Figure 6.
Analysis of dansyl–GSH Conjugates and M1 by LC–MS/MS and UPLC–UV Detection. Sunitinib (25 μM) was incubated with pooled human liver microsomes (1 mg/mL) in the presence of dGSH (1 mM) and an NADPH-regenerating system for 30 min. Control incubations were without NADPH, without dGSH, or without substrate (blank sample). Each reaction condition was performed in triplicate. Samples were analyzed by LC–MS/MS utilizing MRM for the (A) quinoneimine–dGSH conjugate of sunitinib: m/z 935 > 862, (B) M1–dGSH conjugate: m/z 911 > 371, and sunitinib–dGSH conjugate(s): m/z 939 > 399. (D) Relative levels of M1 measured by UPLC–UV absorbance at 430 nm. Levels of M1 were compared from incubations with and without dGSH by unpaired two-tailed t test (n = 2-3).