The ―COOH of nanoparticles was activated using EDC/NHS and then covalently attached to the ―NH2 of the enzyme, forming ―CO―NH―. Now, which peaks number will show that the conjugation successfully done?
Hi rozita,
imdont have have personal experience with this system, but I always start with 2d surfaces as a model system for identifying important peaks.
My colleagues shuster vaish Andrews and Weiss at Ucla wrote several articles on this topic. The following should be particularly helpful as it pertains to extremely dilute functional molecules being chemically modified by that chemical system on flat gold surfaces.
http://pubs.acs.org/doi/abs/10.1021/jp207396m?journalCode=jpccck
it should help you identify which peaks in your composite system are most important.
Other papers cover protein conjugation, but I fear the protein may bring too many additional functional groups to the system for ftir to be useful In a quantitative sense.
i wish you luck with your analysis!
Hey Rozita,
Nice question and there are several articles on this area. I think there might also be hydrogen bonding in your system so it would be interesting to look at the shift in the -CO frequency. I have done some work in this area where have been shift in the -CO freq due to hydrogen bonding and even in my system i had some amide linkages which i think you are also having.
regards
Shashank
Usually, the spectrum of the nanoparticle should be taken before and after the intervention (enzymatic or chemical intervention). Then the two spectra should be compared to check for the appearance of new peaks, the disappearance of some peaks, and the shift (dislocation) of other peaks. From these changes, you can deduce if your intervention was successful or not.
Hello, the -COO-NH peak will appear at 1716 cm-1.
This reference might be useful for you.
http://www.sciencedirect.com/science/article/pii/S0167577X12014826
Salam
I think you should take FTIR spectra of the enzyme before and after incorporated by the activated nanoparticle. Please you pay more attention in 3500-3200 cm-1 wave number broad band range for N-H stretching vibration representative, in around 1646 cm-1 for C-N bending vibration, and in around 1700 cm-1 for C=O stretching vibration. The different pattern and intensity of the spectra between before and after incorporated in the wavelengths ranges can be used to justify the successful of your experiment. It will be better if you also take FTIR spectra of the associated nanoparticle and compare it to the FTIR spectra of nanoparticle-incorporated enzyme.
When N-H is present we observe a pointed small peak at around 2600 to 2800 cm-1
The functional group appearing is an amide bond (―C(O)―NH―).
There are three regions in an IR spectra assigned to this functional group:
amide A at around 3300 cm-1, amide 1 at 1700 - 1600 cm-1 and amide 2 at 1580 - 1480 cm-1
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