Caffeic acid and caffeine attenuate toxicity associated with malonic or methylmalonic acid exposure in Drosophila melanogaster.

The deficiency in the activity of the mitochondrial enzyme methylmalonyl-CoA mutase (MCM, EC 5.4.99.2) leads to a condition called methylmalonic academia, which is characterised by the accumulation of methylmalonic (MMA), malonic (MA) or other organic acids. Importantly, we have recently found that supplementation with Ilex paraguariensis aqueous extract offered protection against toxicity associated with MMA or MA exposure to Drosophila melanogaster. Of note, caffeic acid (CA) and caffeine (CAF) were the major phytochemicals found in our Ilex paraguariensis crude extract. Therefore, here, we have exploited CA and/or CAF to test the hypothesis that supplementation with the isolated compounds (either alone or combined) could exert a protective effect against MMA or MA-induced toxicity in flies. Therefore, flies were exposed to MA (5 mM) or MMA (5 mM) and concomitantly treated with CA (1.39 µg/mL), CAF (1.27 µg/mL) or CA + CAF for 10 days for survival, and for 4 days for behavioural and biochemical assays. CA, CAF and CA + CAF treatments completely abolished the mortality associated with either MMA or MA exposure. Moreover, CA and CAF, either alone or combined, completely abolished behavioural changes, and completely protect against changes in thiobarbituric acid reactive substances (TBARS) levels, catalase (CAT) activity and MTT reduction ability, associated with MA or MMA exposure. In turn, CAF restored SOD activity in the head of flies exposed to MA or MMA. However, CA and CAF (either alone or combined) significantly decreased acetylcholinesterase (AChE) activity per se, while CAF alone protected from changes in AChE activity (in head tissue) associated with MA or MMA. Finally, CA and/or CAF were able to protect from a decrease in glucose and triglyceride levels associated with both MA and MMA exposures in haemolymph. Together, our data confirm the hypothesis that supplementation with CA and/or CAF offers protection against detrimental changes associated with MMA or MA exposure in flies, being responsible, at least in part, for the protective effect of I. paraguariensis crude extract which was reported previously.

Ilex paraguariensis Attenuates Changes in Mortality, Behavioral and Biochemical Parameters Associated to Methyl Malonate or Malonate Exposure in Drosophila melanogaster.

Methylmalonic acidemia is a genetic disease characterized by accumulation of organic acids, such as methylmalonic (MMA) and malonic (MA) acids. Considering that the accumulation of MMA and MA causes several damages due to oxidative stress, antioxidants are thought to play a pivotal role in preventing deleterious effects associated with exposure to such compounds. Ilex paraguariensis (IP) was used here to test the hypothesis that supplementation with the aqueous extract of this plant could exert protective effect against MMA or MA induced mortality, behavioral and/or biochemical changes in Drosophila melanogaster (DM). Initially, a curve time- and dose-response to MMA (1-10 mM), MA (1-10 mM) and IP (63-500 µM) was performed. Thereafter, flies were concomitantly exposed to MA (5 mM), MMA (5 mM) and/or IP (250 µg/mL) during 15 days for survival assay, and for 48 hs to MA (1 or 5 mM), MMA (1 or 5 mM) and/or IP (250 µg/mL) for subsequent investigations. Both MMA and MA exposure resulted in higher incidence of mortality, a worse performance in the negative geotaxis assay and increased locomotion in open-field test as compared with control group. Furthermore, a marked increase in non-protein thiol (NPSH) and in thiobarbituric acid reactive substances (TBARS) levels, decrease in superoxide dismutase (SOD), catalase and acetylcholinesterase (AChE) activities, and decrease in MTT and resazurin reduction were noted in MMA or MA treated groups. IP treatment offered significant protection against all alterations associated to MMA or MA exposure. This study confirm the hypothesis that supplementation with IP offers protection against changes associated to MMA or MA exposure in DM, due, at least in part, to its antioxidant effect.

Ilex paraguariensis crude extract acts on protection and reversion from damage induced by t-butyl hydroperoxide in human erythrocytes: a comparative study with isolated caffeic and/or chlorogenic acids.

BACKGROUND: Studies comparing the effects of phytochemicals under different regimens of exposure are necessary to give a better indication about their mechanism(s) of protection. Hence, in the present study, we investigated the preventive (pre-incubation), protective (co-incubation) and/or remediative (post-incubation) activity of chlorogenic acid and caffeic acids, in comparison with Ilex paraguariensis crude extract, against t-butyl hydroperoxide (t-BHP)-induced damage to human erythrocytes. RESULTS: We found that both caffeic and chlorogenic acids were able to prevent and revert the hemolysis associated with t-BHP exposure. By contrast, isolated compounds (alone or in combination) presented no effect on basal and/or t-BHP-induced non-protein thiol (NPSH) oxidation or production of thiobarbituric acid reactive substances (TBBARS). In turn, I. paraguariensis extract was effective to prevent, protect and revert the hemolysis associated with t-BHP exposure. Moreover, I. paraguariensis significantly protects and reverts t-BHP-induced NPSH oxidation and TBARS production. CONCLUSIONS: We have found that I. paraguariensis extract acts better with respect to the protection and reversion of t-BHP-associated changes, whereas isolated compounds are more active in preventing and reverting t-BHP pro-hemolytic action. Moreover, our data suggest that the pro-hemolytic activity of t-BHP may occur via mechanism(s) other(s) than lipid peroxidation and/or NPSH oxidation. © 2016 Society of Chemical Industry.