Depolymerization, of hyaluronic acid

Even less is known about the effects of ozone on carbohydrates. Buell et al. observed a decrease in the depolymerization of hyaluronic acid after treatment of the lungs of ozone-exposed rabbits (1 ppm for 1 h) with hyaluronidase. B. Goldstein et al. reported a loss in membrane neuraminic acid of red cells exposed in vitro to high concentrations of ozone. It would be important to study the effects of ozone on respiratory tract mucus, which is rich in carbohydrates, including neuraminic acid. This could indude determination of foe extent to which ozone is able to penetrate mucus that is unaltered, whether foe reaction of ozone with mucus results in the formation of cytotoxic intermediates, and evaluation of the interaction in mucus of ozone with other air pollutants, particularly sulfur dioxide. Of possible pertinence is a study by Falk et who observed that ozone produced a loss in foe viral hemagglutinating ability of snail mucus. 

Fox R B, Fox W K 1983 Dimethyl sulfoxide prevents hydroxyl radical-mediated depolymerization of hyaluronic acid. Annals of the New York Academy of Sciences 411 14-18... 

Other effects of ascorbic acid are probably also dependent upon the monodehydroascorbic free radical. One is the depolymerization of hyaluronic acid, first shown by Robertson et al. (R14). This is particularly likely, since subsequent studies have shown that the depolymerization was not prevented by catalase, as should have occurred if peroxide were the active component. There is no reason to believe that this hyaluroni-dase-like action is physiological. There is every reason to believe, however, that the monodehydroascorbic acid radical is formed in vivo. It is unlikely that all of the reactions in the body of such a reactive molecule are catalyzed by enzymes. 

Caputo, A. (1957) Depolymerization of hyaluronic acid by X-rays. Nature, 179, 1133-1334. Jiang, D., Liang, J., Noble, P.W. (2011) Hyaluronan as an immune regulator in human diseases. Physiological Reviews, 91, 221-264. 

Hultin (77) has derived an expression for the enzymic depolymerization of hyaluronic acid based on Staudinger s (182) equation relating specific viscosity and molecular weight. This equation permits the calculation of a microunit for enzymic activity. The formula was applied to data published by Madinaveitia and Quibell (112) and Swyer and Emmens (183). The values obtained showed fairly good agreement for the higher substrate concentrations. 

Glycosidie linkages are not entirely stable to heat even at close to neutral pH and degradation of hyahironic acid at elevated temperatures has been reported (Pantlitschko, 1952), particularly in phosphate buffer (Pigman and Rizvi, 1959). The depolymerization of hyaluronic acid by ascorbic acid is a well-known jihenomenon and the extensive literature on this subject has recently been reviewed (Suiidblad and Balazs, 1966). A... 

While all hyaluronidases can bring about the depolymerization of hyaluronic acid, the individual enzymes differ considerably in specificity, reaction mechanism, and the products of the catalyzed reaction (Meyer el al., 1960 Gibian, 1966). Apart from an enzyme from leech (an endo- glucuronidase) (Meyer el al., 1960), all other known hyaluronidases act upon the ondohexosarainide linkages. 

Interestingly, reaction of hyaluronic acid with EDC in the presence of primary amines affords only theN-acyl urea derivatives, as proven by C- and N-labeling experiments. Water soluble carbodiimides are also used in the depolymerization of polyuronides via reduction of their carbodiimide activated carboxyl groups. ... 

The in vitro anti-inflammatory effects of Cu-Zn SOD are now attributed to the prevention of hyaluronic acid depolymerization [106] and the prevention of chemotactic factor elaboration as a result of PMN produced superoxide [616-619]. However, the modest in vivo anti-inflammatory activity of Cu-Zn SOD in a variety of inflammatory diseases of man [620-628] has not been encouraging. Its lack of effectiveness is most likely due to its very short half-life (about 6 min), which may arise from the antigenicity of this intracellular enzyme in extracellular spaces, and its inability to cross cell membranes and protect intracellular targets of oxygen radicals due to its large mass and polar character. 

Thus, the free-radical-induced degradation of hyaluronic acid, another carbohydrate biopolymer, is a major aspect of rheumatoid arthritis, which has provoked radiation-chemical studies to elucidate the kinetics of its depolymerization (the lubricating property of hyaluronic acid in a joint is rapidly lost when its molecul2u- weight is decreased). 

Effect of chondroitinase ABC. This enzyme (chondroitin ABC lyase, EC A.2.2.A.) also depolymerizes the hyaluronic acid backbone of aggregate. In addition, it can degrade chondroitin sulfate chains of monomer, but not its keratan sulfate chains. 

