I am trying to conjugate some compounds having -COOH groups with my MNPs, using EDC_NHS coupling reaction but I am not getting the peaks around 1700 cm-1 for free -cooh groups on MNPs. I used MES buffer for EDC and NHS activation at pH 6, and later raised the pH with PBS to ~ 7. My particles get dissolved at pH 8 in
CO3--/HCO3- buffer and I can not separate them. The pKa of -NH2 group of amino acids are near 9, so at pH7 that I am using for coupling they must be positively charged. My question is how can I couple the -COOH group effectively to my MNPs? I can not go beyond 7 pH, and at >8 the EDC ester would be very unstable also. Thanks in advance for kind suggestions.
hello,
when you say you are trying to couple -COOH with the -NH2 functionalized MNPs, why are you searching free -COOH group peak at 1700 cm-1 on your MNPs. (defiantly free -COOH will not be on the MNPs)
I don't get the purpose and route you are following for the synthesis, but i would suggest that you can separate out the -NH2 functionalized MNPs and then add it in the solution of EDC activated -COOH derivative. keep the solution under inert atmosphere with continuous nitrogen flow. stir the mixture for 24 hours at room temperature. EDC derived acid amide coupling is very simple reaction. i hope this will work.
Best of luck.!!
Thanks Gaurav for the answer, I have separated the -NH2 functionalized MNPs and for further application purposes I need the -COOH groups on the surface, so I am using the chemicals with 2/more -COOH groups for conjugation with these MNPs, so that after EDC conjugation at least one/two are there for further use. That's why I am searching the peaks at around 1700 cm-1.
Why are you suggesting for inert atmosphere I could not understand. I performed the reaction two times, but I am not getting results.
Most EDC coupling reactions are carried out in organic solvent. inert atmosphere is advised when reaction is carried out in organic phase to avoid deactivation of EDC activated -COOH group.
but if you are carrying the reaction in water media then you need to take care of other precautions like use of HOBt, COMU, etc.. to stabilize the EDC activated acid and avoid hydrolysis. in the water media case you need not to used inert atmosphere by N2 purging, as other reagents are being used to stabilize the EDC activated -COOH group.
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