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Fig. 4
Isopimpinellin and disodium cromoglycate do not have antioxidant activity. (A) Isopimpinellin is a structural isomer of khellin, from which a series of functional benzopyrone analogues were designed, including disodium cromoglycate and nedocromil. (B,C) FRAP assay, performed at 37°C for 5min. Graphs show the rate of increase in absorbance at 593nm, corrected for reagent blank readings, at (B) 100µM and (C) 400µM of each compound, as indicated. (D) The in vivo activity of the compounds tested (expressed as the percentage decrease in the mean neutrophil numbers at the wound at 12hpi), in comparison to the vehicle control (refers to data from Figs 3A, 6A and Fig. S3), plotted against the chemical reducing ability value (refers to data from Table S1) (linear regression analysis; R2=0.8960; P=0.535). (E) In vivo antioxidation assay. Larvae were pretreated with DMSO, isopimpinellin or DPI for 2h and pentafluorobenzenesulfonyl fluorescein for 1h prior to tail-fin injury. Imaging commenced 5min post-injury (mpi) and mean fluorescence intensity at the injury site was measured at 5 and 20mpi. Mean fluorescent intensity at 20mpi is expressed as a percentage of the mean fluorescent intensity at 5mpi. This was significantly reduced by DPI but isopimpinellin had no effect (one-way ANOVA with Dunnett′s multiple-comparison post-test; ****P<0.0001; ns, non-significant; n>15, performed as two independent experiments). Illustrative images are shown in F. Scale bar: 50 µm.