(76f) Demonstrated Neuroprotective and Antioxidant Effects of Sugarcane Derivatives: In Vitro Rotenone-Induced Oxidative Stress on Neuroblastoma Cells (SH-SY5Y)

Plant-derived extracts from traditionally medicinal plants have proven to be an important source of bioactive compounds, many of which have been the basis for the development of novel pharmaceuticals. In Latin America, sugarcane (Saccharum officinarum) has been extensively used ancestrally to combat disease. In particular, non-centrifugal sugarcane (NCS), one of its primary products in the region, is traditionally used for the treatment of skin infections and malnutrition [1]. Some studies have determined the importance of the manufacturing process in the nutraceutical and antioxidant activities of non-centrifugal sugar [2] [3] [4]. The nutritional components enclose bioactive substances such as minerals, vitamins, and amino acids that are involved in beneficial contributions to human health [5]. The most interesting and potentially therapeutic activities of sugarcane and its derivatives (juice, syrup and non-centrifugal sugar) are their neuroprotection and antioxidant effects, which can be mainly attributed to the presence of a large amount of phenolic and flavonoid compounds in sugar cane juice [4] [6] [7]. The aim of this study is to confirm that these effects are observable in an in vitro neurodegenerative model where oxidative stress is induced by rotenone on neuroblastoma (SH-SY5Y). The inhibitory effect of rotenone is due to the blockage of the Complex I of the mitochondrial electron transport chain. In turn, this can ultimately cause degeneration of dopaminergic neurons by an exacerbated production of ROS [8] [9] [10]. The samples of sugarcane-derived products analyzed in the present study were obtained under controlled thermal processing conditions by Corporación Colombiana de Investigación Agropecuaria (Agrosavia), and were compared to commercially available NSC products fabricated under traditional methods. Antioxidant activity present in all five sugarcane derivatives was confirmed by DPPH and ABTS radical scavenging activity assays. In addition, neuroprotective effects were determined on SH-SY5Y cells treated with 150µM rotenone and cultured in a CO2 incubator for 24 hours. The cell response was evaluated by looking at changes in the mitochondrial membrane potential after exposure to a range of concentrations of juice, syrup, non-centrifugal sugar, pulverized sugarcane stalk and commercially available non-centrifugal sugar prepared in DMEM and incubated for an additional 24 hours. Changes in mitochondrial membrane potential were determined by JC-1 quantification using confocal microscopy and by analyzing a total of 25,000 events in a FACS CANTO flow cytometer. According to the results shown in Table 1, all the samples exhibit radical free scavenger activity. Syrup and non-centrifugal sugar, exhibited a significant DPPH radical scavenging activity, which can be related to their polyphenolic content. The observed trend was in accordance with the total phenolic content quantified by Folin–Ciocalteu method, where the highest concentration of the phenolic compounds corresponded to syrup and NCS. It is hypothesized that some of the phenolic content is lost during the lyophilization of the assayed samples. Furthermore, confocal and flow cytometry results allowed for the quantification of mitochondrial dysfunction and provided insights as to if the neuroprotective effect of sugarcane derivatives was mediated by biological events on the mitochondria. Treatment with sugarcane-derived samples led to a noticeable preventive effect against rotenone-induced mitochondrial malfunction in all the concentrations assessed as shown in Figure 1A. Confocal microscopy results shown in Figure 1B focused on the changes in red to green fluorescence (R/G) ratios, which were 3.759±0.977 for the control, 1.310±0.29 for traditional NCS, 2,219±0.784 for Agrosavia NCS, 2.979 ±0.758 for syrup, 2.267±0.537 for juice, and 3.455±0.442 for sugarcane. In rotenone-treated cells, red fluorescence decreased while green fluorescence increased, which strongly suggests loss of mitochondrial membrane potential. However, the R/G ratio of sugarcane derivatives, particularly of juice, demonstrated that mitochondrial membrane potential remained very close to that of the control (untreated samples).

In the present work, we found that sugarcane derivatives can protect neuroblastoma (SH-SY5Y) cells against rotenone-induced oxidative stress. Antioxidant compounds present in all samples appear capable of modulating ROS levels and thereby maintaining mitochondrial membrane potential, which suggests their prospective use in the treatment of neurodegenerative diseases. In conclusion, natural antioxidants are promising candidates as chemopreventive agents for neurodegenerative disease treatment.

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