Masatake Hori

Analytical and preparative methods for polymyxin antibiotics using high-performance liquid chromatography with a porous styrene-divinyllbenzene copolymer packing

Abstract Polymyxin antibiotics, such as polymyxin A, B, D, E, K, M and P, circulin and colistin, are a complex of two or three components. These components have been succesfully separated on a porous styrene—divinylbenzene copolymer column with methanol—0.2M potassium chloride/hydrochloric acid buffer (pH 2.0) (1:1) as mobile phase. Hitachi gel 3011 (particle size 10 μm) gave a good analytical separation of polymyxins using the isocratic mobile phase. In the preparation, all of the components of colistin and polymyxin B are separated by use of Hitachi gel 3010 (particle size 25 μm)or Amberlite XAD-2(100-200 mesh) packings. A new component of polymyxin B, named polymyxin B0, was deduced from the results of the amino acid and fatty acid analyses.

Leakage of immobilized IgG from therapeutic immunoadsorbents.

AbstractIn developing therapeutic immunoadsorbents (IAs), antibodies (IgG molecules) covalently immobilized on porous carriers, a leak of IgG was determined both in the storage test with buffers at 25 and 4°C and in contact with plasma at room temperature (RT). The amount of antibody released from therapeutic IAs must be minimized to avoid side effects during treatment. The amount of IgG released wasa.Dependent on the amount of IgG immobilizedb.Much greater with CNBr-activated Sepharose 4B, or FormylCellulofine as a support material than with aminopropyl CPG (controlled pore glass)c.Found to yield again during another storage in buffers after the IAs were washed and their buffers replaced with fresh ones andd.Decreased after the IAs were treated with glutaraldehyde (GA) solutions. Whereas treating the IAs with GA solutions significantly reduced the amount of IgG released, it caused some deterioration of the adsorption characteristics of the IAs. An irradiation dose of 2.5 Mrad as a crosslinking procedure also reduced the amount of IgG released; its effect was comparable to that of 0.025% GA, the lowest concentration used.

Studies of antipyretic components in the Japanese earthworm

Abstract Antipyretic components in the Japanese earthworm ( Lumbricus Spencer, Perichaeta communishima, Goto and Hatai ) were investigated through a study of the antipyretic effect on rabbits having pyrogen-induced fevers caused by Esherichia coli pyrogen and Chromatographic separations. The main proven antipyretic components are considered to be all- cis -5,8,11,14-eicosatetraenoic acid and all- cis -5,8,11,14,17-eicosapentaenoic acid.

Isolation and characterization of euglobals from eucalyptus globulus labill. by preparative reversed-phase liquid chromatography

Abstract The isolation of eleven novel acetogenin mevalonates, which have strong granulation-inhibiting activity, from Eucalyptus globulus Labill. is reported. These eleven compounds, which have closely related structures, were successfully isolated with a preparative reversed-phase column using acetonitrile or methanol as the eluent.

Specific removal of IgE by therapeutic immunoadsorption system

A therapeutic immunoadsorption system was developed that can remove IgE effectively and specifically from the plasma of patients with an allergy or other hyper-IgE syndrome. The immunoadsorbent (IA) consists of immunoaffinity purified anti-IgE antibody (a-IgE ab) immobilized on controlled pore glass beads (50 nm pore size). Adsorption isotherms for IgE, which were reduced by the Freundlich adsorption equation, were obtained with IA that immobilized various amounts of a-IgE ab. An optimum amount of a-IgE ab to be immobilized was selected. IA worked sufficiently in a wide range of IgE concentrations. Clinical treatment requires an amount of 41 mg of IgE to be removed from a patients plasma for 3 h. An IA for clinical use was designed to contain 10 g of the support binding 325 mg or more of the antibody. In fact, our study in vitro simulating a clinical case showed that serum IgE was removed by IA, as expected: the level decreased from 11,000 to 3000 U/ml after a 3 h perfusion (1 U = 2.3 ng). A very small amount of a-IgE ab (goat IgG) was found to be detached from IA by flowing plasma; the average level was 20 ng/ml, which seems to be safe. However, we installed the second column in a circuit that adsorbs a-IgE ab leaked into plasma, because the amounts of a-IgE ab infused into the patient must be minimized. The second column contained IgE immobilized on the same support, since IgE as a ligand adsorbed more a-IgE ab than did anti-goat IgG antibody. This is an effective and safe therapeutic immunoadsorption system and has been subjected to clinical tests.

A new fluorometric analysis of citric acid

Abstract A new fluorometric method for analysis of citric acid in blood and plasma was established, where the citric acid react with o -aminothiophenol in 35% phosphoric acid solution, at 125°C, for 15 hr. The fluorescent substance was extracted by ethylacetate and measured with excitation at 415 nm and emission at 450 nm. Only 0.05 ml of human blood or plasma is required.

Sterilization of therapeutic immunoadsorbents with aqueous propylene oxide solution.

Propylene oxide (PO), in the form of its aqueous solution, was evaluated as a chemical sterilant for therapeutic immunoadsorbent to be used in on-line plasmapheresis. Generally, PO is characterized as less toxic and less antimicrobial than ethylene oxide. PO was significant sporicidal to B. subtilis at 25°C and 10β spores/ml were killed with 1% (V/V) PO solution after storage for 3 weeks when 75% to 80% of PO decomposed; the death-rate conformed to the kinetics of a first-order reaction although PO decomposed rapidly. Thus an apparent D values were estimated to be 13 to 16 days with 0.5% PO solution and 5 to 7 days with 1% PO. An immunoadsorbent consisting of Sepharose 4B immobilizing anti-human IgE antibody was used to study the effect of PO on adsorption characteristics. PO affected the IgE removal of the immunoadsorbents depending on its concentration ranging from 0% to 4%. The IgE-adsorptions were lowered by only 17% and 30% of the intact and glutaraldehyde-treated immunoadsorbents respectively after storage for 3 months in the 2% PO solution at 25°C, and these numbers are considered to be acceptable for a clinical use. A 2% PO solution was chosen based on the results of the shelf life test of immunoadsorbents and the sporicidal test of PO.