Dibenzylfluorescein  (Synonyms: DBF)

Dibenzylfluorescein (DBF) is a fluorogenic probe (Fluoresecent dye) that acts as a substrate for specific cytochrome P450 (CYP) isoforms, including CYP3A4, CYP2C8, CYP2C9, CYP2C19, and aromatase (CYP19). Dibenzylfluorescein is typically used near its K_m value of 0.87-1.9 µM (Ex=485nm，Em=535nm). Dibenzylfluorescein is used to detect changes in CYP catalytic activity caused by drugs or disease^[1][2][3][4].

The protocol of P450-catalyzed metabolism of Dibenzylfluorescein and effect of base^[3]:
Reaction Process: Dibenzylfluorescein is dealkylated by P450 to form a fluorescein benzyl ester, which is further hydrolyzed to fluorescein by NaOH (if present). Addition of 2 M NaOH causes also decomposition of Dibenzylfluorescein to fluorescein benzyl ether.
1. Incubation mixtures for CYP2C19 enzyme-catalyzed samples each 150 μL contains 0.1 M Tris-HCl buffer (pH 7.4), 10 μM Dibenzylfluorescein, 15 pmol of CYP2C19 enzyme, and 50 μL of NADPH-regenerating system.NADPH-regenerating system contains 1.13 mM NADP, 12.5 mM isocitric acid, 56.33 mM KCl, 187.5 mM Tris-HCl, pH 7.4, 12.5 mM MgCl2, 0.0125 mM MnCl2, and 0.075 U/ml isocitrate dehydrogenase.
2. The samples were incubated for 30-60 min at 37°C. The reactions were terminated by rapid cooling to 4°C and after centrifugation, the supernatants were analyzed by LC-MS.
3. Pure Dibenzylfluorescein, fluorescein benzyl ester, fluorescein benzyl ether, and fluorescein (all 10 μM) were used as standards and were analyzed in the absence and presence of 2 M NaOH.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

512.55

C₃₄H₂₄O₅

97744-44-0

Solid

Yellow to orange

O=C1C=C2OC3=CC(OCC4=CC=CC=C4)=CC=C3C(C5=CC=CC=C5C(OCC6=CC=CC=C6)=O)=C2C=C1

Room temperature in continental US; may vary elsewhere.

4°C, protect from light

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light)

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (protect from light). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

• [1]. Stresser DM., et al. Substrate-dependent modulation of CYP3A4 catalytic activity: Analysis of 27 test compounds with four fluorometric substrates. Drug Metabolism and Disposition 28(12), 1440-1448 (2000).  [Content Brief]

  [2]. Donato MT., et al. Fluorescence-based assays for screening nine cytochrome P450 (P450) activities in intact cells expressing individual human P450 enzymes. Drug Metab. Dispos. 32(7), 699-706 (2004).  [Content Brief]

  [3]. Salminen KA, et al. Simple, direct, and informative method for the assessment of CYP2C19 enzyme inactivation kinetics. Drug Metabolism and Disposition 39(3), 412-418 (2011).  [Content Brief]

  [4]. Moutinho D, et al. Altered human CYP3A4 activity caused by Antley-Bixler syndrome-related variants of NADPH-cytochrome P450 oxidoreductase measured in a robust in vitro system. Drug Metabolism and Disposition 40(4), 754-760 (2012).  [Content Brief]