Isolation of the acid-soluble nucleotides of liver.

In the course of studies on the acid-soluble phosphorus compounds of liver and kidney, it became evident that the fraction comprising the nucleotides represented a complex mixture (1). In an attempt to isolate the individual nucleotides, methods designed to minimize the breakdown of labile phosphorus compounds were employed. The isolation of adenylic acid and of a material representing a mixture of adenosine diand triphosphate from rabbit and dog liver is reported in the present communication. The following steps were taken in the isolation of the products. The organs were removed quickly from narcotized animals and put into a dry ice-ether mixture. In the trichloroacetic acid ext,ract the nucleotides together with glutathione and an unknolr-n nitrogen-free phosphorus compound were precipitated \l;ith mercuric acetate. After removal of the mbrcury, the barium sa1t.s were prepared and divided into a soluble fraction, containing adenylic acid and glut.athione, and an insoluble fraction, containing adenosine triphosphate and the bulk of the adenosine diphosphate. From the soluble fraction free adenylic acid was isolated after a preliminary precipitation with phosphotungstic acid to separate it from glutathione. Adenosine triphosphate and the bulk of the adenosine diphosphate, present in the insoluble fraction of the barium salts, were isolated after repeated fractionation as silver salts from 0.2 s nit.ric acid to remove the nitrogen-free cont,aminant . The preparations were analyzed for nitrogen by a Kjeldahl procedure, for adenine (2), and for easily hydrolyzable and total P (3). Determination of the total nucleotide phosphorus was performed with rattlesnake venom as a specific nucleotidase according t.o Hitchings (4). The content of adenosine triphosphate was assayed by three methods, (a) rattlesnake venom (4), (b) myosin (5, G), and (c) washed lobster muscle (7).