Pullulan is a linear glucosic polysaccharide produced by the polymorphic fungus Aureobasidium Pullulans, which has long been applied for various applications in medical and food industry due to its security, stability and low adhesive ability.At present, the two problems in restricting pullulan industrial production are the low polysaccharide production and melanin secreted which is hard to erase completely, giving the following process some problem.As a starting point, this review article collects and analyzes the progress on the breeding of pullulan high-yield strain without melanin in recent years, in order to find more efficient strains breeding methods, laying a foundation for further breeding of pullulan high-yield strain without melanin.

Objective To investigate the role of interleukin-1 beta (IL-1?) in glomerulosclerosis secondary to adriamycin (ADR)-induced nephropathy in rats, and the effect of simvastatin on the expression of IL-1?. Methods Male Sprague-Dawley rats were randomly divided into normal control, ADR-induced nephropathy (model), and simvastatin-treated ADR nephropathy (treatment) groups. After ADR-induced nephropathy establishment in model and treatment groups, and eleven weeks of intragastrical administer of normal saline in normal control and model groups and of simvastatin in treatment group, the expression of IL-1? was detected by immunohistochemistry, RT-PCR and ELISA. Histopathological change of renal tissues was observed under light microscope, and glomerulosclerosis index (GSI) was also evaluated. Results Higher expression of IL-1? in kidney and GSI, as well as more severe loss of renal function were observed in model group than those in control group (all P

Objective To construct a eukaryotic expression vector in Pichia pastoris containing human tissue factor( hTF) gene,in order to achieve high level secretory expression in extracellular.Methods Expression plasmid, pGAPZaA-hTF, was constructed by inserting the synthesized sequence encoding human extracellular tissue factor into yeast expression vector pGAPZaA and transformed into Pichia pastoris SMD1168H with electroporation.Having been selected by Zeocin, transformants containing hTF cDNA were expressed in YPD, extracellular proteins were detected by SDS-PAGE and confirmed by Western bolt.Results Successfully constructed the recombinant pGAPZaA-hTF expression system in Pichia pastoris.SDS-PAGE showed that the molecular weight of the expression product was about 37 -40 kDa.Western-blot indicated that it was human extracellular tissue factor.The crude yield of total protein in medium was up to 1 g/L, more than 80% of which was hTF.Conclusion Truncated rTF gene is expressed in Pichia pastoris and the active products are secreted into the medium which have the same activation as the native TF.

We enrolled 60 patients with American Association of Anesthesiologists grade Ⅰ - Ⅱ undergoing unilateral total knee arthroplasty. All patients received combined epidural and spinal anesthesia,and a nerve stimulator was used to guide placement of a femoral nerve catheter. Patients were divided into three groups according to the catheter location on X-ray : psoas muscle group ( n = 18 ), iliacus muscle group (n = 19) and local group (n =23). Visual analog scale (VAS) pain scores were recorded at rest and with movement at 4, 24 and 48 h postoperatively and sensory blockade of the femoral, obturator and lateral femoral cutaneous nerves was recorded at 24 h.There were no significant differences in femoral nerve blockade among the three groups. Obturator nerve blockade was significantly better in the psoas muscle group than in the iliacus muscle and local groups, and was also better in the local group than in the iliacus muscle group. There was no significant difference in lateral femoral cutaneous nerve blockade between the psoas muscle and iliacus muscle groups, but there was better blockade in both these groups than in the local group. At 4 h postoperatively, VAS pain scores at rest were significantly lower in the psoas muscle group than in the iliacus muscle and local groups, but there were no significant differences in VAS pain scores with movement among the three groups. At 24 and 48 h postoperatively, VAS scores at rest and with movement were significantly lower in the psoas muscle group than in the iliacus muscle and local groups.

OBJECTIVE:To optimize the extraction technology of glucosinolates from Uighur medicine Brassica rapa L.. METHODS:Ethanol refluxing method was adopted to extract glucosinolates from Uighur medicine B. rapa L.. With the comprehen-sive score of glucosinolates and dry extract yield as the index,L9(34)orthogonal test based on the single factor test was adopted to observe the effects of ethanol volume fraction,extraction time and material-liquid ratio on the extraction technology of glucosino-lates from B. rapa L.,and verification test was conducted. RESULTS:The optimal extraction technology was to add 95% ethanol 8 times as much as the amount of the herbs,twice for 1.0 h reflux extractions. The extraction amount of glucosinolates and dry ex-tract yield were 7.36 mg/g and 25.29% at average,respectively.The comprehensive score of RSD was 0.52%(n=3). CONCLU-SIONS:The optimal extraction technology is stable and feasible and can be used for the extraction of glucosinolates from B. rapa L..

Objective To retrospectively study the high risk factors for biliary complication (BC) and the application of the Clavien system to classify BC in a large cohorts of subjects undergoing liver transplantations (LT).Methods The clinical data of 181 patients who received LT from Jan.2004 to Dec.2008 were studied.BC was classified using the Clavien system.The risk factors of biliary complication were evaluated by using a binary forward stepwise logistic regression analysis.Results 14.4％ (26/181) recipients developed BC (BC group).In 84.6％ (22/26) patients the BC was above the Clavien Ⅲ b.Regression analysis of BC revealed that the placement of a T tube (P =0.0090,OR=31.177),RIld (P=0.0094,OR＜0.001).RI1w (P=0.0013,OR＞999.999) were significantly associated with the development of BC.Regression analysis of BC above Clavien Ⅲ b revealed that RIld (P=0.0065,OR＜0.001,RI1w (P=0.0022,OR＞999.999) were significantly associated with the development of BC above Clavien Ⅲ b.Conclusions The Clavien classification system was useful to classify BC.The placement of a T tube was an independent risk factor to predict BC,it was not a factor for BC above Clavien Ⅲ b.Hepatic arterial insufficiency (HAI) was an independent risk factor for BC and BC above Clavien Ⅲ b.

