ABSTRACT
This chapter focuses on different analytical methods that are used to detect and measure Npn N’s. The possibility of continued degradation of dinucleotides in the extraction solution and the amount of other losses can be examined by including labeled dinucleotide standard for recovery determination. Trichloroacetic acid or perchloric acid have been the most common solutions for dinucleoside polyphosphate and mononucleotide extraction. Baker and Jacobson reported a novel extraction procedure which relies on the stability of dinucleoside polyphosphates in base. Dinucleoside polyphosphates, in contrast to mononucleotides, are resistant to phosphomonoesterases. The challenge of separating the dinucleotide polyphosphates from each other and from other similar compounds in cell extracts has made HPLC a logical choice due to its high separation efficiency and the relatively high sensitivity of detection with UV monitors. Coupling enzymes were usually required and the particular enzymes used depended on the particular reaction product measured.
