دانلود رایگان مقاله تجزیه و تحلیل کمی پیوند عمده منطقه تترا ساکاریدها در هپارین

ABSTRACT

Heparin is a polysaccharide based anticoagulant drug composed of a complex mixture of glycosaminoglycan chains and peptidoglycosaminoglycan chains. In an effort to better characterize this important polysaccharide based drug, we examined the peptide components of the minor peptidoglycosaminoglycan chains comprising heparin. Three different the glycan-peptide linkage regions tetrasaccharide fragments were isolated from pharmaceutical heparin using heparin lyase II and characterized the structure of these tetrasaccharides using nuclear magnetic resonance spectroscopy and mass spectrometry. A sensitive and quantitative assay was developed for these linkage regions using multiple reactionmonitoring tandem mass spectrometry. These three different linkage regions were found in heparins coming from porcine intestine and bovine lung. Two of these were also present in the low molecular weight heparin, enoxaparin.

4. Conclusions

While wehave a generalunderstanding oftheprocess toprepare heparins and LMWHs, the specifics of individual manufacturing processes are trade secrets (Bhaskar et al., 2012; Linhardt & Gunay, 1999). Moreover, such process steps can also take place to the crude heparin, at either the slaughterhouse or consolidator levels, before it moves to a cGMP facility under regulatory oversight. This makes their detection only possible through forensic analysis. The analyses described in this manuscript provide some insights into the differences in such processes. The strength ofthe oxidation step can lead to the oxidation and the use of peracetic acid in the bleaching step can lead to acetylation of the serine residue in the pG chains of a heparin product. Thus, the modification of the serine residue can serve as an indication of these process steps. These pG chains are only present in very minor amounts within a heparin product, which is primarily composed of GAG chains. Thus, modern analytical methods together with the preparation of oligosaccharide standards offer an approach to better understand and ultimately control these process artifacts.