Abstract
BackgroundThe purpose of this study was to develop and validate an analytical method to determine the presence of hyaluronic acid derived disaccharides in equine synovial fluid.FindingsA high-performance liquid chromatography method for the determination of hyaluronic acid derived unsaturated disaccharides in equine synovial fluid was developed and validated. The method is based on the measurement of unsaturated disaccharides released by digestion of linear hyaluronic acid molecules. The method showed linearity (r2 = 0.996) over the full working concentration range 0.89-30 mg/l. Relative standard deviation of intra- and inter-day precision ranged from of 4.3-6.7% and 7.1-7.8% respectively. The detection limit was 0.3 mg/l corresponding to 20 mg/l in synovial fluid. Accuracy of the assay was 97-103%. This method was evaluated by determining the concentration of unsaturated disaccharides from hyaluronic acid in synovial fluid of horses with lameness in the metacarpo-/metatarsophalangeal joint localized with positive response to intra-articular anesthesia.ConclusionsThe described method is valid for determination of hyaluronic acid derived disaccharides in equine synovial fluid. This method was applied to a larger research project dealing with a new form of intra-articular therapy in horses with arthritic diseases.
Highlights
The purpose of this study was to develop and validate an analytical method to determine the presence of hyaluronic acid derived disaccharides in equine synovial fluid
The described method is valid for determination of hyaluronic acid derived disaccharides in equine synovial fluid
Hyaluronic acid (HA) is a linear polysaccharide formed of disaccharide units which are linked together to form the HA chain
Summary
The purpose of this study was to develop and validate an analytical method to determine the presence of hyaluronic acid derived disaccharides in equine synovial fluid. Conclusions: The described method is valid for determination of hyaluronic acid derived disaccharides in equine synovial fluid. By digestion of HA to unsaturated disaccharides and labelling with a hydrophobic fluorescent agent, the analysis is enabled by reverse-phase high-performance liquid Reverse-phase HPLC has been used to analyze HA-disaccharides in biological samples [10], to our knowledge the method is not validated for equine SF.
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