Because of the great and continuing biological interest in the acid carbohydrates of connective tissue, there has been an intensive effort to locahze by histochemical methods the various carbohydrates that have been characterized by the chemical studies of Meyer and others (see Chapter XII). Although it is not yet settled completely whether hyaluronic acid is stainable by any of the customary methods, most workers have assumed it to be metachromatic with toluidine blue. Hyaluronidases (see Chapter XII) from various sources have been used (50), Much of the earlier work was done with impure preparations and needs verification. Bull-testis hyaluronidase can be obtained in concentrated form and has been most often used. As bull-testis hyaluronidase depolymerizes both hyaluronic acid and chondroitin sulfates of types A and C, metachromasia that is abolished by this enzyme has been considered due to one of these (114)-As the optimal conditions for the histochemical application of bull-testis hyaluronidase have not been systematically explored, technical directions vary greatly and need to be standardized. 

The hyaluronidases of bacterial origin are said to depolymerize only hyaluronic acid (114) and have been applied to a few histochemical studies (115), Unfortunately, only impure or crude filtrates of cultures have sometimes been used. There is need for more histochemical observations made with purified preparations of these specific hyaluronidases. The attempts to demonstrate hyaluronic acid illustrate the confusion that can arise from an insufficiently critical interpretation of results obtained with impure enzymes whose actions are poorly understood and with demonstration... 

The coprecipitation of hyaluronic acid and acidified protein in the form of a stringy mucin clot can be prevented by the depolymerizing action of hyaluronidase. This phenomenon forms the basis of the mucin clot prevention test (M. C. P.) developed by Robertson et al. (163) and modified by McClean (104,107). The method is well suited for serial determinations but gives only relative values since the assay results vary inversely with the concentration of the substrate (104). 

The use of heat to denature the proteins has been employed by several workers (92,107,187). Heating is recommended only for aqueous suspensions of ground cords where the tissue proteins are in suSicient excess to protect the polysaccharide from depolymerization. The extraction of hyaluronic acid with buffered trichloracetic acid has been proposed recently (82). 

Reaction Kinetics. The relationship between enzyme concentration and the amount of unreacted substrate is linear for a range of approximately 0.15 T. R. U. to 1.2 T. R. U. per milliliter (1,119,172,190). Furthermore, Harris and Harris (71) have observed a direct proportionality between the initial amount of hyaluronic acid and the amount of enzyme necessary to depolymerize all but a small, constant amount of the substrate. 

Assay by Reaction Rates. Lundquist (99) found that for low concentrations of hyaluronic acid in Mcllvaine sodium chloride buffer, the relative viscosity between 3 and 4 is approximately a straight-line function of the substrate concentration. In this viscosity range, the initial reaction rate is constant provided that the enzyme concentration is chosen to produce only 20% depolymerization. The activity of bull semen hyaluronidase was determined with several different substrate preparations. The assay results were in good agreement with the exception of one substrate fraction which was hydrolyzed six times slower than the other samples. The reaction rate constant was also used by Werthessen et al. (198) to determine hyaluronidase activity. 

Thus, ionizing radiation provides an excellent tool to reduce the molecular weight of a carbohydrate polymer whenever required for medicinal application, e.g. in the case of chitosan [83-85]. On the other hand, an increased stability against free-radical induced depolymerization might be required. This may be achieved by chemically crosslinking the native linear polymer the radiolytic behaviour of cross-linked hyaluronic acid, called hylan, has been studied [86]. 

The spontaneous, /8-eliminative depolymerization is a rapid procedure that can be measured either by the change in viscosity or by the increase in the u.v. absorption. It was furthermore observed that the ability for /3-eliminative depolymerization of the pectic material depends on the number of esterified D-galactopyranuronate residues in the glycuronan chain. This depolymerization proved to be an endocyclic, enolacetal-forming j8-elimination, similar to the enzyme-catalyzed process for hyaluronic acid, chondroitinsulfuric acids, and heparin,... 

Effect of Streptomyces hyaluronidase. The effects of some depolymerizing enzymes on the viscoelastic properties of proteoglycan were examined. The first example was that of Streptomyces hyaluronidase (hyaluronate lyase, EC 4.2.2.1.), which is shown as strictly specific to hyaluronic acid (j6). Thus this enzyme can degrade only the hyaluronic acid backbone of aggregate and should not attack the proteoglycan monomer. To 0.1% solution of aggregate 0.05TRU of Streptomyces hyaluronidase was added. As... 

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