I was extracting total plant protein from a cotton leaf sample following TCA/Acetone precipitation method given in the paper linked below. After taking the aqueous phase along with the upper phenol phase I added 80% methanol containing 0.1M ammonium acetate and kept for overnight stand at -200C. In the morning before centrifugation dense whitish substances were seen trying to settle down. I thought after centrifugation pellet will definitely form. But after centrifugation I found all of them had disappeared, perhaps dispersed. Even after the 2nd centrifugation no pellet was formed. What was the problem? Please help. I have attached a file showing the phenolic separation stage what I had got. I believe the phase in between the lower phenol phase and the aqueous phase contains the major amount of protein. Please suggest. This is the first time I am using this protocol.  

http://onlinelibrary.wiley.com/doi/10.1002/elps.200500722/full