Objective To construct a eukaryotic expression vector in Pichia pastoris containing human fibrinogen gene, in order to achieve high level secretory expression in extracellular.Methods Expression plasmid,pGAPZαA-FGB-FGG-FGA-AOX1,was constructed by inserting the synthesized sequence encoding human fibrinogen(FGA, FGB,FGG) and then introduced into Pichia pastoris SMD1168H by electroporation.Transformants were availably screened by Zeocin resistance,the expression of recombinant protein was identified by SDS-PAGE and Western blot analysis, the protein yield was tested by ELISA assay.After ultrafiltration and purification, the biological activity of protein was detected.Results The crude yield of human fibrinogen in Pichia pastoris supernatant reached 15 mg/L in flask and the biological aggregation activity was determined.Conclusion The human fibrinogen gene was obtained and successfully expressed in Pichia pastoris and the active products were secreted into the medium.

Objective To establish a method of HPLC assay for determining stilbene glucoside in Zishen Ningshen Pills(ZNP),and to study the influence factors on the content of stilbene glucoside in the process of preparation.Methods HPLC was used for the determination of stilbene glucoside in ZNP.Through simulation the process of preparation,the stilbene glucoside content in the intermediate products was determined by HPLC,and its retention rate and metastasis rate were also investigated.Results The resolution and the linearity of stilbene glucoside were fine,the average recoveries being 98 % ～ 102 %.The retention rate of stilbene glucoside in the drying powder was 60.3 %,lower than that in the original medicinal powder.Conclusion The quantitative method for determining the ingredients in ZNP is simple,feasible and reproducible,and is beneficial for quality control of ZNP.The drying process under normal pressure is the main influence factors of the decrease of stilbene glucoside content,and the decompression drying can be taken into account to take the place of the atmospheric drying.

Objective To construct a prokaryotic expression vector in BL21 to secretorily expressα-Cyclodextrin Glycosyltransferase(α-CGTase). Methods α-CGT gene was amplified from Bacillus macerens genome by PCR.pET26b and α-CGT gene were connected after digested with Nco I, Xho I respectivly, and then transformed into Escherichia coli BL21 strain.α-CGTase was expressed in fermentation culture medium and AA-2G was prepared by using α-CGTase, VC and starch.Results α-CGTase was expressed secretorily and the enzyme activity was up to 120 U/mL.AA-2G was prepared by the biotransformation of VC and starch using α-CGTase which proved to be correct by HPLC.Conclusion AA-2G was prepared by using self-madeα-CGTase, after optimized the preparation conditions the yield of AA-2G was 17.46 g/L, and the conversion rate reached 58.2%(mg/mg).

Objective To investigate the effect and possible mechanism of bone marrow stem cell mobilized by stem cell factor (SCF) with granulocyte colony-stimulating factor(G-CSF)on renal peritubular capillary, fibrosis and renal function in unilateral ureteral obstruction (UUO) rats. Methods One hundred and twenty eight healthy male Wistar rats were randomly divided into four groups: Sham group, SCF-G group, UUO group and UUO+SCF-G group. Eight rats of each group were randomly selected and killed on the 5th, 14th, 21st and 28th day. Serum creatinine, CD34 positive cells and factor Ⅷ positive cells in renal interstitium, histopathologic lesion scores of interstitial fibrosis and interstitial pathology in kidney were measured. The mRNA expression of vascular endothelial growth factor (VEGF). and thrombospondin-1 (TSP-1) in the renal cortex was detected. Results (1) The renal interstitial fibrosis anti the loss of peritubular capillary were observed in UUO group after two weeks. (2) The number of bone marrow stem cells homing to renal interstitium in UUO +SCF-G group was significantly higher than that in UUO and Sham groups (P<0.05). (3) The loss of peritubular capillary in UUO+SCF-G group appeared later than that in UUO group (P<0.05). (4) The interstitial fibrosis and tubule injury was milder in UUO+SCF-G group than that in UUO group (P<0.05). (5) The decrease of VEGF mRNA expression of renal cortex in UUO +SCF-G group was seen later than that in UUO group. VEGF mRNA expression in UUO+SCF-G group was higher than that in UUO group. (6) The increase of TSP-1 mRNA expression of renal cortex in UUO+SCF-G group was seen later than that in UUO group. TSP-1 mRNA expression in UUO+SCF-G group was lower than that in UUO group (P<0.05). (7) In UUO and UUO+SCF-G groups, peritubular capillary index was negatively correlated with serum creatinine, interstitial fibrosis and interstitial lesion scores. VEGF mRNA expression of renal cortex was positively correlated with peritubular capillary index, and TSP-1 mRNA expression of renal cortex was positively correlated with peritubular capillary index. Conclusions (1)The loss of peritubular capillary is found in UUO group, and is correlated with interstitial fibrosis and interstitial lesion. (2) Application of SCF with G-CSF can effectively motivate stem cells to injured renal tissue, contribute to decrease the loss of peritubular capillary, lessen interstitial fibrosis and interstitial lesion, and ameliorate renal function. (3) Application of SCF with G-CSF can up-regulate VEGF mRNA expression and down-regulate TSP-1 mRNA expression, which may contribute to promote the repair of endothelial cells and protect peritubular capillary.