Roles of Glutathione (GSH) in Dopamine (DA) Oxidation Studied by Improved Tandem HPLC Plus ESI-MS

Abstract

In this study, new procedure with improved tandem HPLC plus ESI-MS was utilized to decipher the protective role of glutathione (GSH) against dopamine (DA) oxidation. We demonstrated that auto-oxidation of DA could produce aminochrome (AM, a cyclized DA quinone), which could be effectively abrogated by reductants, especially by GSH. Furthermore GSH was demonstrated to be able to conjugate with AM to form various conjugates via condensation reactions without enzymatic catalysis. The GSH-AM conjugates tend to aggregate, possibly mediated by conjugated AM structures, but could be inhibited by GSH. We hypothesized that proteins conjugated by AM might facilitate Lewy body formation of Parkinson's disease (PD) in dopaminergic neurons via similar polymerization. We proposed that GSH could protect dopaminergic neurons against DA-induced toxicity via various mechanisms. The imbalance between DA oxidation and GSH protective capacity could be a key factor contributing to PD. Strategies to use GSH analogues, GSH inducers or to control DA oxidation might work to control PD onset and development.

Glutathione Conjugates with Dopamine-Derived Quinones to Form Reactive or Non-Reactive Glutathione-Conjugates

“…The Improved HPLC A Procedure [20] The HPLC analysis was performed using AKTA purifier HPLC system (Amersham, Sweden) with a UV detector (detection wave length was set as 280 nm) and a reversedphase column (DENALI TM Monomeric RPC C18 4.6 mm 9 250 mm, 120Å Pore Size VYDAC 238DE TM Series, Grace Vydac, U.S.A.) and analyzed under the control of UNICON 4.11 program. Our HPLC A procedure used two elution buffer systems (elution buffer A and elution buffer B).…”

“…Furthermore GSH transferase could catalyze GSH conjugation with AM to form 4-S-GSH-5,6-dihydroxyindoline [19]. In our recent study using an improved tandem HPLC procedure coupled with ESI-MS, we found that GSH could actually react with AM to form various GSH conjugates without any enzymatic catalysis [20]. We have also shown in a separate study that DA-derived quinones are associated with cellular toxicity induced by mutant alpha-synuclein which is strongly associated with PD [21].…”

Analysis of endogenous glutathione‐adducts and their metabolites

“…These findings suggested autoxidation of catechols to quinones had occurred in a similar manner to that described above for catechol estrogens and that GST-mediated GSH addition had occurred at C-5 to give a dopamine–GSH-adduct. Recently, off-line LC separation followed by MS analysis was employed to show that GSH-adducts were formed from the reaction of GSH with aminochrome, a dopamine-derived quinone (Zhou and Lim, 2009). …”

Separation of intermediates of iron-catalyzed dopamine oxidation reactions using reversed-phase ion-pairing chromatography coupled in tandem with UV–visible and ESI-MS detections

“…This observation can be rationalized on the basis that H 2 O 2 is absent in the solution. Among the identifications of the intermediates/products shown in Figures 2 and 3, detection of 6-OHDA is the most significant given its neurotoxicity and the fact that it has not been separated and identified by other HPLC methods [6,8,14–16]. We believe that in previous studies either 6-OHDA was completely oxidized or the sensitivity of the methods used was not sufficiently high to detect the trace amount of 6-OHDA.…”