8 1. lODOACETATE AND lODOACET AMIDE 



The reaction rate constants for (A) and (C) are similar, and about 8.7 X 10"^ 

 liters/mole/sec, with an activation energy around 16-17 kcal/mole, corre- 

 sponding to an exchange rate of 0.0234 mole/mole/min. Reaction (B) is 

 about one-tenth and reaction (D) about one-seventeenth as rapid. A reduc- 

 tive reaction with iodoacetate: 



ICH^COO- + H+ + 2e- -> CH3COO- + I- 



may also occur under certain conditions and is the basis for the polaro- 

 graphic determination (Brdicka, 1936). The stability in tissue preparations 

 is of course much less, due to reaction with several components; thus, bro- 

 moacetate in ox blood at 37° is 23% reacted in 2 min, 50% in 35 min, and 

 67% in 64 min (Hansen, 1956 b). The methyl and ethyl esters of bromo- 

 acetate and iodoacetate hydrolyze very rapidly in solution (Bergmann and 

 Shimoni, 1953) and this should be taken into account whenever these 

 esters are used to facilitate penetration. 



lodoacetic acid which has been obtained commercially or which has been 

 standing for some time should be tested for free iodine with starch, and 

 repurified if a positive test is given. Some enzymes are very sensitive to 

 iodine (see Chapter II-5).* Lotspeich and Peters (1951) found that isocit- 

 rate dehydrogenase is not inhibited by recrystallized iodoacetate but is by 

 discolored samples, which could explain some of the earlier discrepancies, 

 lodoacetic acid may be readily recrystallized from carbon tetrachloride 

 (Hellstrom, 1931), or dissolved in benzene and precipitated with petroleum 

 ether. The preparations of the sodium salts of bromoacetate and iodoacetate 

 have been described by Drushel and Simpson (1917) and Goldberg (1943). 

 lodoacetamide may be prepared from chloroacetamide and sodium iodide 

 by the method of Anson (1940) and iodoacetamide-I^^^ obtained by the 

 method of Friedman and Rutenburg (1950 a). 



The substitution of a halogen atom on acetic acid increases the acid 

 strength by reason of the electronegative nature of these atoms (Table 1-1). 

 /^-Substitution in propionic acid has less effect because of the greater dis- 

 tance between the halogen and the carboxyl group. Thus lodoacetic and 

 bromoacetic acids are quite strong acids so that at physiological pH's there 

 is very little of the undissociated species, which may be of importance in 

 work with cells relatively impermeable to anions. The ratio (iodoacetate^)/ 

 (lodoacetic acid) at pH 7.4 is 18,600 and, at a total concentration of 1 mM, 

 (lodoacetic acid) = 5.4 X 10"^ mM. Of possible significance in permeabi- 



* Cross references to chapters, figures, equations, etc., in other volumes of this 

 treatise will appear in the following manner: Chapter 1-6; Fig. II-6-7; Eq. II-3-1; etc. 

 The Roman numeral refers to the volume; the first Arabic numeral to the chapter 

 number; and the second Arabic numeral to the figure or equation number, as the 

 case may be. 



