Ninhydrin is commonly used for the quantification of amine groups and proteins. The method is also based on the measurement of Vis absorbance and quantification using a calibration curve.

Apart form the former methods, you can also use the fluoroscamine test, which uses the binding of fluoroscamine to amines to examine the fluorescence increase of the dye. For quantification, a calibration curve is also needed.

you can use XPS tehnique. From C, O, N and Fe spectra you can have information you needed.

Thermogravimetric Analysis (TGA) would be a quick way to find out how much organic material burns off from your iron oxide particles. From there, it's an easy calculation to mmol amine per gram of particles.

You may use Angle resolved XPS, to probe quantitatively and qualitatively the extreme surface after functionalization. In the same time if you wanna check the change in the surface properties such work function, or HOMO level change you may combine your analysis to UPS. Good luck

Thank you all for your valuable suggestions

@ Ana Luísa Daniel-da-Silva:

Thank you mam. Can you please provide me any reference or protocol for using ninhydrin to detect amino functional groups?

I dont have XPS facility .. so i have to try other options. All XPS suggestions were very informative. Thank you all

@ Jeremy Steinbacher . Thank you sir. I shall try TGA. Please provide me any reference if possible.

Hi, well, TGA is so ubiquitous that I don't think a citation is really needed. I've made up a PDF of the procedure for you to read. Let me know if I can help further or if you need an better explanation of anything I wrote. Cheers!

Oops, I made one small mistake on that PDF. Here's the corrected version.

@Jeremy Steinbacher Thank you for your concerned reply and effort. I understood your simple and precise explanation.

Glad I could help! Let me know how your TGA experiments go.

@ Prof. Zoran Vujcic : Dear sir , i am not able to understand why you are asking such a question.. what makes you doubtful .. Please answer these questions.

Answer for your question:

Yes I have prepared Amino functionalized Fe2O3 nanoparticles as well as graphene nanosheets coated with Fe2O3 nanoparticles further modified with aminopropyltriethoxysilane

Ref: J. Phys. Chem. B 2006, 110, 1553-1558

ACS Applied Materials & Interfaces VOL. 2 • NO. 11 • 3201–3210 • 2010

@ Prof. Zoran Vujcic : Yes sir i prepared magnetic nanoparticles ..Magnetic Fe2O3

( magnetic saturation (Ms) value AminoFunctionalized-MNPs 11.22 emu g-1 ) were prepared by co-precipitation method "He, Y. P.; Wang, S. Q.; Li, C. R.; Miao, Y. M.; Wu, Z. Y.; Zou, B. S.Synthesis and characterization of functionalized silica-coated

Fe3O4 superparamagnetic nanocrystals for biological applications.

J. Phys. D: Appl. Phys. 2005, 38 (9), 1342–1350."

NO aggregation was observed as it was immediately treated with tetraethylorthosilicate followed by aminopropyltriethoxysilane.. Pls see attachment for Scheme.

Hi, If you have Atomic absorption or ICP-MS, the copper is easy to react with amine and by calculating the total mass of copper that was up-taken by amine; you can calculate the number of amine groups on the surface

@Nezar Khdary : Thank you Sir for the above interesting answer. Unfortunately in my institute there is no ICP-MS or Atomic absorption spectroscopy. I shall possibly give my sample to nearby institute and try to obtain the data. Your answer shall certainly be useful for other followers those have ICP-MS or Atomic absorption spectroscopy in their institute.

Hi. How accurate do your amine measurements have to be?

The amine reactive dyes listed above will probably give you a fair estimate, barring quenching, FRET, steric hindrance, etc.

If you need an absolute value for the amine groups you will probably have to use small 3H or 14C amine reactive compounds, like acetic anhydride.

As well as prof Vujcic said do HCl titration to determine the number of present free amines.

@Prof. Zoran Vujcic : Thank you very much for your suggestion. I shall try the experiment and shall share my observations/results in this discussion.

You can used XPS to measure it

Try TGA and titration method. In the latter you will find less or equal values to the TGA because maybe not all the titrant reacts with the amine groups.

please see the following paper, may be it will help you.

Slavko Kralj, Miha Drofenik and Darko Makovec

Controlled surface functionalization of silica-coated magnetic nanoparticles with terminal amino and carboxyl groups

Journal of Nanoparticle Research

Volume 13, Number 7 (2011), 2829-2841, DOI: 10.1007/s11051-010-0171-4

XPS is a useful method to determine the functional groups on the nanoparticles surface.

Hi Ana ! i quantified amine sites by doing Back Titration ..

Disperse 10 mg of amino terminal fe3o4 nanoparticles in 20mL of 1.0mMHCl solution and stir for 30min.

Separate nanoparticles by centrifugation. Titrate 10mL of supernatant with

the standardized 1.0mMNaOH solution in the presence of

phenolphthalein indicator. From the difference of HCl concentration, you can calculate

the molar concentration of amine sites.

This value can be converted into the number of amine sites per unit area (nm2)

of the surface of nanoparticle based on the density of and the surface area of nanoparticle.

Other options...

TGA could be done with even 1 mg of sample..

Use RITC Rhodamine isothiocyanate or Flourescein isothiocyanate and do quantification by fluorescence intensity

Gud luck

Maybe somebody has mentioned this already. There is a commercially available dye called fluorescamine, which is suitable to quantify surface amine groups. It binds to amines and becomes fluorescent. For your specific application you will need to calibrate this, but in principle it should work.

Try the simple Schiff base reaction of your amine groups with salicylic aldehyde - it`s very sensitive :) With UV-Vis detection.

@Gokulakrishnan Subramanian:

Hi Sir, in your back titration technique. Does your fe3o4  supernatant liquid a coloured solution, how were you able to titrate it cause my nanoparticle produces brown coloured supernatant (cobalt ferrite solution) once dispersed and centrifuged. It seems to be very hard to see end point during titration.