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Isolation, purification, and quantitation of cyclic AMP and cyclic GMP in canine bile

1977; Elsevier BV; Volume: 81; Issue: 1 Linguagem: Inglês

10.1016/0003-2697(77)90605-4

1096-0309

Mary J. Ruwart, Donald L. Kaminski,

Adenosine and Purinergic Signaling

A method has been established to purify and separate cyclic AMP and cyclic GMP from canine bile. Perchloric acid extracts are treated with Dowex 1 resin. This batch processing binds the nucleotides completely, but very little biliary pigment (less than 5%). The Dowex 1 resin is then layered onto a Dowex 50 column, and nucleotides are eluted with 1 n (cyclic AMP) and 4 n formic acid (cyclic GMP). Bile pigments are retained on the Dowex 50, Samples can be concentrated in vacuo. Cyclic nucleotides are then measured via radioimmunoassay. Isolates should be treated with phosphodiesterase for consistent results. This method can be used with small samples, resin volumes, and clution volumes. Cyclic AMP and cyclic GMP are both resolved. Canine bile averages 8097 ± 1118 pmol/ml of cyclic AMP and less than 0.8 pmol/ml of cyclic GMP.