Preserving the original heparin structure of a novel low molecular weight heparin by gamma-irradiation.

Several preparations of low molecular weight heparins (LMWHs) obtained by physical depolymerisation (irradiation with gamma-rays) of pig mucosal heparin have been characterised by mono- and two-dimensional NMR spectroscopy. Integration of typical 1H- and 13C-NMR signals provided a useful quantification of their sulfation pattern. The availability of the corresponding parent heparins showed that the original structure (including that of the active site for antithrombin, as also confirmed by affinity chromatography) had not been significantly modified by the depolymerisation procedure. This process involves only a slight decrease of undersulfated sequences. A peculiarity of gamma-LMWH is the absence of the 'linkage region', commonly present in unmodified heparins and most LMWHs.

Characterization of sulfation patterns of beef and pig mucosal heparins by nuclear magnetic resonance spectroscopy.

Though differing only slightly in their degrees of sulfation, heparin preparations from pig mucosa and those from beef mucosa have consistently different 13C- and 1H-NMR spectra, which provide useful fingerprints for distinguishing the two types of heparin. Integrated areas of NMR signals associated with minor, undersulfated sequences (assigned by comparison with mono-dimensional spectra of selectively desulfated heparins and by analysis of two-dimensional spectra of heparins prepared from pig and beef mucosa) permit quantitation of differences in sulfation patterns. Undersulfation of pig mucosal heparins at position 6 of the hexosamine units, determined by 13C-NMR and expressed as percent glucosamines nonsulfated at C6 referred to total glucosamines, is substantially lower for pig mucosal heparins than for beef mucosal heparins (16.9-21.7% vs 36.7-40.7%; average values: 18.6% vs 40.3%). By contrast, undersulfation at position 2 of the iduronic acid units, determined by 1H-NMR and expressed as percent nonsulfated iduronic acid referred to total (sulfated + nonsulfated) iduronic acid is significantly higher for pig mucosal preparations (9.6-13.5% vs 2.1-2.7%; average values: 12.7% vs 2.3%). Pig mucosal heparins also have a significantly higher content of 3-O-sulfated glucosamine units, which are markers for the active site of heparin for antithrombin-III.

Pharmacologic profile of a low-molecular-weight heparin depolymerized by gamma-irradiation.

Low molecular weight heparins (LMWHs) are considered to be the agent of choice for the prophylaxis of DVT in medical and surgical patients. Conventionally, these agents have been produced by fractionation of or by chemical or enzymatic depolymerization of native heparin. The fractionated heparin retains many of its biological properties such as AT III affinity and sulfate content gamma-irradiation (60Co) has been used to depolymerize GAGs (De Ambrosi et al. In: biomedical and Biotechnological Advances in Industrial Polysaccharides, pp. 45-53). This procedures has now been used for the preparation of LMWH derivatives of varying molecular weight. The current studies examine the biochemical and pharmacologic profile of one such gamma-irradiated depolymerized heparin. In standard clotting and amidolytic antiprotease assays (PT, APTT, AXa, Alla), gamma-irradiated depolymerized heparin produced equal or stronger activity when compared to a LMWH produced by nitrous acid depolymerization and retained the ability to active AT III and HCHII. Initial results indicate that LMWHs produced by gamma-irradiation exhibit comparable antithrombotic actions to those produced by chemical depolymerization when measured in animal models of thrombosis. gamma-Irradiation may be a useful method for the production of LMWHs.

Pharmacodynamic properties of adrenal cortical extracts in comparison to synthetic corticosteroid mixture in the rat.

The effects of a total extract of adrenal cortex on corticosterone and ACTH plasma levels have been studied in the rat, either during circadian rhythms or in conditions of stress, in comparison to the effects of a synthetic corticosteroid mixture. A total extract of adrenal cortex, acutely and chronically administered at low and high doses, showed an inhibitory effect on plasma steroids and ACTH, whereas a synthetic corticosteroid mixture was more effective in producing such an inhibition. On the other hand, rats treated with a total extract of adrenal cortex responded better to stress than animals treated with a synthetic corticosteroid mixture. This difference may be due to the presence in the total extract of the adrenal cortex of certain substances having stimulatory properties on the steroidogenesis and/or on the activity of the hypothalamus-hypophysis-adrenal axis.