Figure 1

The enhanced catalytic horseradish peroxidase cycle (adapted from [25]). Luminol (L) is used as substrate for the light generating process. A di-aza-quinone (AQ) is formed as intermediate. This in turn is oxidised by hydrogen peroxide (H₂O₂) to form an excited state of aminophtalate (AP*). The final step is the emission of blue (420 nm) light (h·ν) when the excited AP* returns to its ground state. Luminol can work as a substrate of the horseradish peroxidase (HRP). However, for analytical purposes an intermediate aromatic hydrogen donor (AH) is added. This enhancer serves as primary substrate for the HRP and its radical (A^·) subducts electrons from luminol (L) and thus forms the radical form L^·. AQ is formed by electron transfer between two L^·. The HRP compound I-state (CMP I) is sensitive to excess of H₂O₂ [33] and can undergo a peroxide inactivation (so-called 'suicide reaction'; grey arrows) when the concentration of H₂O₂ added to start the light emitting reaction is too high (compare Fig. 1.4 in additional file 1).