Objective To establish a RP-HPLC method for determination of ammonium glycyrrhizinate in Shufei Mixture .Methods The sample was analyzed on an Agilent Zorbax SB-C18 column (4.6mm ×150 mm, 5μm), and acetonitrile-0.0025 mol/L sodium heptanesulfonate solution–0.05 mol/L potassium dihydrogen phosphate solution(20:45:45) ( adjusted pH value of 7.2 ±0.05 with 20% sodium hydroxide solution) was used as mobile phase.The flow rate was at 1.0 mL/min.The detective wavelength was at 250 nm.The column temperature was 30 ℃.Results With this chromatographic condition, the ammonium glycyrrhizinate peak in Shufei Mixture sample chromatogram could be separated with other ingredient peaks completely.The negative sample had no interference.The calibration curve was linear at a ranges of 23.6-118.1μg/mL for ammonium glycyrrhizinate, and equation of regression was Y=0.1133X–0.00110,r=0.999 8.The average recovery from sample was 97.8% and RSD was 0.88%(n=6).The content range of ammonium glycyrrhizinate in three batch Shufei Mixture sample was 0.2532-0.2865 mg/mL, and average content was 0.2721 mg/mL. Conclusion This method is simple, accurate, and useful for control method of this preparation.

Objective To summarize the clinical characteristics,diagnosis and treatment of autoimmune pancreatitis(AIP).Methods The retrospective descriptive study was adopted.The clinical data of 43 patients with AIP who were admitted to the First Affiliated Hospital of China Medical University between January 2010 and December 2014 were collected.All patients underwent laboratory tests and imaging examinations.For the patients who had the definite diagnosis of AIP,corticosteroid therapy was adopted,and for the patients who had the suspicion of pancreatic cancer,surgical operation was carried out.Observed indexes included clinical manifestations,laboratory findings,imaging findings,treatment,pathological findings and follow-up status.The follow-up of imaging examinations and laboratory tests via outpatient and inpatient examinations was performed on the patients until January 2016.Measurement data with skewed distribution were presented as average (range).Results (1) Clinical manifestations:of the 43 patients,33 demonstrated obstructive jaundice,19 had loss of weight,18 had upper abdominal pain and 5 had acute mild pancreatitis (the same patient with a variety of symptoms).Extrapancreatic organ involvement was detected in 23 of the 43 patients,including focal nephritis in 15 patients,sicca syndrome in 7 patients,pulmonary involvement in 5 patients,autoimmune thyroiditis in 5patients,hilar IgG4-associated cholangitis in 3 patients,retroperitoneal fibrosis in 3 patients and ulcerative colitis in 1 patient (the same patient with a variety of diseases).(2) Results of laboratory tests:17 patients underwent serum IgG4 detection and increased expression of IgG4 was found in 15 patients.(3) Imaging findings:all the 43patients underwent enhanced computed tomography (CT) scan of pancreas.Diffused enlargement of pancreas with appearance of sausage was observed in 34 patients and localized pancreatic head enlargement was observed in 9patients.Twenty patients had peripancreatic vascular involvement including stenosis or occlusion of splenic vein,stricture at the confluence of superior mesenteric vein,portal vein and splenic vein resulting in regional portal hypertension which led to gastric venous varices and splenomegaly.Three patients showed that superior mesenteric artery was surrounded by tissue masses with massive ascites.Five patients had pancreatic pseudocyst with splenic vein involvement and peripancreatic effusion.The abdominal CT examination of 15 patients with renal involvement showed renal cortical wedge or nodular lesions.Pulmonary inflammatory pseudotumor and interstitial pneumonia appeared in 5 patients,CT of whom showed solid pulmonary nodules or diffuse ground-glass opacities and patchy shadows in pulmonary interstitium.Thirty-two patients underwent magnetic resonance cholangiopancreatography and 28 patients underwent endoscopic retrograde cholangiopancreatography.The results showed the focal and diffuse or segmental stenosis of the main pancreatic duct were detected in 28 patients (diffuse stenosis in 23patients,local stenosis in 5 patients),stenosis of the intrapancreatic common bile duct in 32 patients,stenosis of hilar bile duct in 3 patients.Nine patients underwent endoscopic ultrasonography examination.The results showed 3 patients had hypoechoic mass in the head of pancreas and 6 patients had diffuse or heterogeneous hypoechoic of the pancreatic parenchyma.