C60 is a carbon nanomaterial that is widely used in chemical industries ranging from electronics to medicine. This widespread use means that humans will very likely be exposed to the substance. When a new material becomes available, it is always wise to study the possible effects that it may have on human health to evaluate the risks associated with the benefit. C60 is a nonpolar molecule that can lodge in biological membranes due to hydrophobic interactions. Mitochondria can be used to study the effects of C60 on health because, having two membranes, they are particularly susceptible to membrane integration by C60. Mitochondria are also used as indicators of toxicity because they produce the energy currency of the cell, adenosine triphosphate (ATP), and reactive oxygen species (ROS) which are damaging side products of ATP production. Mitochondria use oxygen in a membrane-dependent process involving electron transfer to produce ATP. In this study, beef heart mitochondria were used to study the effects of C60. We hypothesized that because C60 is able to lodge in membranes, it will have a dose dependent adverse effect on mitochondrial electron transfer activity. When electron transfer activity is disrupted, ROS production can increase. Thus, we also predicted that ROS production could increase in mitochondria that are exposed to C60. Mitochondria were isolated from beef heart and exposed to 2 to 35 ppm C60 for 0 to 30 minutes at 25° C in a HEPES buffer at pH 7.4. Electron transfer was measured by oxygen consumption using a Clark electrode and succinate as the electron source. This portion of the electron transfer chain contains three enzyme complexes (II, III and IV, known as succinoxidase) which must interact in the membrane to allow electron transfer. As an indicator of ROS, hydrogen peroxide (H2O2 ) production was monitored by reacting it with amplex red and measuring the fluorescent products with a Synergy II plate reader. A 66% inhibition of succinoxidase activity between the control and the 20 ppm treated mitochondria was observed. An increase in H2O2 production was also observed. The results show that C60 had a dose-dependent adverse effect on the mitochondria as demonstrated by decreased mitochondrial electron transfer activity and increased production of H2O2.

Dose-dependent inhibition of succinoxidase by buckminsterfullerene (C60)

251st National Meeting (2016)

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