64 FIXATION 



cause underlying the continued use of potassium dichromate in 

 microtechnique (see p. 128). 



Acetic acid behaves quite differently. The ionization constant 

 being low (i-8 X io~^), the hydronium ion is not present in suffi- 

 cient abundance to cause denaturation, but is able to exert its 

 swelling effect. Indeed, this effect will be more readily produced if 

 proteins are not coagulated by denaturation. Collagen fibres are 

 swollen most by acids at pH 2-5,^^* and this is near the pH of 5% 

 acetic acid (see p. 134). Acids will tend to break the salt-links 

 between the chains; that is, the links binding together an amino- 

 group in one with a carboxyl group in another. The breakage will 

 expose hydrophil groups of atoms, and it is the entry of water 

 attracted by these groups that is the cause of swelling.^** 



Weak organic acids swell wool much more than mineral acids do 

 at the same pH. This is thought to be due to the action of the 

 undissociated part of the acid, which tends to attach itself to 

 amide groups through hydrogen bonds. The amide groups on 



r° 



HC^R 



I 

 NH • • 0=( 



=? 



HCR NH 



C=0 HCR 



Parts of tico protein chains held together by a hydrogen botid (...) between 

 two amide groups. The latter are indicated by bold lettering. 



neighbouring protein chains are supposed to associate with one 

 another through hydrogen bonds, and the breakage of these by un- 

 dissociated acid would permit swelling. The fact that acetic 

 acid has much more capacity to swell proteins than mineral acids 

 have is well shown in fig. i (p. 36). 



In addition to the hydronium ion and undissociated acid, there 

 is the acetate ion to consider. This has a separate effect in fixation, 

 which has been carefully studied by the American botanist 

 Zirkle.^^^ He found that all acetates, including hydrogen acetate 

 (acetic acid), tend to act similarly at the same pH (except that the 

 salts of heavy metals show the effects of the cation rather than that 

 of the anion). The acetate ion does not fix ordinary proteins; 

 indeed, it opposes the action of denaturing agents. *^^ It can react, 

 however, with nucleoproteins. Whether it does so or not depends 

 on whether the pH is on the less or the more acid side of pH 4-4 



