Purified hemoglobin was modified with pyridoxal 5-phosphate(PLP) and polymerized with glutaric dialdehyde(GDA) to get the products. By comparison of the physical, chemical and biological properties of different procedures for modification before and after polymerization, there is no significant difference in molecular distribution, methemoglobin(MetHb) concentration, oxygen carrier capacity, P50 and spectra. Furthermore, the procedure of modification after polymerization can save PLP greatly and decrease cost greatly. So the procedure of modification after polymerization is a better way in research and production. The addition of GDA could control the increasing of MetHb. By comparison on the physical, chemical and biological properties of different procedures, there is no significant difference in molecular distribution, MetHb concentration, oxygen carrier capacity and spectra between the procedure of adding GDA before PLP and that after PLP. But the P50 of adding GDA before PLP is much lower than that after PLP. So the procedure of adding GDA after PLP is a better way.
Blood Substitutes ; Chemical Phenomena ; Glutaral ; chemistry ; Hemoglobins ; chemistry ; Humans ; Methemoglobin ; chemistry ; Pyridoxal Phosphate ; analogs & derivatives ; chemistry

OBJECTIVETo investigate the influence of cross-linking agents genipin and glutaraldehyde on the composite bio-sponge.

METHODSThe composite bio-sponge was prepared with the technology of lyophilization. The degree of cross linking was determined using absorptiometry of trinitrobenzenesulfonic acid; the cytotoxicity was tested by MTT assay; the degradation rate in vitro was valuated by lysozyme degradation.

RESULTS(1) The degree of cross linking of composite bio-sponge crosslinked using genipin and glutaraldehyde increased with the crosslinking time, and reached 26.43% and 54.63% respectively after crosslinking 3 d. (2) The water absorption rate of composite bio-sponge crosslinked using genipin was better than that of crosslinked using glutaraldehyde. (3) In the initial stage of cells incubation, all extracts of composite bio-sponges crosslinked using genipin and glutaraldehyde inhibited the growth of the cells, and the inhibition decreased with the incubation time; but the cytotoxicity of composite bio-sponge crosslinked using glutaraldehyde was higher than that of crosslinked using genipin. (4) After soaking in saline for 4 weeks, the degradation rate of composite bio-sponge crosslinked using genipin or glutaraldehyde was 32.1%, 28.4%, respectively; however, after soaking in saline containing lysozyme for 40 h, the degradation rate of composite bio-sponge was 36.7%, 31.2%, respectively.

CONCLUSIONCompared with the composite bio-sponge crosslinked using glutaraldehyde, the degree of cross linking and the cytotoxicity of the composite bio-sponge crosslinked using genipin decreased; however, the water absorption rate and the degradation rate increased.

Five influencing factors of the polymerization process of glutaraldehyde with hemoglobin were studied in cluding the approach of feeding glutaraldehyde, hemoglobin concentration, the molar ratio of glutaraldehyde and hemoglobin, reaction time and temperature, in order to reduce the average molecular weight and to improve the effective polymerization ratio. The results showed that the optimal process reduced the average molecular weight of hemoglobin based oxygen carriers (HBOCs) from (350.20 +/- 35.45)kD to (158.60 +/- 8.70)kD and improved the effective polymerization ratio from 53.27% +/- 4.95% to 69.50% +/- 3.70%. When the experiments expanded 30 folds, the results of the effective polymerization ratio and the average molecular weight mentioned above could be achieved.
Female ; Glutaral ; chemistry ; Hemoglobins ; chemistry ; Humans ; Molecular Weight ; Oxygen ; chemistry ; Placenta ; Polymerization ; Pregnancy

Concanavalin A (ConA) is immobilized on a pre-activated chitosan microspheres, and then oriented immobilization of urease is carried out based on the strong interaction between ConA and glycoprotein. The optimum immobilization conditions are as follows: glutaraldehyde concentration is 3.5%, ConA concentration 1 mg/mL, ConA pH 7.0 and urease concentration 0.4 mg/mL. For orientedly immobilized urease, the highest activity was allowed at pH 5.0-6.0 and temperature 77 degrees C, and the Michaelis constant (Km) was disclosed to be 11.76 mmol/L by Lineweaver-Burk plot. Compared with the free urease and the randomly immobilized urease, the optimum pH of the orientedly immobilized urease becomes smaller and the pH domain wider. Orientedly immobilized urease presents higher temperature resistance, higher affinity to the substrate, and higher stability of operation.
Chitosan ; chemistry ; Concanavalin A ; chemistry ; Enzymes, Immobilized ; Glutaral ; chemistry ; Glycoproteins ; chemistry ; Hydrogen-Ion Concentration ; Microspheres ; Temperature ; Urease ; metabolism

