130 P. George, J. Beetlestone and J. S. Griffith 



high- and low-spin forms calculated in Section IV. At 540 m/< the difference 

 in e^^ is about 4, which gives an increase of between 0-05 and 0-07, i.e., 

 between 5 and 7 %. However, in order to obtain the individual spectra of the 

 high- and low-spin forms from the difference spectrum, an independent 

 determination of the fractions present at the two temperatures is required. 

 This can be seen from the equations, 



fg = agSj -}- (1 - a5)£^ (13) 



£35 = aaaCj + (1 - a35)e;, (14) 



where £5 and £35 are the extinction coefficients of the hydroxide at 5° and 

 35°, a5 and cc^^ are the fractions of the low-spin form at the two temperatures, 

 and £f^ and £j are the extinction coefficients of the high- and low-spin forms. 

 Since £5 is known and £35 can be obtained from the difference spectrum, 

 provided cc^ and cc^^ can be determined, jUj^ and jUi can be evaluated from the 

 equations rearranged in the form, 



^3b'^5 ~ ^5*^35 



"■35 



(15) 



€5(1 - ^35) - ^35(1 - 0^5) .... 



El = (16) 



0^5 - ^35 



The variation of a with temperature has been obtained in the following 

 way. Using a sensitive Gouy balance, constructed from a Varian electro- 

 magnet V4004 and a Sartorius Microbalance MPR 5 II, and equipped with 

 a coaxial glass thermostat surrounding the sample tube and suspension fibre, 

 the change in Aw was measured as a function of temperature over the range 

 1° to 30°C for the fluoride, cyanide and hydroxide derivatives of ferrimyo- 

 globin. Calibration with nickel chloride solution enabled these changes in 

 Aw to be converted into changes in molar susceptibility, Xu- The value of 

 Xu obtained by Theorell and Ehrenberg (1951) for the three derivatives at 

 20°C were adopted, namely, 14,790, 2,340 and 11,040 x 10-^ c.g.s. units 

 respectively, and hence values of^Xu o^^^ the temperature range were obtained. 

 The variation of Xu for the fluoride and cyanide derivatives was found to 

 follow very closely the simple Curie law, x = constant/r. The magnitude of 

 the change is illustrated by the following data: from 20° to 1°, Xu for the 

 fluoride and cyanide derivatives increases by 1,046 x 10~^ and 82 x 10~^ 

 c.g.s. units respectively. On the other hand, Xu for the hydroxide, although 

 it has a high value approaching that of the fluoride, only increases by 

 145 X 10~^ c.g.s. units for the same decrease in temperature. As a conse- 

 quence, the values of Xm do not follow the Curie Law, and the type of devia- 

 tion is just what would be expected if, on lowering the temperature, the 

 fraction of the high-spin form decreases. 



