I've been trying to synthesize NH2-HA recently, but it failed. Can someone please tell me why and how I can improve it?
The synthesis steps were as follows: I dissolved 500mg of HA in ddH2O and adjusted the pH to around 6 using dilute hydrochloric acid. Then I added 500mg of EDC and 800mg of sulfo-NHS to activate the carboxyl groups of HA, and activated for 15 minutes. After that, I added 300μL of ethylenediamine, which brought the pH to around 7-8 (this is the appropriate pH value for the reaction, theoretically). Then the reaction was carried out for 2 hours. The whole procedure was carried out at room temperature.
First of all, EDC with or without NHS (or sulfo-NHS) sucks. Second of all, the pH should be kept around 6 with a buffer. Hydrolysis is your enemy, if the amino groups do not react with the activated carboxylic group, water will do.
You can have water, but you need to determine the best EDC concentration, the best pH, and the best time to add the amino-containing molecule. A pain in the ass. If possible use a dried organic solvent.
Hi, the concentration of your HA also the concentration of your solution should be considered
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