AS1. 398 neutral protease was immobilized onto carboxymethyl chitosan with glutaraldehyde as cross-linking agent. The effects of pH, time of cross-linking, amount of cross-linking agent and the ratio of enzyme to carrier on the activity of the immobilized enzyme were demonstrated, and the optimum immobilization condition of AS1. 398 neutral protease was established. Also studied was the characterization of immobilized enzyme,including pH, temperature, Km and the stability of storage.
Bacterial Proteins ; chemistry ; Chitosan ; analogs & derivatives ; pharmacology ; Endopeptidases ; chemistry ; Enzymes, Immobilized ; chemical synthesis ; chemistry ; Glutaral ; pharmacology ; Microspheres

Nano magnetic microspheres prepared by chitosan and poly acylic acid were applied to purifying superoxide dismutase from blood erythrocyte. Chitosan-polyacyilc acid graft copolymer was synthesized by free radical graft copolymerization with potassium persulfate as inititator. To prepare Fe3O4 magnetic fluids with chemical coprecipitation, chitosan-polyacylic nano magnetic microspheres were prepared with glutaraldehyde as crosslinking agent. Structure of nano magnetic microspheres was detected by FT-IR spectrometer. Particle size and morphology were characterized by JEM-4000EX technology. Chitosan-polyacylic nanometer microspheres have good paticle cize distribution, magnetic responsiveness and protein adsoption. Activity, product yield and activity recovery of SOD after purification reached 6 727 U/mg, 21.1%, and 85.7% respectively. Purification of blood superoxide dismutase by chistosan-polyacylic acid microspheres has its renewable and feasible nature.
Chitosan ; chemistry ; Erythrocytes ; enzymology ; Glutaral ; chemistry ; Magnetics ; Microspheres ; Polymers ; Spectroscopy, Fourier Transform Infrared ; Superoxide Dismutase ; isolation & purification

Chitosan microsphere has been wildly researched in controlled release of protein and peptide drug because of its excellent mucoadhesive and permeation enhancing effect across the biological surfaces. The control of the size and size distribution of microspheres is necessary in order to improve reproducibility, bioavailability, and repeatable release behavior. In this work, uniform-sized chitosan microspheres containing insulin were prepared by a novel membrane emulsification technique combined with glutaraldehyde crosslinking method. In order to prepare uniform-sized chitosn microspheres, it is necessary to modify hydrophilic membrane into hydrophobicity. It is found that there exists a linear relationship between the size of chitosan microspheres and pore size of the membrane used, so it is easy to control the size of microspheres by using membranes with different pore size. In this study, the effect of different amount of crosslinker and crosslinking time on microspheres' morphology, encapsulation efficiency (EE) and release profile of drug in vitro were investigated. It is shown that the morphology of microspheres is more smooth and spherical, and the release rate is slower with the increase of amount of glutaraldehyde and prolongation of crosslinking time. When the molar ratio of amino group of chitosan to aldehyde group of glutaraldehyde is 1:0.7, and crosslinking time is 1 h, the highest EE was obtained (about 65%). Date obtained suggest that chitosan microspheres prepared by this new method would be a promising system for controlled release of protein drugs.
Biocompatible Materials ; chemistry ; Chitosan ; chemistry ; Cross-Linking Reagents ; Delayed-Action Preparations ; chemical synthesis ; Drug Carriers ; chemical synthesis ; Emulsions ; Glutaral ; chemistry ; Humans ; Insulin ; pharmacokinetics ; Microspheres ; Particle Size

With sodium alginate as a carrier and glutaraldehyde as the crosslinking agent, an improved immobilization method of beta-glucosidase for production of soybean genistein was developed. As compared with entrapment or entrapment-crosslinkage, crosslinkage-entrapment that beta-glucosidase was treated with glutaraldehyde and then entrapped in sodium alginate remained high loading efficiency and activity recovery, Effects of bead sizes, concentrations of alginate and glutaraldehyde as well, on the loading efficiency and activity recovery were assessed. When compared with the free enzyme, the optimum temperature, pH value and Km of the immobilized beta-glucosidase were respectively shifted from 50 degrees C to 40 degrees C, 4.5 to 4.0 and 2.57 microg/mL to 2.02 miocrog/mL. The stabilities of the immobilized beta-glucosidase were considerably better than that of the native enzyme. The immobilized beta-glucosidase was employed to genistein production, 84.94% of the activity and 56.04% of conversion were kept after consecutive use of 6 times.
Alginates ; chemistry ; Aspergillus niger ; enzymology ; genetics ; Enzyme Stability ; Enzymes, Immobilized ; metabolism ; Genistein ; chemical synthesis ; chemistry ; Glucuronic Acid ; chemistry ; Glutaral ; chemistry ; Hexuronic Acids ; chemistry ; Isoflavones ; chemistry ; Soybeans ; chemistry ; beta-Glucosidase ; chemistry ; metabolism

