The effect of organic solvents on the proteins depends on the type of protein  it self , As you know some of them dissolved and other not  ( molecular weight and polarity ) .what is the type of your protein ?

Thank you Alaa J Mahrath 

Ubiquitin,myglobin,BSA,Transferrine and cytochrome are my proteins.

Gedda, what exactly are you trying to do?

Not every organic solvent is immiscible with water. Ubiquitin, myglobin, BSA, transferrine and cytochrome can all be dissolved in water and then mixed with organic solvents if that is what you need to do.

You list this question here under Proteomics, it sounds like you are interested in some standard proteins and I would almost suspect that you want to do an MS experiment with them. But then you state that you are worried about them being denatured. For most MS experiments, this would not matter.

Yet,  you could try to do activity studies with these proteins and then denaturation should be a concern.

Could you please be a bit more specific what you are trying to do?

Dear sonja Hess thank you so much .

I'm trying to concentrate and separation of  proteins biological samples  using  nano particles dispersion  liquid liquid micro extraction.

Here i will disperse nano particles in organic solvents such as Toluene  and proteins in water.

So i will trying to extract from aqueous phase to organic phase. Here i will use organic sovlents 50uL to 100uL and 1-2mL of aqueous proteins sample.

I see, and why are you concerned about whether the proteins are denatured or not? If the goal is to concentrate and separate, it should not matter, unless it means that they precipitate. But denaturation is not the same as precipitation.

Yes my goal was concentrate and separation. But i'm afraid that if the  organic solvents  denature the proteins  its may destroyed the protein structure . 

According your suggestion i no need to worry for  denature of protein. However can you explain me  these water immiscible(like toluene ) solvents can influence on proteins structure or not.

Yes, organic solvents will influence the tertiary (and secondary) structure of proteins. I have once studied the effect of hexafluorisopropanol (HFIP), which is miscible with water in low and high concentrations (but inbetween is a miscibility gap).

HFIP reportedly enhances the alpha helicity of the secondary structure, and the

tertiary structure of proteins is disordered by this solvent.

I have not used toluene and I do not know what it does to the alpha helicity of the secondary structure, but I would assume it to disorder the tertiary structure as well. As long as it does not precipitate, that should not be a problem, unless you want to do biological assays with it afterwards.

Hope this answers your question