Noriko Motohashi

Superoxide‐dependent formation of hydroxyl radical catalyzed by transferrin

Hydroxyl radicals are generated in the hypoxanthine‐xanthine oxidase system in the presence of iron‐saturated transferrin isolated from human serum. This has been demonstrated by colorimetrically measuring the hydroxylation of salicyclic acid and by EPR using the spin trap DMPO (5,5‐dimethyl‐1‐pyrroline‐N‐oxide). A Fenton‐type Harber‐Weiss reaction catalyzed y transferrin is proposed.

Thiol-induced hydroxyl radical formation and scavenger effect of thiocarbamides on hydroxyl radicals

The effects of thiols and thiocarbamides on hydroxyl radical (.OH) formation by the hypoxanthine(HYP)-xanthine oxidase(XOD)-Fe3+ .EDTA system were investigated in the range of 0.5-5 mM by colorimetrically measuring salicylate hydroxylation. Thiocarbamides powerfully inhibited the hydroxylation while thiols showed a paradoxical effect, enhancing it at low concentrations, but inhibiting it at high ones. Thiols in the presence of Fe3+ .EDTA generated superoxide anions (O2-.) and .OH during the oxidation, but thiocarbamides did not. A study of the effect of ergothioneine, a thiocarbamide present in mammals, on the .OH spin adduct of 5,5-dimethyl-1-pyrroline-N-oxide(DMPO) by EPR spectrometry showed that it effectively decreased the .OH spin adduct without causing the appearance of other signals. Reaction mechanisms are proposed for the O2-. evolution and .OH formation by the thiols themselves in the presence of Fe3+ .EDTA and .OH with thiols and thiocarbamides.

Analysis by high-performance gel permeation chromatography of hyaluronic acid in animal skins and rabbit synovial fluid

The simultaneous analysis of the molecular weight and concentration of hyaluronic acid in biological samples using high-performance liquid chromatography with two gel permeation columns is described. The elution volumes of various molecular weights of hyaluronic acids were linearily related to the logarithms of their molecular weights up to 600,000. The concentration of hyaluronic acid could be determined in the range from 20 to 100 micrograms/ml, i.e., from 4 to 20 micrograms per 200 microliter injected. The method was applied to the analysis of several animal skin extracts and rabbit synovial fluid. Skin extracts from mouse, rat, guinea-pig and rabbit could be chromatographed without prior isolation and purification. Hyaluronic acids in skin were separated clearly from chondroitin sulphates and their concentrations were determined. The molecular weights were estimated simultaneously to be more than 10(6). Rabbit synovial fluids from intact joints and saline- and carrageenin-treated joints could be chromatographed directly. The chromatograms showed that the concentration of hyaluronic acid in carrageenin-treated synovial fluid is lower than that in saline-treated fluid and the molecular weight distribution is broader. This technique enabled the rapid analysis of hyaluronic acid present at low levels in biological samples.

The effect of synovial fluid proteins in the degradation of hyaluronic acid induced by ascorbic acid

The degradation of hyaluronic acid induced by ascorbic acid and the effect of synovial fluid proteins, such as ceruloplasmin, transferrin, and albumin, were investigated on the basis of the elution volume and the molecular weight of hyaluronic acid using high-performance gel permeation chromatography. Hyaluronic acid was degraded to less than one-third of the original molecular weight in the range of the physiological concentrations of ascorbic acid. Synovial fluid proteins protected against the ascorbate-dependent degradation of hyaluronic acid at their physiological concentrations. It is suggested that the inhibitory activity of ceruloplasmin mainly depends on the ferroxidase activity and that of transferrin is probably due to iron binding property.

Uricase-like catalytic activity of ion-exchange resins modified with metalloporphyrins.

The uricase-like catalytic activity of the ion-exchange resins modified with metalloporphyrins has been investigated through the oxidation of uric acid. The anion-exchange resins modified with Mn(3+)-tetrakis(sulfophenyl)porphine and the cation-exchange resin modified with Mn(3+)-tetrakis(1-methylpyridinium-4-yl)porphine exhibited the highest uricase-like activity among the modified resins tested. The fact that these resins accelerated the oxidation of uric acid even after ten cycles of use indicates that the modified resins act as catalysts in the reaction catalysed by uricase. Some of the modified resins may be effectively used for the determination of uric acid in place of uricase.

Modification of gamma-irradiation-induced change in myoglobin by alpha-mercaptopropionylglycine and its related compounds and the formation of sulfmyoglobin.

The effect of some sulfhydryl-containing amides such as mercaptopropionylglycine (MPG), dimercaptopropionylglycine (DMPG), and α-mercaptopropionyl-L-cysteine (α-MPC) in the modification of γ-irradiation-induced change in myoglobin was investigated, and the results were compared with those for cysteine and cysteamine, and discussed in terms of their chemical structures. The effect of these thiols in the reduction of radiation-induced change in metmyoglobin decreased in the order, DMPG = α-MPC > α-MPG > cysteine = cysteamine ⪢ β-MPG. An abnormal green pigment, sulfmyoglobin, was formed from metmyoglobin solution by the sulfhydryl-containing amide but not by corresponding mercaptocarboxylic acids. When the metmyoglobin solution was irradiated in the presence of some thiols, the formation of sulfmyoglobin and metsulfmyoglobin was observed. The degree of the formation of sulfmyoglobin was found to be dependent on the ability of the thiols to release sulfur. The presence of the -CONH- bond in their molecules se...