Following the enzymatic digestion of chondroitin sulfate by a simple GPC analysis

摘要: Abstract We describe the use of gel permeation chromatography (GPC) setup with four size exclusion columns for analysis enzymatically digested glycosaminoglycans (GAGs). This provides information about molecular weight (Mw) and concentration all species (low high Mw) present in digests a single measurement. The data fraction Mw (often omitted GAG digests) provide direct evidence mechanisms action enzymes. proved feasibility this methodology by applying it to chondroitin sulfate (CS) substrates different sulfation pattern using enzymes (hyaluronidase chondroitinase). NMR obtained fractionated ultrafiltration confirmed results GPC reveal further details degradation mechanisms: (i) both preferentially attack 4-sulfated (ii) additionally well documented endolytic activity hyaluronidase we also observed low lyase enzyme reflected detected minor exolytic breakage. Finally, demonstrate that CS medium (12–60 kDa) which is sulfated mainly at 6-position can be good yields enzymatic digestion following ultrafiltration.