7-Ethoxyresorufin (Resorufin ethyl ether) (7-ER) is a fluorometric substrate and competitive inhibitor of cytochrome P450, particularly CYP1A1.

7-ER inhibited responses to NO and nitrergic nerve stimulation through generation of superoxide radicals. A 7-ER sensitive P450 system may be involved in the bioactivation of GTN and SNP in rat aortic rings, but not in rabbit aorta or rat anococcygeus muscles.

Instructions
I. Solution preparation
1. Stock solution preparation: 7-Ethoxyresorufin is dissolved in DMSO or ethanol and prepared to 1–10 mM (adjusted according to experimental needs).
2. Working concentration: Usually 0.5–5 µM. (Adjusted according to experimental needs)
Notes:
1) Powder: Store dry and away from light at -20°C.
2) Stock solution: Store away from light at -20°C, and can be stored at 4°C for a short period of time to avoid repeated freezing and thawing.
II. CYP1A1/CYP1A2 enzyme activity detection
Operation steps
1. Sample preparation:
Use liver microsomes or cultured cells as samples. Cells need to be pretreated to induce the expression of CYP1A1 or CYP1A2 (such as treatment with benzo[a]pyrene or 3-methylcholanthracene).
2. Reaction system:
1) Prepare reaction buffer (e.g. 50 mM Tris-HCl, pH 7.4).
2) Dilute the stock solution of 7-Ethoxyresorufin to a working concentration (e.g. 2 µM) and add to the buffer.
3) Add NADPH generating system (e.g. NADPH or NADPH regenerating system) to start the reaction.
3. Reaction conditions:
1) Incubate at 37°C for 10–30 minutes (the specific time is optimized according to the experiment).
2) Stop the reaction: The reaction can be terminated by adding an equal volume of cold ethanol or methanol.
3. Fluorescence detection:
Resorufin (deethoxylated product) generated by the reaction emits strong green fluorescence:
Excitation wavelength: ~530 nm
Emission wavelength: ~585 nm
Measure the fluorescence signal using a fluorescence spectrophotometer or microplate reader.