Methacrylation of Hyaluronic Acid (HA)
Materials
hyaluronic acid from streptomyces equi. (Fluka)
methacrylic anhydride (Aldrich)
5 N NaOH
95% EtOH
dialysis supplies
Methods
Make 1% solution (w/v) of HA in diH2O. Heat at 60°C with stirring until dissolved. Solution will be very viscous. Let cool to room temp. before proceeding with reaction.
In cold room, add 10-fold molar excess methacrylic anhydride dropwise to HA solution.
Maintain pH at 8-9. This will require frequent addition of 5 N NaOH to the reaction mixture over several hours. Allow reaction to occur on ice.
Let react at 4°C overnight with stirring.
Precipitate methacrylated HA (HA-MA) into chilled 95% EtOH (10-fold volume ratio of EtOH:HA solution).
Resulting precipitate is white and gooey. If desired, reprecipitation may be done.
Dissolve precipitated HA-MA in diH2O.
Add HA-MA solution to dialysis tubing (MWCO 10,000) and dialyze against several changes of diH2O for 2 days.
Lyophilize and analyze via NMR.