White-rot fungus manganese peroxidase (MnP) oxidizes a wide range of substrates, rendering it an interesting enzyme for potential applications. The stability of MnP can be improved by immobilization. With sodium alginate, gelatin, or chitosan as a carrier, and glutaraldehyde as the crosslinking agent, MnP was co-immobilized using the embed-crosslinked method and the adsorb-crosslinked method. The immobilization conditions and the partial properties of the three immobilized enzymes were investigated. When compared with the free enzyme, the optimum pH values and the temperatures of the three immobilized MnPs carried by alginate, gelatin, and chitosan were respectively shifted from 7.0 to 5.0, 5.0, 3.0 and from 35 degrees C to 75 degrees C , 55 degrees , 75 degrees C . The thermostabilities of the three immobilized MnPs were considerably better than that of the native enzyme. The chitosan-decreased by less than 5% even after repeated use for 6 - 9 times. The ability of decolorizing azo dyes in static and shaky situation by gelatin-immobilized MnP approached to the free enzyme, and there was no loss of enzyme activity during 2 repeated batch reactions.
Adsorption ; Alginates ; chemistry ; metabolism ; Biocatalysis ; drug effects ; Chitosan ; chemistry ; metabolism ; Dose-Response Relationship, Drug ; Enzymes, Immobilized ; chemistry ; metabolism ; Fungal Proteins ; chemistry ; metabolism ; Gelatin ; chemistry ; metabolism ; Glucuronic Acid ; chemistry ; metabolism ; Glutaral ; pharmacology ; Hexuronic Acids ; chemistry ; metabolism ; Hydrogen-Ion Concentration ; Kinetics ; Peroxidases ; chemistry ; metabolism ; Schizophyllum ; enzymology ; Substrate Specificity ; Temperature

Urease was immobilized in a simple and effective way by physical aggregation using a precipitant-ammonium sulfate, followed by chemical cross-linking using a bifunctional reagent-glutaraldehyde to form insoluble Cross-linked urease aggregates (CLUAs). The optimum pH, optimum temperature and Km of CLUAs were 8.0, 70 degrees C and 0.021 mol/L respectively. Compared with that of free urease, the thermal stability, storage stability and resistance of cross-linked urease aggregates to the exogenous proteolysis were enhanced. The efficacy of CLUAs for the treatment of rats with chronic renal failure was also studies. The rats with chronic renal failure caused by adenine were divided into 3 groups randomly:the control group (fed with 10 mL water /kg per day), Coated Aldehyde Oxystarch (CAO) group (fed with 20 g CAO /kg and 10 mL water /kg per day) and CLUAs + CAO group (fed with 20 g CAO /kg and 10 mL CLUAs /kg per day) in which CAO was used to absorb the ammonia produced from urea. The contents of BUN and Scr in serum before and after 2 weeks treatment were determined. In three groups, the level of Scr decreased slightly (P = 0.922, 0.972 and 0.225 > 0.05 respectively) after treatment. The level of BUN was not changed (P = 0.211 > 0.05) in the control group, but decreased greatly BUN in both CAO group and CLUAs + CAO group (P = 0.004 < 0.05 and P < 0.001 respectively). Furthermore, the decrease of the BUN level after treatment in the CLUAs + CAO group was more remarkable than that in the CAO group (P = 0.016 < 0.05), which showed that the CLUAs + CAD system was more efficient than the CAO system for the removal of urea in serum.
Adenine ; toxicity ; Animals ; Enzyme Stability ; Female ; Glutaral ; chemistry ; Hydrogen-Ion Concentration ; Kidney Failure, Chronic ; chemically induced ; drug therapy ; Male ; Random Allocation ; Rats ; Rats, Wistar ; Temperature ; Urea ; blood ; Urease ; blood ; chemistry ; metabolism ; therapeutic use