[A novel fluorophotometric method for assaying free glutathione of both reduced and oxidized in plasma].

To investigate the measurement of free glutathione of both reduced or oxidized (i.e. GSH and GSSG) in plasma, and to evaluate the redox state of GGSH/GSSG in human plasma, both GSH and GSSG in plasma were measured by fluorophotometry based on the facts that one molar GSSG can be reduced to two molar GSH by dithiothreitol(DTT) under the condition of the pH being about 6.0, and GSH can provide both primary amine and thiol groups to react with the two carbonyl groups of O-phthaldehyde (OPA) to form a fluorescent ternary isoindole complex at pH 8.0. This method can at least measure 16 picomole GSH and 8 picomole GSSG respectively in the tube. The variation coefficient (CV) for intra-ssay and intera-ssay is about 4. 6% and 3.9% for GSH and 3.5% and 4.1% for GSSG respectively. The recovery of GSH and GSSG added to the plasma is (99.77% +/- 5.70)% and (99.28% +/- 4.73)% respectively. The concentration of GSH and GSSG in the plasma of young healthy volunteer is (16.5 +/- 2.4) nmol x mL(-1) and (1.7 +/- 0.35)nmol x mL(-1) respectively, without significant difference between male and female. This measurement method is simple with great sensitivity and selectivity for rapid measuring GSH and GSSG in human plasma simultaneously.

[Superoxide dismutase biosensor for screening substances possessing scavenging superoxide anion activity].

AIM: To establish a method for screening active substance with scavenging effects on superoxide anion in vitro by designed superoxide dismutase biosensor. METHODS: The enzyme sensor was built by connecting the immobilized CuZnSOD with optical oxygen sensor through a special way. Superoxide anions were generated by auto-oxidation of pyrogallol. The auto-oxidation speed was examined before and after adding samples into the system, and the Vit C having the scavenging radical activities was served as a positive control. RESULTS: The limit of biosensor detection was 7.0 U in activity, and lifetime of the immobilized enzyme in the reaction-cell was above 2 weeks. The scavenging effects on superoxide radicals of fifteen active substance were studied in vitro by the sensor, and some of them presented scavenging activities. CONCLUSION: The signal from biosensor is stable, easy to be determined, and the kinetic information on scavenging superoxide radicals could be obtained directly. The biosensor system can be used for screening drugs simply and rapidly.

[Preparation and transfection of GFP plasmid DNA/cationic liposome complex].

OBJECTIVE: This work was directed at obtaining a better gene carrier to improve the effects of gene delivery. METHODS: Cationic liposomes made from cholesterol, lecithin and Eighteenth Amic by reverse evaporation technique were used for encapsulating plasmid DNA containing gfp gene. The DNA/liposome complexes differed in quantity of plasmid DNA. The sizes of complexes and the efficiency of encapsulation were detected. MTT assay was used to measure the cytotoxicities of complexes. The efficiency of transfection into COS1 cells was shown by observation of green fluorescent and measurement of fluorescent intensity. RESULTS: The average size of complexes was 562 nm, the encapsulating efficiency of DNA in microspheres reached 55%-65%. These DNA/Cationic liposome complexes could be transfected into mammalian cells, but they were different from each other in efficiency of transfection. The cytotoxicities of these complexes varied with the concentration of DNA in complexes, the quantities of complexes and the time of treatment by complexes. CONCLUSION: DNA/Cationic liposome complexes prepared by reverse evaporation technique could be applied in DNA delivery system.

[Study on the tumor cell targetability of folate-conjugated albumin nanoparticles].

OBJECTIVE: To study folate-conjugated albumin nanoparticles targeting to tumor cells via folate receptor-mediated endocytosis. METHODS: The activated folic acid (N-hydroxysuccinimide ester of folic acid) was conjuated to the surface of bovine serum albumin nanoparticles (BSANP) via the amino groups. The extent of the influence that concentration, incubation time and free folate exerted on the uptake of Folate-BSANP by ovarian cancer cells (SKOV3) was determined using fluorescence spectrophotometer. RESULTS: Folate-conjugated BSANP (Folate-BSANP) was successfully achieved. Uptake of Folate-BSANP by cancer cells was gradually increased with the extension of incubation time or the increase of Folate-BSANP concentration, and the uptake could be competitively inhibited by excess free folate. CONCLUSION: Folate-BSANP could be delivered into tumor cells via folate receptor-mediated endocytosis and significantly targeted to tumor cells with rich folate receptors.