To establish a detection method of oncolytic adenovirus/p53 and standard of quality control, human telomerase reverse transcriptase (hTERT) promoter, CMV fusion promoter containing hypoxia reaction element (HRE) and p53 gene were identified by vector DNA restriction enzyme digestion and PCR analysis. The result conformed that all modified regions were in consistent with theoretical ones. Particle number was 2.0 x 10(11) mL(-1) determined by UV (A260). Infectious titer was 5.0 x 10(10) IU mL(-1) analyzed by TCID50. In vitro p53 gene expression in human lung cancer cell H1299 was determined by ELISA, and A450 ratio of nucleoprotein in virus infection group to control group was 5.2. Antitumor potency was evaluated by cytotoxicity assay using human lung cancer cell A549, and the MOI(IC50) of this gene therapy preparation was 1.0. The tumor cells targeted replication ability of recombinant virus was determined by TCID50 titer ratio of filial generation virus between human lung cancer cell A549 and human diploid epidermal fibrolast BJ cells after infected by virus with same MOI. TCID50 titer ratio of tumor cell infection group to normal cell infection control group was 398. The IE-HPLC purity of virus was 99.5%. There was less than 1 copy of wild type adenovirus within 1 x 10(7) VP recombinant virus. Other quality control items were complied with corresponding requirements in the guidance for human somatic cell therapy and gene therapy and Chinese pharmacopeia volume III. The detection method of oncolytic adenovirus/p53 was successfully established for quality control standard. The study also provided reference for quality control of other oncolytic viral vector products.

Objective To evaluate the quality status of recombinant human interferon α1b injection and find out some quality problems.Methods Totally 31 batches of recombinant human interferon α1b for injection and 11 batches of recombinant human interferon α1b injection from two enterprises were examined according to Chinese Pharmacopoeia Volume Ⅲ (2010),and the quality status of recombinant human interferon α1b injection was evaluated by statistical analysis of the results.Results All 42 batches of samples were qualified.The production process of each enterprise was steady.Conclusion At present the quality of recombinant human interferon αlb injection is generally good.The current standards are feasible,but the specified standard of osmolality needs to be improved.

An approach was proposed to evaluate preparation technology by means of fingerprint-peak matching technology of high performance liquid chromatography with diode array detector (HPLC-DAD).Similarity and hierarchical clustering analysis (HCA) were applied to identify the 15 batches of Xiaochaihu granules from different manufacturers and our laboratory,and peak pattern matching between the composite formulae and Radix Bupleuri Chinensis,which was one of the main ingredients of Xiaochaihu granules,was utilized to evaluate the preparation technology of Xiaochaihu granules via the indexes of the relative deviation of retention time (RT) and UV spectrum feature similarity of their corresponding peaks.Eleven matching peaks were found between Xiaochalhu granules and Radix Bupleuri Chinensis.However,the saikosaponin A and saikosaponin D,which are the important active components in Radix Bupleuri Chinensis,were not found in Xiaochaihu granules from any manufacturers.The peak areas of 11 characteristic peaks of Xiaochaihu granules samples formed a matrix of 11 × 15.The result of HCA showed that Xiaochaihu granules samples were divided into four kinds of category.Xiaochaihu granules samples from the same manufacturer were basically clustered of the same category.The results suggested that the saikosaponin A and saikosaponin D are prone to structural transformation under the condition of decoction and in the presence of the organic acidic components.These active components,existing in raw herb,might transform to a series of non-active secondary saikosaponin due to unfavourable preparation technology.So the conventional decoction-based preparation technology of Xiaochaihu granules might greatly affect its quality and therapeutic effectiveness. This study demonstrates that fingerprint-peak matching technology can not only be used for quality control of this composite formulae,but also provide some guidance for preparation technology of Xiaochaihu granules.

OBJECTIVETo develop a HPLC method for determination of matrine, oxymatrine; ferulic acid, L-shikonin and beta, beta-dimethylacrylshikonin in compound preparatioti of Dangguishuan.

METHODThe chromatogtaphic separation was performed on a Kromasil ODS C18 column (4.6 mm x 250 mm, 5 microm) maintaining at 30 degrees C during the whole process. The mobile phase consisted of methanol and 0.1% triethylamine aqueous solution (adjusted with phosphoric acid, pH 3) at a flow rate of 1.0 mL x min(-1). The detection wavelength was set at 220 nm for matririne and oxymatrine, 316 nm for ferulic acid, 516 nm for L-shikonin and beta, beta-dimethylacrylshikonin, respectively.

RESULTAll the compounds showed good linearity (r > 0.9996) in the range of the test concentrations, and the average recoveries of the method is betwuen 96.92% and 102.22%, RSD < 3.1%.

CONCLUSIONThe method is proved to be credible, sensitive, accurate and repeatable. It can be applied to determine of matrine, oxymatrine, ferulic acid, L-shikonin and beta, beta-dimethylacrylshikonin in compound preparation of Dangguishuan simultaneously, and provide a basal method of quality control to this preparation and other relative preparations.

Alkaloids ; analysis ; Chromatography, High Pressure Liquid ; methods ; Coumaric Acids ; analysis ; Drugs, Chinese Herbal ; analysis ; Naphthoquinones ; analysis ; Quinolizines ; analysis

Objectives: To establish an effective method to isolate germinal vesicle frommammalian oocyte and make the isolated germinal vesicle undergo meiotic resumptionin vitro. Methods: The germinal vesicles were isolated from oocytes directly througha physical method by drawing oocyte in and out of fine-bore pipette.The isolatedgerminal vesicles were cultured in cell-free system which was prepared from the celllysate of synchronized HeLa cells.DNA fluorescence dye Hoechest 33342 was used tomonitor the condensation of chromatin during germinal vesicle breakdown. Results:The cell lysate of metaphase HeLa cell induced chromatin condensation while the cellextract of early G2 phase could not. Conclusions: The germinal vesicle can beisolated successfully in such a physical method and it can undergo chromatincondensation in vitro just as the oocyte does in vivo. Natl J Androl,2001,7(2):79～83

Objective To discuss the diagnosis and treatment of normotensive pheochromocyto-ma. Methods The clinical data of 22 patients with normotensive pheochromocytoma were reviewed. Inclusion criteria for normotensive pheochromocytoma were no previous history of hypertension and episode of symptoms suggesting high blood pressure. The blood pressure on admission was 90-130/ 60-90 mm Hg with an average of 113/72 mm Hg. Seven patients were found adrenal mass by routine ultrasonic examination. Twelve patients presented with superior abdominal or flank pain. Four pa-tients were present with fatigue, and 2 patients had fever. Headache and palpitation were found in 1 patient. Most of patients were present with large and round mass with low density area in the center of the tumor by uhrosonography and CT. Four patients had elevated level of plasma epinephrine and nor-epinephrine. 24 hours urine CA and VMA were elevated in 5 and 4 patients respectively. Seven pa-tients were prepared with infusion preoperatively to expand intravascular volume, and 2 patients were given prazosin 1.5 mg/d for 5 to 7 days. Results During the operation, seventeen patients had ele-vated blood pressure and 5 patients had no changed. One of seven patients with preoperative prepara-tion had obvious hypertension during operation, and 11 of 15 patients without preoperative preparation had obvious hypertension. The tumors were removed successfully in 21 patients. All the patients were diagnosed pheochromocytoma pathologically. Twenty-one patients had normal blood pressure with no recurrence during the follow-up from 1 month to 7 years. Conclusions The patients with normotensive pheochromocytomas may have lower catecholamine in their plasma and urine. The application of α-blockers and the expanding intravascular volume before operation could be important for the patients safe.