RADioBioT.or,ic:,\i. mf.c;h.\\ism at thi: cfi.ia'i.ar i.fa kl 



of the l)ulk of tlic specimen or tlic paramagnetism of elements such as 

 iron, manganese, and oxygen. 



(3) That in fa\()urable circumstances some informalic^n can be obtained 

 about the nature of the free radicals as well as about the numbers 



present. 



(4) The method offers a possibility in principle of examining the kinetics of 

 radical reactions in living systems at short intervals of time after irradia- 

 tion. 



The fraction of the energy of a resonant electromagnetic wave which is 

 removed by even a millimolar concentration of radicals in a 100 mg speci- 

 men, is exceedingly small, and the art of the spectroscopist consists in 

 achieving such stability of operation that when the incident and absorbed 

 waves are balanced in some kind of null disposition, the difference between 

 them stands out above the 'background noise' and may be amplified and 

 recorded. Instrumentation has been dealt with comprehensively by a 

 number of authors, including Gordy, Smith, and Trambarulo^^ and 

 Ingram^^. 



We have not considered our own electron spin resonance spectrometer, 

 which has been built in the laboratory at comparatively low cost, to be in any 

 way remarkable. It uses 3-2 cm waves (9375 Mc) generated by a 723 A/B 

 Klystron and employs a superheterodyne detection system. The main 

 Klystron is stabilized by an automatic frequency correction circuit in which 

 the sample cavity itself is used as a standard of reference. The local oscillator 

 operates with a frec}uency 60 Mc different from the main oscillator, and this 

 difference in frequency is amplified and passed to a phase sensitive detector 

 modulated to 180c/sec, and thence to a pen recorder. The magnet at present 

 in use is very small and inexpensive. A steady field of some 3300 gauss is 

 produced by permanent magnets behind the pole pieces. These are initially 

 magnetized in situ, and the magnetizing coils are then used to vary the field 

 by ± 300 gauss. The pole diameter is only 2 -5 in. and the gap 1 in., but by 

 the use of Rose shims the field variation on the central plane near the axis 

 is kept below 1 part in 10^ over a region 5 mm in diameter. The sample is 

 usually smaller than this. 



To the radiobiologist the important figure of merit of any equipment 

 would be the smallest number of radicals in a specimen of given size which 

 can be estimated with a given accuracy. Such figures are to be found in the 

 literature but they are not always very meaningful. Since the number of 

 spins which have contributed to a given signal is proportional to the 

 area of an absorption spectrum, sensitivity depends on the shape of the 

 spectrum, being greatest when most of the absorption lies within one or two 

 sharp peaks as is the case with the stable free radical diphenyl picrichydrazyl 

 (DPPH) and with irradiated glycine, and least for a broad structureless band 

 such as is obtained by irradiating seeds. Indeed, it is probable that many of 

 the radicals formed by the irradiation of biological materials escape detection 

 altogether. 



The performance of difTerent types of equipment may be compared on the 

 basis of the signal/noise ratio for a standard paramagnetic substance such as 

 a manganese salt which gives a six line spectrum, or DPPH which gives a 

 single sharp peak. Commoner el al.*^ quote their limit of detection as 04 



294 



