PAPER CHUOMATOGKAPHY 8(')I 



matography, became recojiiiiized as a pi-act ical and iii\';ilual)le analytical 

 tool for the biochemist. 



In esseiKie, the procedure is carried out by the application o{ a small 

 drop of test solution a short distance t'loni one end of a strip of filter paper. 

 After the drop has dried, this end of the strip is placed in an appropriate 

 solvent so that the latter moves past the spot by capillary action and 

 along the paper. This results in a differential movement of the compo- 

 nents of the test solution along the paper. The solvent usually consists 

 of a stationary aqueous phase which has a strong affinity for the filter 

 paper, and an organic or mobile phase which tends to move along the 

 paper. As the mixed solvent flows through the section of paper contain- 

 ing the test substances, the latter are distributed between the organic 

 phase, which is moving rapidly ahead, and the stationary aqueous phase. 

 After completion of the separation, the individual zones or spots are iden- 

 tified by color reactions, radioactivity, or other methods. 



The potentialities of paper chromatography, especially when used in 

 conjunction with radioisotopes, are so vast that the worker in biochem- 

 istry can ill afford not to have these techniques at hand. It is first noted 

 that, as a separation procedure, paper chromatography, on account of its 

 countercurrent nature, is highly efficient as compared with batch proce- 

 dures. The sensitivity is high, the detection of the spots usually being 

 the limiting factor; this is precisely where radioisotope technicjues may 

 offer considerable advantage. Another important feature is that the 

 pi'ocedure is simple and rapid, requiring no equipment except filter paper, 

 chemicals, and glassware, all of which are to be found in the usual lab- 

 oratory. This is in contrast to other chemical separation procedures such 

 as fractional crystallization or distillation. Also the latter procedures 

 can be used only with substances that can be crystaUized or distilled, and 

 they often reciuire elevated temperatures, which may degrade the test 

 substance. Large numbers of samples can be analyzed by paper chro- 

 matography even with fimited facilities. Williams (9), for example, 

 reports that his laboratory averaged about 700 analyses each working day 

 for over a year, an output that would have been prohibitive by other 

 methods. 



The primary usefulness of paper chromatography lies in the following: 

 (a) separation of mixtures into their constituents, (b) demonstration of 

 homogeneity of chemical substances, (c) demonstration of identity of sub- 

 stances, and (d) quantitative or qualitative estimation of one or more sub- 

 stances present in a mixture. The method has become of particular value 

 because a great many important biochemical compounds occui- in nature 

 as complex mixtures of substances of similar properties and structure and 

 are therefore most difficult to resolve by other means. 



The general references may be consulted for a discussion of the theory 



