L. H. GRAY 



ml. of a 10"^ molar aqueous solution of Mn + +, i.e. 2-4 x 10^^ spins. Conger 

 and Randolph*^ quote 3 x 10^^ radicals of DPPH as giving the 'minimum 

 detectable signal'. The corresponding figure quoted by Hirshon and 

 FraenkeH- is 10^^ radicals. Our own equipment gives about 10:1 signal/ 

 noise ratio with 10^^ radicals of DPPH. Figures of this kind can at present 

 be taken only as rough order of magnitude estimates. The theoretical limit 

 of sensitivity has been computed by Fraenkel*^ as 10^° spins (2 • 10"^'' moles) 

 at a concentration of 10^^ to 10"^" moles/1. 



Perhaps the best indication of the useful levels of sensitivity so far achieved 

 is given by the minimum dose for which quantitative results have been 

 reported by different workers, viz. barley seeds 50 kr*^, wheat germ (150 

 mg sample) 10 kr*^ and glycine 2 kr*^. The many problems associated 

 ^vith the use of quantitative electron spin resonance spectroscopy for evaluat- 

 ing radical yields in irradiated materials have been discussed informatively 

 by Zimmer, Ehrenberg, and Ehrenberg**, Zimmer*^, and Livingston^^. 



As mentioned above, radicals only show hyperfine structure when the 

 unpaired electron is associated to some degree with nuclei which have a 

 magnetic moment. As far as biological materials are concerned, this excludes 

 i-C, ^•^O, ^^P, and ^-S, and leaves only ^H and ^^N among the major consti- 

 tuents of tissue. However, the nuclei -H, ^^C (0-5 per cent abundance), 

 ^"O, and ^^S (1 per cent abundance) have magnetic moment, and these 

 isotopes may prove useful. Smaller and Avery ^'^ have used the differences 

 between the spectra which result from irradiation of normal and of deuterated 

 yeast to good effect in analysing radicals formed in the water and in the 

 organic constituents of the cell. It should be noted, however, that while 

 biological systems are not in general greatly disturbed by growth in up to 

 20 per cent heavy water, a sharp toxicity often sets in at about the 40 per 

 cent level'*^. It is also to be noted that Laser has reported*^ that when E. coli 

 B. is irradiated in DoO the ratio of aerobic to anaerobic sensitivity is increased 

 from the usual 2-5 : 1 to about 8:1. 



It is in some ways unfortunate that oxygen, which is a paramagnetic 

 molecule and a bi-radical, does not give an observable spectrum except in 

 the gas phase at low pressures^*^' ^^' ^^. The coupling of the electron spin 

 to the molecular axis of oxygen produces a complicated set of magnetic 

 energy levels that depend on the rotational state of the molecule. Since the 

 perturbations caused by collisions are not sufficiently strong to break down 

 the coupHng but do broaden the rotational levels, no spectrum is observed at 

 high pressures or in solution*^. 



One of the most exciting things about the communication from Gordy 

 et al. in 1955, to which reference has been made above^", was that all the 

 amino acids which had been examined gave rise on examination to para- 

 magnetic centres with distinguishable spectra. The second was that proteins 

 did not show the expected superposition of spectra corresponding to the 

 component amino acids. Some of the sulphur-containing proteins showed 

 no peaks corresponding to many of the constituent amino acids, but instead, 

 a rather simple spectrum closely resembling that of irradiated cystine, 

 suggesting that though the atoms originally ionized and excited must have 

 been distributed in a random manner among all the amino acids, the free 

 radicals appear preferentially in the cystine. This internal 'protection' 



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