Oligodcoxyaucleotide-doxorubicin conjugate is a novel modified oligodeoxynucleotide (ODN) to inhibit the expression of mdrl gene. The present study was undertaken to determine the stability of the conjugate in virto and its pharmacokinetics in vivo using a reversc-phase HPLC assay. The method employed a Sort C18 reverse phase column combined with a C8 prc-column and a linerar gradient elution with acetonitrile containing 0.1M aqueous triethylammonium acctate, pH7.0 Detection was carried out using a UV-didoe array detector at 254 and 480nm. Minimum sensitivity of ～0.15μg/mL in plasma and 0.1μg/mL in PBS of hte conjugate was achieved. After incubation in 10% activated fetal calf serum for 24 hours, 15.8% of the conjugat was degraded. As a comparison however, 97.5% of the control ODN was degraded witin the same incubation time. The phamacokinetics stuies show that the halif-lives of the conjugate is about 8 hours, 4 times longer than ODN as a conro. Assay validaton studies revealed that the method is accurate, reproducible for dcterminatin of the conjugate, and can be used for a pharmacokinttic study of the conjugate.

Reconmbinatn antibodies, especially ScFv fragments can be applied as detection reagents and even substitute for some reagents used in immunoassays such as antibodyenzyme conjugates. For ScFv fragments there is no such universal system available up to now. A vector ystem was onstructed based on pPIC9-Fc, in which the hinge, CB2 and CB3 domin (Fe fragment) of mouse IgG1 and HIS-tag wee eloned into the Pichia expression vectro pPIC9. A model SeFv was introduced into pPIC9-Fc. which can bind Gluathione-S-transfrease (GST) from Schistsoma Japonicum to yield the fusion was expreessed at high levels in the methylotrphic yeast Pichia Pastoris, secreted as a dimeric form in the culture, ad purified by Ni2+-NTA column chromatography. The expression yield can reach 10-30mg/liter of culture medium. The SeFv-Fc fusio retains the bioglogyical binding ability of the parent Scfv, and can be applied as anti-GST antibodies for the detection of GST and GST-fusion proteins. Furthermore. the successful expression and maintenance of the binding activitv verify the efficacy of the vector system for use as detection reagents in vitro, by reacting with the specific antigens and being readily detected using general anti-mouse antibodies.

Oleic acid esters were shown to be the best carbon source for both cell growth and lipase production by Candlida rugosa. Use of cosolvent, dodecane, in fermentations improved the solubility of solid substrates and incresased oxygen solubility. This resulted in the hightst lipase ativity in batech fermentation with glycerol triolcate and dodecane. Lipase activity reached 77.1 units ml1.

Reactive oxygen species ate beliceved to play a role in the development of several diseascs incloding vascular disenased and theaging process. It is reported that increased reactive oxygen species were impliated as an important mechanism that contrhibutes to endothelial dysfunciton, So, human umbilical cord vein endothelial cells were used to study the antioxidative effect of L-ascorbic acid and its derivative. The study indicalted that L-ascorbic acid as a tradilonal antioxidant was instable and could protect the cells against hydrogen peroxide induced cytotoxicity as its concetration beiow 50 μg/ml, but hadly could rotect the cells against tert-butyl hydroperoxide induced cytotoxicity, Ascortic acid-2-o-phosphate-6-o-palmitate could effctively protect the cells against hydrogen pcroxide and tert-butyl hydropemoxide induced cytotoxicity, and exhibited no cytotoxicity within the tested concentratin rage. The study indicated that ascorbic acid2-2o2phosphate-6-o-palmite could not only signficantly reduce the intracellular reactive oxygen species level in 3h culture. but also increase the cell viability in 15h culture. In additon. ascorbic acid-2-o-phosphate-6-oi-palmtate could keep stable in RPM1-1640 medium and water for 4 dya. permecat the cell membrane. which in tum may seavenge the intracllular reactive oxygen species. increase the cell viability and the plasminogen activatrs'activity. All above rsults suggested that addition of some hydrophobie groups to the traditonal antioxdants could from novel compounds with better properties.

L-ascorbic acid 2-phosphate-6-palmiate (Asc2P6p) was synthesized and its effect on the damage of PC12 cells induced by H2O2 was investigted. 200μM H2O2 in a treatment pcriod of 4 hours in our experiment resulted in substantial cell loss. With the increasing concetration of antioxidants. such H2O2-induced cytotxicity was significantly prevented and the corresponding intracellular and extracellular ROS levels decreased concurrently by pre-treatment with Asc2P6P and Asc. It was found that Asc2P6P was superior to L-ascorbic acid in its protective role and showed a dose-depcndent manner during a 24-hour tretment. The higher potency of Asc2P6P's protective role on PC12 cells was correlated with its more effective ROS seavenging ability. HPLC assay its distinguished rle in protechiting PC12 cells againts H2O21-induced cytotoicity.