An important index determination for clinical diagnosis of renal function is to assay the creatinine concentration in serum. In the analytical process applied with coupled-enzyme, the quality control of sarcosine oxidase (SOX) as a key enzyme is the first problem to be solved. In order to establish an efficient and laboratory-scale production of SOX, the recombinant sarcosine oxidase (r-SOX) gene was a high-level expression in E. coli induced with lactose on a large-scale fermentation in 300 L fermenter. The results suggested that the biomass concentration reached OD600 of 22 and the expression of recombinant sarcosine oxidase in E. coli accounted for about 25% of total soluble protein in culture after fermentation. The cell-free extract obtained from high pressure homogenizer was processed by selective thermal denaturation and then purified with Ni-Sepharose FF chromatography. The sarcosine oxidase with 97% purity, 25 U/mg specific activity and 92.4% activity recovery was obtained. The molecular weight with single peptide chain of 53 kD and 55 kD of recombinant sarcosine oxidase was assessed by SDS-PAGE in presence or absence of 2-mercaptoehanol and Sephacryl S-200 chromatography. This sarcosine oxidase was found to be a conjugated protein, yellow enzyme, which combined with FAD as prosthetic group by covalent linkage. The contaminant of catalase was not detected in the sample pool of this enzyme. In addition, a further test to the thermal stability of sarcosine oxidase was done. According to the above results, the development and utilization of this enzyme has been set up on a reliable foundation.
Escherichia coli ; Fermentation ; Recombinant Proteins ; biosynthesis ; Sarcosine Oxidase ; biosynthesis

Objective To evaluate the safely and efficacy of therapeutic endoscopic retrograde cholangiopancreatography (ERCP) and its related procedures in patients older than 85 years.Methods Data of patients older than 85 who underwent ERCP between 2004 and 2009 (group A, n =52) were reviewed and compared with those of patients aging 70-84 years (group B, n = 329).Results The rate of patients with underlying disorders of other systems in group A was significantly greater than that of group B (100.0% vs.77.5% , P < 0.05) , as well as the rate of multiple procedures to achieve stone clearance (42.3% vs.28.8% , P <0.05).There were no significant differences between 2 groups regarding the rates of complication (7.7% vs.4.9% ) and post-procedure mortality (1.9% vs.0.6% ).Conclusion Therapeutic ERCP and related procedures are safe and effective in patients aging over 85, and advanced age should not be regarded as the contraindication of the procedure.

Objective To investigate the diagnostic value of intraductal ultrasonography for the quality of biliary stricture.Methods Data of the patients who had received operation because of biliary stricture after IDUS examination from 2006 to 2010 were collected.IDUS results were compared with those of operation.Results There were 43 cases of malignant strictures and 6 benign strictures in total.The sensitivity,specificity,positive predictive value,negative predictive value and diagnostic accuracy of intraductal uhrasonography for the quality of biliary stricture were 97.7％ ( 42/43 ),83.3％ ( 5/6 ),97.7％ ( 42/43 ),83.3％ (5/6) and 95.9％ (47/49),respectively,which were significantly higher than conventional imaging like ultrasound B,CT and MRCP.Twenty one cases in 32 were diagnosed as malignant biliary stricture with cytological brushing,with the diagnostic accuracy of 65.6％.All cases had been diagnosed by IDUS.Conclusion Intraductal ultrasonography is of high diagnostic value for biliary stricture.However,cytological brushing based on IDUS is of limited diagnostic value for malignant biliary stricture.

ObjectiveTo investigate the diagnostic value of single balloon enteroscopy (SBE) for obscure gastrointestinal bleeding.MethodsA total of 78 SBE procedures was conducted on 72 patients with obscure gastrointestinal bleeding,with 40 via oral route and 38 via anal route.The procedure time,insertion depth and rate of positive finding were recorded.ResultsFor 40 SBE procedures performed via oral route,the mean procedure time was 60 minutes ( 15-110 minutes),and the mean insertion depth was 195 cm at the distal end of Trentz ligament (30-240 cm).For 38 SBE procedures performed via anus,the mean procedure time was 75 minuets (30-120 minutes),and the mean insertion depth was 160 cm at the proximal end of ileocecal valve (50-200 cm ).The whole diagnostic yield of obscure gastrointestinal bleeding was 62.5％.ConclusionSBE is a safe and useful tool for the diagnosis of obscure gastrointestinal bleeding.

Objective To investigate the prognostic value of preoperative neutrophil lymphocyte ratio (NLR) in patients with gastrointestinal stromal tumors (GIST).Methods The clinical-pathological data from 85 GIST cases were collected from the Second Affiliated Hospital of Wenzhou Medical University and retrospectively analyzed from June 2009 to December 2013.According to the exclusion criteria,79 cases were enrolled.Preoperative neutrophil and lymphocyte count was collected and NLR was calculated.According to the receiver operating characteristic (ROC) curve of NLR,GIST patients were divided into low NLR group (NLR ＜ 2.30) and high NLR group (NLR ≥ 2.30).Clinic-pathological features and five year disease free survival (DFS) were compared between the two groups.Results There was a statistical significant difference in tumor size and tumor risk between high NLR group and low NLR group (respectively,x2 =9.517,12.411,all P ＜ 0.05).Univariate analysis showed that the five year disease free survival rate of low NLR and high NLR group were 78％ and 32％ (x2 =18.749,P =0.000).By multivariate analysis a high NLR was identified as an independent risk factor of poorer prognosis for patients with GISTs (RR:3.516,95％CI:1.453-8.506,P=0.005).Conclusion A high preoperative NLR is an independent risk factor for the prognosis of GISTs.

Abscisic acid, a plant hormone that inhibits growth, is a key factor in balancing plant endogenous hormones and regulating growth and metabolism. Abscisic acid can improve the drought resistance and salt tolerance of crops, reduce fruit browning, reduce the incidence rate of malaria and stimulate insulin secretion, so it has a broad application potential in agriculture and medicine. Compared with traditional plant extraction and chemical synthesis, abscisic acid synthesis by microorganisms is an economic and sustainable route. At present, a lot of progress has been made in the synthesis of abscisic acid by natural microorganisms such as Botrytis cinerea and Cercospora rosea, while the research on the synthesis of abscisic acid by engineered microorganisms is rarely reported. Saccharomyces cerevisiae, Yarrowia lipolytica and Escherichia coli are common hosts for heterologous synthesis of natural products due to their advantages of clear genetic background, easy operation and friendliness for industrial production. Therefore, the heterologous synthesis of abscisic acid by microorganisms is a more promising production method. The author reviews the research on the heterologous synthesis of abscisic acid by microorganisms from five aspects: selection of chassis cells, screening and expression enhancement of key enzymes, regulation of cofactors, enhancement of precursor supply and promotion of abscisic acid efflux. Finally, the future development direction of this field is prospected.
Abscisic Acid/metabolism* ; Plant Growth Regulators/metabolism* ; Plants/metabolism* ; Yarrowia/metabolism*