminated after the NaOH treatment.

Abstract:The production of propionic acid by Propionibacterium freudenreichii CCTCC M207015 was investigated in a Fibrous-bed bioreactor (FBB). The FBB was constructed by packing spiral cotton fibrous and immobilized into a bioreactor. By applying this bioreactor to propionic acid fermentation, the propionic acid yield had a significant improvement and reached 20.41 g/L, compared with the cell-free culture of 14.58 g/L (40 g/L of glucose). At the same time, the glucose exhausting time decreased from 120 h to 60 h. Batch fermentations at various glucose concentrations were carried out with FBB. Based on the analysis of the time course of production, fed-batch fermentation was also applied to produce propionic acid with FBB, the maximal propionic acid yield reached 45.91 g/L, and the proportion of propionic acid to total acids was about 72.31%.

Abstract:Biodenitrogenation of petroleum oil was investigated by a previously isolated carbazole-degrader Pseudomonas sp. XLDN4-9. In a tetradecane-aqueous phase system, biodegradation of carbazole was enhanced by the presence of n-tetradecane. And strain XLDN4-9 was capable of absorbing 95.2% of 2 g/L carbazole dissolved in diesel within 15 hours. Significant denitrogentation of crude oil, diesel and lubricanting oil was detected by strain XLDN4-9. Removal of carbazole, methylcarbazole, and dimethylcarbazole in diesel was confirmed by using GC-MS. After 3 days, 99% of carbazole and 15% of dimethyl carbazole was degraded. And the removal rate of 1-, 2-, 3-, and 4-methyl carbazole was determined to be 63.4%, 87.6%, 78.4%, and 66.5% respectively.

Abstract:Biohydrogen production from corncob by dark fermentation was reported for the first time. The effects of the pretreatment condition, substrate concentration and initial pH on the hydrogen production were investigated in batch cultivations. The maximum hydrogen yield of 107.9 mL/g-TVS and hydrogen production rate of 4.2 mL/g-TVS·h-1 were obtained under the condition of 1% HCl pretreating substrate for 30 min, 10 g/L substrate concentration and initial pH8.0. The content of hemicellulose in corncob decreased significantly from 42.2% to 3.0% after HCl pretreatment. The contents of cellulose, hemicellulose and lignin in the acid pretreated corncob decreased slightly in hydrogen producing process. The results indicate that the acid pretreatment of the substrate plays a key role in the conversion of corncob into biohydrogen. Fourier transform infrared spectroscopy (FTIR) was used to study the changes in the corncob composition during the treatment of chemical-microbial process. It was shown that the amorphous domains of cellulose and hemicellulose were hydrolyzed into fermentable asccharides through HCl pretreatment and the microorganisms had a devastating effect on the crystallinitiy of the cellulose.

Abstract:The inactivation efficiencies of microorganisms were found to be enhanced by using silver solution together with ultraviolet light (UV-A, 395 nm) irradiation. The inactivation efficiencies were improved remarkably especially in eukaryotic microorganism. To make clear the inactivation mechanism of microorganisms by the combination effect of silver and ultraviolet light irradiation, the resultant solution was characterized by ESR (Electron spin resonance, ESR). Scanning electron microscopy (SEM) and the method for measuring enzyme activity of mitochondria for eukaryotic cells were used to conjecture the mechanism, by analysis of the morphological and physiologic changes in eukaryotic cells. It is proposed that silver oxide (Ag2O) can be activated by ultraviolet light irradiation and react with water molecules to produce hydroxyl radical (?OH). Hydroxyl radical could damage cell wall of eukaryotic microorganisms, and inactivate the enzyme activity of mitochondria of eukaryotic microorganism cells. Accordingly, eukaryotic microorganism cells would die. In the experiment, Staphylococcus aureus was employed as the representative of prokaryotic microorganisms, and Candida albicans and Trichophyton mentagrophytes as the representative of eukaryotic microorganisms, respectively. Moreover, the results of the technology applied to washing machine were presented and discussed.

