Highly selective visual distinction of pyrophosphate from other phosphate anions with 4-[(5-chloro-2-pyridyl)azo]-1,3-diaminobenzene in the presence of copper(II) ions

Pyrophosphate (PPi), as a biologically related phosphate anion, plays very important roles in organisms. Here, a highly selective visual method for distinction of PPi was made with commercial available 4-[(5-chloro-2-pyridyl)azo]-1,3-diaminobenzene (5-Cl-PADAB) in the presence of copper(II). The yellow solution of 5-Cl-PADAB exhibits strong absorption at 450.5 nm, and addition of Cu(II) results in a red solution with a new absorption band at 506.0 nm. Upon titration with PPi, the absorption band at 506.0 nm decreases with blueshift, while another new absorption band in the region from 562.0 nm to 750.0 nm appears which gradually splits into two peaks, and the color accordingly changes from red to cyan. Further addition of PPi, the new absorption peaks gradually disappear, and the mixture shows the absorption of 5-Cl-PADAB and recovers to yellow from cyan. This process is highly selective for PPi since other phosphate anions such as nucleotides cannot induce such spectral and color changes. With this method, the detection of PPi concentration in human urine was made with satisfactory results.