Methacrylation of Hyaluronic Acid (HA)

Materials

hyaluronic acid from streptomyces equi. (Fluka)

methacrylic anhydride (Aldrich)

5 N NaOH

95% EtOH

dialysis supplies

Methods

• 1. 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.

  2. In cold room, add 10-fold molar excess methacrylic anhydride dropwise to HA solution.

  3. 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.

  4. Let react at 4°C overnight with stirring.

  5. Precipitate methacrylated HA (HA-MA) into chilled 95% EtOH (10-fold volume ratio of EtOH:HA solution).

  6. Resulting precipitate is white and gooey. If desired, reprecipitation may be done.

  7. Dissolve precipitated HA-MA in diH₂O.

  8. Add HA-MA solution to dialysis tubing (MWCO 10,000) and dialyze against several changes of diH₂O for 2 days.

  9. Lyophilize and analyze via NMR.