52 FLUORESCENCE SPECTROPHOTOMETRY 



fluorescence spectrum of the red alga Porphyra lacineata in a study of 

 energy transfer between photosynthetic pigments. This study was 

 extended to pigments of blue-green algae, purple bacteria and Chlo- 

 rella (Duysens, 1952). French and Young (1952) also measured 

 fluorescence spectral energy distribution curves of photosynthetic pig- 

 ments in live cells and in extracts. They have reviewed the work prior 

 to 1951 on absorption, action, and fluorescence spectroscopy of photo- 

 synthetic pigments ( Hollaender, Ed. ) . 



Although the experiments here described have been limited to the 

 study of photosynthetic pigments, the possible uses of these methods 

 in other fields of biology and biochemistry, particularly in the field of 

 luminescence, are evident. This paper will describe briefly the appa- 

 ratus constructed for automatically plotting the spectral energy dis- 

 tribution of weakly emitting light sources such as fluorescing leaves 

 and will illustrate the use of such measurements in studying the prop- 

 erties of pigments in solution and in living cells. Most of the material 

 has been taken from more detailed papers already published or now 

 in preparation; only a superficial survey will be attempted here. Some 

 progress reports in this field have been published (Carnegie Insti- 

 tution of Washington Year Books, 1948-1953). 



Apparatus 



Quantitative measurement of a fluorescence excitation spectrum re- 

 quires illumination of the sample with bright light in a narrow spec- 

 tral range and of known total energy as measured by a thermopile. 

 This incident light is converted in the sample, usually with a very 

 low efficiency, into fluorescent hght that is emitted in all directions. 

 To measure the spectral energy distribution of the weak fluorescent 

 light, as much as possible of it must be put through a monochromator 

 and the intensity of its various wavelengths determined. 



In order to make such measurements practical it is necessary to 

 start with a bright light source. Our high intensity source is a Bol type 

 of high-pressure mercury lamp made by the Huggins Laboratories in 

 Menlo Park, California. This is similar to the GE H6, but it dissipates 

 twice the power in a narrower capillary. This lamp gives a very high 

 intensity continuous spectrum as well as the mercury lines which are 

 very greatly broadened. Even in the red part of the spectrum this 



