Coal waste-derived synthesis of yellow oxidized graphene quantum dots with highly specific superoxide dismutase activity: characterization, kinetics, and biological studies

The disintegration of coal-based precursors for the scalable production of nanozymes relies on the fate of solvothermal pyrolysis. Herein, we report a novel economic and scalable strategy to fabricate yellow luminescent graphene quantum dots (YGQDs) by remediating unburnt coal waste (CW). The YGQDs (size: 7-8 nm; M.W: 3157.9 Da) were produced using in situ "anion-radical" assisted bond cleavage in water (within 8 h; at 121 degrees C) with yields of similar to 87%. The presence of exposed surface and edge groups, such as COOH, C-O-C, and O-H, as structural defects accounted for its high fluorescence with epsilon(max) similar to 530 nm at pH 7. Besides, these defects also acted as radical stabilizers, demonstrating prominent anti-oxidative activity of similar to 4.5-fold higher than standard ascorbic acid (AA). In addition, the YGQDs showed high biocompatibility towards mammalian cells, with 500 mu M of treatment dose showing <15% cell death. The YGQDs demonstrated specific superoxide dismutase (SOD) activity wherein 15 mu M YGQDs equalled the activity of 1-unit biological SOD (bSOD), measured using the pyrogallol assay. The K-m for YGQDs was similar to 10-fold higher than that for bSOD. However, the YGQDs retained their SOD activity in harsh conditions like high temperatures or denaturing reactions, where the activity of bSOD is completely lost. The binding affinity of YGQDs for superoxide ions, measured from isothermal calorimetry (ITC) studies, was only 10-fold lower than that of bSOD (K-d of 586 nM vs. 57.3 nM). Further, the pre-treatment of YGQDs (similar to 10-25 mu M) increased the cell survivability to >75-90% in three cell lines during ROS-mediated cell death, with the highest survivability being shown for C6-cells. Next, the ROS-induced apoptosis in C6-cells (model for neurodegenerative diseases study), wherein YGQDs uptake was confirmed by confocal microscopy, showed similar to 5-fold apoptosis alleviation with only 5 mu M pretreatment. The YGQDs also restored the expression of pro-inflammatory Th1 cytokines (TNF-alpha, IFN-gamma, IL-6) and anti-inflammatory Th2 cytokines (IL-10) to their basal levels, with a net >3-fold change observed. This further explains the molecular mechanism for the antioxidant property of YGQDs. The high specific SOD activity associated with YGQDs may provide the cheapest alternative source for producing large-scale SOD-based nanozymes that can treat various oxidative stress-linked disorders/diseases.