Preparation of low molecular weight heparin using an ultrasound-assisted Fenton-system

Heparin, a high-molecular weight acidic polysaccharide, has raised much interest in the field of biomedical research due to its multiple bio-functions. The anticoagulant application of heparin in routine clinical practice, however, has been limited as the large molecular size of heparin can reduce its subcutaneous bioavailability and lead to severe adverse consequences such as thrombocytopenia. Here, we report a highly efficient and convenient method to depolymerize high-molecular weight, unfractionated heparin (UFH), into low molecular weight heparin (LMWH) by combining physical ultrasonic treatment with the chemical Fenton reaction, referred to as sono-Fenton. We found that this combination treatment synergistically degraded UFH into a LMWH of 4.87 kDa within 20 min. We characterized the mechanism of sono-Fenton heparin degradation through multiple approaches, including HPLC-SAX, disaccharide composition, FT-IR, NMR and top-down analysis, and found that the uronic acid residue in heparin was the most susceptible site attacked by OH radicals produced in the sono-Fenton process. Importantly, the LMWH prepared by this method had significantly higher anticoagulant activity than UFH and other LMWHs. This approach represents an effective method to produce heparin with improved activity and should be potentially useful for heparin production in the pharmaceutical industry.