HPLC methods applying the reversed phase technique, where the eluating buffer solution is supplemented by a large hydrophobic cationic, typically a tetrabutylammonium (TBA) compound, were probably first used for the separation of nucleotides by Juengling and Kammermeier (1980), who applied two separate isocreatic runs in different conditions to resolve Cr, PCr, and adenine nucleotides.