Abstract:Through studying the process of glycerol fermentation to 1, 3-propanediol(1, 3-PD) by Klebsiella pneumoniae, it was found that the cell growth and product (or by-product) production were under salt stress. Cell growth and product formation kept high rate at low salt concentration. High salt concentration led to low growth of cells, final concentration of 1, 3-PD and conversion from glycerol to 1, 3-PD, and, 1, 3-propanediol oxidoreductase activity decreased. When the salt concentration in 5 m3 bioreactor was controlled under appropriate manner, the concentration of 1, 3-PD production was markedly enhanced. The final 1, 3-PD concentration ,the conversion of glycerol to 1, 3-PD and productivity were 64 g/L, 61% and 2.1 g/(L·h).

Abstract:A strain ZG0656 producing α-amylase inhibitor was isolated from soil in this study. Polyphasic taxonomic studies were performed, including appearance characteristics, culture characteristics, phenotypic characteristics, cell walls chemical composition, nearly complete 16S rDNA sequence alignment with those of representative Streptomyces species. These results revealed that strain ZG0656 represents a novel variation of Streptomyces coelicoflavus, for which we propose the name S. coelicoflavus var. nankaiensis. After fermentation in a 10 L fermentor, a-amylase inhibitors were accumulated in the harvested broth of strain ZG0656. The a-amylase inhibitors we obtained were identified as aminooligosaccharides after concentration, resin-adsorption, gel-filtration, and desiccation. They could intensively inhibit α-amylase, depress postprandial blood glucose elevation obviously. Thus, the α-amylase inhibitors are expected to act as drugs or functional food against diabetes.

Abstract:The xylose reductase of Pichia stipitis is one of the most important enzymes. It can be used to build up recombinant Saccharomyces cerevisiae strain for utilizing xylose and producing ethanol. Intercellular redox imbalance caused by NADPH preference over NADH for Pichia stipitis xylose reductase (PsXR) has been considered to be one of the main factors for poor ethanol productivity. Some key amino acids of PsXR, which affect the activity or coenzyme preference, were investigated in our previous study. In this study, Lys21 were rational designed for site-directed mutagenesis to alter coenzyme specificity of PsXR from NADPH and NADH into single NADH. The wild gene and mutagenesis genes were ligated into pET28b, and were transferred into E.coli BL21(DE3). After induced by IPTG, the xylose reductases were purified. Purified mutants K21A (Lys21→Ala), K21R(Lys21→Arg) were characterized by steady-state kinetic analysis. The results showed that the coenzyme dependence of K21A was completely reversed to NADH.

Abstract:Cellulases are relatively costly enzymes that are sold in large volumes for use in different industrial applications, and a significant reduction in cost will be important for their commercial use in biorefineries. The production of cellulase is a major factor in the hydrolysis of cellulosic materials. Hence it is essential to make the process economically viable. A strain (L-06) with high cellulase activity was screened from rice straw compost and classified as Penicillium decumbens by the analysis of its morphology and 18S rRNA gene sequences. Different conditions of liquid fermentation medium including nitrogen source, carbon source, surfactant, temperature, initial pH, inoculation quantity for the production of cellulase had been studied. The maximal b-1, 4-glucosidase(BGL) activity was 1662 u/mL which is 1.49 times of the previous and the maximal exo- b-1, 4-glucanases(CBH) activity was 2770 u/mL which is 1.36 times of the previous, cultured in the optimal condition for three days. And the maximal endo-b-1, 4-glucanases (EG) activity was 18064 u/mL which is 1.87 times of the previous and the maximal filter paper enzyme(FPase) activity was 4035 u/mL which is 1.47 times of the previous , cultured in the optimal condition for four days. In the optimization experiments, the EG and CBH in the culture condition (pH10) maintained 70% and 43% activity. In the culture condition(50oC) EG and CBH maintained 59% and 68% activity, which showed heat and alkali resistant characteristics.

Abstract:Anoxybacillus sp. WP06 is a thermophilic (optimum temperature for growth, 60oC), facultative anaerobe. Strain WP06 is able to utilize a wide range of carbon sources such as glucose, xylose, arabinose, starch, maltose and sorbitol. Anaerobically, glucose and xylose were fermented to ethanol as minor products. Unlike most thermophilic bacteria isolated to date, strain WP06 is tolerant (maintained viability) to high ethanol concentrations up to 15% at 60oC. The growth rate was slightly inhibited at 8% ethanol. The observation that strain WP06 exhibits higher tolerance of 15% ethanol at 60oC exploits the level of ethanol tolerance in thermophilic bacteria. Strain WP06 may be candidate for mechanisms of ethanol tolerance in thermophilic bacteria.