Mass spectrometric investigation of concentration-dependent effect of curcumin and oxidative stress on intracellular glutathione levels

Herein, we investigated the correlation between curcumin and glutathione (GSH) levels in mammalian cells using gold nanoparticles (AuNPs) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). GSH exists in high concentration in the cytosol and acts as a major antioxidant and reducing agent in organisms. Previous studies showed that curcumin, a well-known antioxidant with anti-inflammatory, anti-proliferative, and anti-carcinogenic activities, affects GSH levels in mammalian cells. However, the correlation between their levels remains controversial and has not yet been completely elucidated. This study used our recent strategy of GSH quantification, where GSH in cell lysate is captured on maleimide groups of AuNPs and analyzed using MALDI-TOF MS with isotopomer GSH (GSH*)–conjugated AuNPs as an internal standard. The comparison between GSH and GSH* relative intensities allows the quantitation of GSH in cells. In this way, GSH levels in mammalian cells were investigated after incubation with curcumin at various concentrations with or without oxidative stress. We observed that intracellular GSH levels were affected by curcumin in a concentration-dependent manner with oxidative stress; GSH levels decrease at a lower curcumin concentration, which can be recovered at increased curcumin concentrations. We also found that the GSH level increased at all curcumin concentrations after a certain incubation time. We believe our strategy can be commonly used to determine GSH levels in cells that are treated differently with various exogenous stimulants like reactive oxygen species, biofunctional natural products, and drug candidates.