(4) Treatment status:of 43 patients,30 patients received initial steroid treatment.After steroid treatment,the clinical symptoms of 27 patients gradually improved and laboratory and imaging findings were both improved.Eleven patients didn 't receive initial steroid treatment,of whom 2 patients with obstructive jaundice underwent endoscopic biliary stent implantation,7 patients underwent surgical biliary drainage and 2 received neither endoscopic treatment nor surgical treatment.Two patients underwent pancreatoduodenectomy due to suspicion of pancreatic adenocarcinoma and were diagnosed as AIP based on postoperative pathological results and treated with steroid after operation.(5) Pathological examination:of the 43 patients,22 underwent pathological examination.Results of pathological examination of superficial lymph nodes showed lymph follicles hyperplasia,and immunohistochemistry staining showed massive IgG4 positive plasmocytes in 5 patients,supporting the diagnosis.Results of fine needle aspiration cytology in 15 patients showed no cytological evidence were detected in 14 patients and epithelial cells of malignant tumors in 1 patient.Results of pathological examination of surgical specimens showed dense infiltration of lymphocyte and immunohistochemistry staining showed massive IgG4 positive plasmocytes.(6) Follow-up status:of 43 patients,41 were followed up for an average time of 28 months (range,12-71 months).Of the 30 patients who received steroid treatment,27 had complete response and 3 had partial response.Of the 27 with complete response,4 patients relapsed during followup and were cured by continuous steroid treatment and extending treatment course,3 patients did not relapse,1 patient relapsed again,and long-term low dose steroid maintenance therapy was needed.The 3 patients with partial response were treated with mycophenolate mofetil for 6,4,4 months and imaging abmormalities disappeaved absolutely.Eleven patients received no steroid for the initial treatment,in whom 2 patients were lost to follow-up,1 patient with common bile duct jejunum anastomosis died in 1 year after surgery.In the other 8 patients,the imaging abnormalities were spontaneous remission or clinical and imaging manifestations were improved after steroid treatment and no recurrence was found.Two patients who underwent pancreatoduodenectomy due to suspicion of pancreatic adenocarcinoma received steroid treatment after operation and no recurrence was observed.Conclusion Clinicians should combine clinical manifestations,serological examination,imaging findings and pathological examination to make definite diagnosis of AIP and make differential diagnosis of pancreatic cancer to avoid unnecessary surgical treatment by misdiagnosis.

Objective:To purify H5N1 influenza virus concentrate prepared by MDCK cells with a new mixed-mode chromatography medium Capto Core700 and the traditional medium Sepharose 4FF, and to compare the separation and purification efficacy of the two media.Methods:Capto Core700 and Sepharose 4FF were used to purify inactivated H5N1 influenza virus concentrate. The morphology of virus particles in different samples was then observed under a transmission electron microscope. Single radial immunodiffusion (SRID), Folin-Phenol (Lowry) method, double-antibody sandwich ELISA and qPCR were used to detect hemagglutinin, total protein, host cell protein (HCP) and host cell DNA (HCD) before and after purification. The recovery rate of virus antigen and the removal rate of impurities were calculated. The immunogenicity of the viruses purified with different media was analyzed using animal experiments. Difference in the purification efficacy of the two chromatography media was analyzed by t-test. Results:H5N1 influenza viruses purified by Capto Core700 or Sepharose 4FF showed the typical influenza virus morphology under transmission electron microscope. There was no significant difference in the recovery rate of hemagglutinin between the two chromatography media ( P>0.05), but compared with Sepharose 4FF, Capto Core700 had a higher removal rate of impurities (total protein, HCP, HCD) and the difference was statistically significant ( P<0.05). Animal experiments showed that the viruses purified by the two chromatography media had good immunogenicity. Conclusions:Compared with Sepharose 4FF chromatography medium, Capto Core700 could more effectively remove process-related impurities such as HCP, HCD and total protein without affecting the recovery rate of viral antigen. This study provided reference for the development of purification technology in the production of H5N1 influenza virus vaccine in MDCK cells.