**3.2 9-[2-(3-Carboxy-9,10-diphenyl)anthryl]-6-hydroxy-3H-xanthen-3-one (DPAX)**

In terms of sensitivity issue a fluorescence probe is always better than probes which work on the basis of absorbance. Umezawa et.al developed a new sensitive and efficient fluorescence probe DPAX namely 9-[2-(3-carboxy-9,10-diphenyl) anthryl]-6-hydroxy-3H-xanthen-3-one, for the detection of <sup>1</sup> O2 by fusing a fluorescein moiety, with DPA which serve the characteristics of fluorescence due to fluorescein as well as a good <sup>1</sup> O2 trap for the presence of DPA [34]. DPAX and its derivatives show very low fluorescence intensity in aqueous solution but once binding with <sup>1</sup> O2 the corresponding endoperoxide (DPAX-ED) shows excellent fluorescence intensity with quantum yield in the range of 0.5–0.7 (**Figure 6**) [35]. The DPAX and its derivatives demonstrate excellent selectivity towards <sup>1</sup> O2 as the fluorescence intensity remains unchanged upon reaction with hydrogen peroxide, superoxide and nitric oxide [35]. DPAXs are suitable for application in neutral and basic aqueous solution but the fluorescence intensity is known to be decreased under acidic condition due to the protonation of phenoxide oxygen atom; thus are not suitable for application in acidic conditions [32]. The stability of the fluorescence intensity can be enhanced by incorporating electron withdrawing group like Cl, F at 2 and 7 position of the xanthenes moiety leading to generation of two

#### **Figure 6.**

*Reaction of 9-[2-(3-carboxy-9,10-diphenyl)anthryl]-6-hydroxy-3H-xanthen-3-ones (DPAXs) with 1 O2 to produce corresponding DPAX endoperoxides (DPAX-EPs) (adapted from [34]).*

derivatives of DPAX namely DPAX-2(Cl derivative) and DPAX-3(F-derivative). This structural change lowered the Pka value of the phenolic oxygen atom [34].

Absorption maxima (A), molar extinction coefficient (ε) and emission maxima (λem) are more or less same for DPAXs and DPAX-EPs but the quantum efficiencies of fluorescence are altered when DPAXs bind with <sup>1</sup> O2. The emission maximum of DPAX-2-EP is shifted to longer wavelength than the corresponding DPAX-2 compound in comparison to other DPAXs compounds (**Table 2**) [34].

DPAX-2 can be utilised as an efficient 1 O2 sensor in both basic and neutral medium. Umezawa et al. used two different 1 O2 generation system namely, MnO4 − / H2O2 and 3-(4-methyl-1-naphthyl)propionic acid endoperoxide (EP-1) which works in different pH values (10.5 and 7.4 respectively). In both the cases an increase in fluorescence intensity is established when this probe reacts with 1 O2 generation system (**Figure 7**).

To confirm the specificity of DPAX-2 towards <sup>1</sup> O2, fluorescence experiment was performed in presence of hydrogen peroxide, superoxide and nitric oxide but no appreciable change was observed for those species. These observations suggest the specificity of this probe for 1 O2 [34].


#### **Table 2.**

*Absorbance and fluorescence properties of DPAXs and DPAX-EPs (adopted from [36]).*

#### **Figure 7.**

*Emission spectra of DPAX-2 at 505 nm in reaction with 1 O2, generated from MnO4 − /H2O2 system. The reaction was performed in 0.1 M sodium phosphate buffer medium of pH 10.5 containing 0.1 mM EDTA at 25°C (adapted from [34]).*
