[Char_lotte]

For this determination I used a Mojonnier-type fat extraction flask, but while proceeding the standard procedure, a problem occured.

After weighing 3 g of cheese in a Mojonnier flask, I added 10 mL of HCl and heated it.
After cooling the flask, I added 10 mL of ethanol and mixed.
Here is the problem: After adding 25 mL of diethyl ether, a white clump appeared in the flask.
I searched for a few hours, but couldn't find out why this white clump appeared.
My guts say it has something to do with the casein in the cheese, but I can't find the answer.

I proceeded the fat extraction, but the results weren't correct.

Could anybody help please?

[Babcock_Hall]

I can't do any better than to make a guess about this, but I would not expect protein to be soluble in ether.  I would think that it might be difficult to extract all of the fat away from the solid without doing something such as centrifugation or possibly an additional extraction, but again I am not speaking from experience.  Are you working from a protocol that is specific for cheese?

[Char_lotte]

We used a protocol specific for cheese.
My colleage told me that there wasn't suppost to be a clump.
So I repeated the experiment, but the same white clump appeared.
I also did a blanco test. This time, there was no white clump.
So I suppose something in the cheese reacts with the diethyl ether to form this clump.

We don't have a centrigufe for Mojonnier flasks in our lab.

[Arkcon]

From the name of the instrument, you appear to be following ISO 1735:2004, or something similar.  The exact protocol is only for people who pay for it, however, I did find something of the technique by Googleing. 

The procedure I saw used a mixture of diethyl ether and petroleum ether, but I don't know if that will help solubilize proteins. 

It doesn't seem likely this is the problem, but a 3 gram sample is for cheese you expect to have no more than 30% fat.  You'd use a 1 gram sample for fattier cheeses.  You may want to try 1g, 2g, samples, just to see if this is the problem, even if you can't report the result from a less than 3 gram sample.

Just to be sure you're following all of the procedure, are you thoroughly grating the cheese sample finely?  If its too soft, mincing it may be enough, but you could always freeze it to make it something you can grate.  Are you thoroughly drying to remove moisture before you add ethanol.  Obviously, you're adding water with the HCl, but water intimately mixed with the proteins may give them something to clump with, and then you get a clump, instead of something that disperses in the ether, even if they won't dissolve in ether.

[Arkcon]

This reference I found is to a competing method, and I know you can't just switch methods.  However, you can see from the three methods specified, how different solvents are used, compared to yours.

http://www.dionex.com/en-us/webdocs/4491-AN340-ASE-Fat-Milk-Products-28Apr2011-LPN1185-01.pdf

[Furanone]

So the clump floats in the diethyl ether phase when you decant into weighing pan? This means its density must be much lower than water, and should not be protein material or lactose. I am thinking it might be clumps of undissolved fat that has not enough other things (water, protein) to not weigh the clump down in the aqueous phase but enough to give you an overestimation in your fat content results. Alternatively, if the clumps are remaining in the aqueous phase then it is likely undissolved protein material with some bound fat not enough to float in ether phase, but enough to give you an underestimation in your fat content results.

Also, I am not sure why you are using HCl. I seem to remember many, many years ago when I did Mojonnier using an NH4OH solution to help dissolve the caseins to where there was no longer any turbidity, and this was at a commercial lab where we did raw milk, cheese, yogurt, etc.

Even in the USDA testing method for Mojonnier, they use ammonium hydroxide not HCl:

http://www.fmmaseattle.com/lab/fat.htm

[Darryl1]

I am speculating here.  It may be unbroken down proteins.  Dairy proteins can be very complex and may not completely break down to amino acids with just HCl.  Could you add a small amount of protease into it and see if it still happens?

-d