## 3 Results and Discussion
### Cytotoxic Effects of C. pepo Seed Extracts towards Intestinal and Neuronal Cells  
Caco-2, an intestinal cell line, was used to evaluate the cytotoxicity of the _C. pepo_ extracts obtained by UAE and MAE. This cell line allows the study the compound absorption across the intestinal epithelium, being commonly employed to evaluate the extracts safety [97]. According to Figure 1, the exposure of this cell line to increasing concentrations (0.1-1000 \(\mathrm{\SIUnitSymbolMicro g}\)/mL) of MAE _C. pepo_ extracts showed a viability around 100%, without significant differences (\(p>0.05\)). On the other hand, only the concentration of 0.1 \(\mathrm{\SIUnitSymbolMicro g}\)/mL of the UAE condition 1 extract maintained a viability of 101.36%, being significantly different from the other concentrations (\(p<0.05\)). At higher concentrations, the viability ranged between 70.20% (1000 \(\mathrm{\SIUnitSymbolMicro g}\)/mL) and 77.82% (1 \(\mathrm{\SIUnitSymbolMicro g}\)/mL). The UAE condition 2 displayed viabilities above 88.78%, being not significantly different (\(p>0.05\)). The lower viability displayed by Caco-2 cells incubated with UAE _C. pepo_ extracts may be explained by the higher content on neochlorogenic acid and caftaric acid present in the extracts. These compounds have antiproliferative activity and may effectively inhibit the growth of colorectal cancer cells [98; 99]. In the present study, NSC-34 cell line was used to evaluate the potential neurotoxicity of _C. pepo_ extracts obtained by UAE and MAE (Figure 2). NSC-34 is a hybrid cell line that has similar morphological and physiological properties to motor neurons [100]. This cell line in culture mimics the synthesis and storage of acetylcholine (ACh) and the expression of neurofilament proteins [100]. After 24 h of incubation in the presence of different concentrations (0.1-1000 \(\mathrm{\SIUnitSymbolMicro g}\)/mL) of _C. pepo_ extracts, it was possible to observe a decrease of the NSC-34 cell viability, mainly in the MAE and UAE condition 1 extracts (Figure 2). Indeed, at the highest concentration tested (1000 \(\mathrm{\SIUnitSymbolMicro g}\)/mL), the cells' viability was 9.92% and 19.58%, for MAE condition 1 and UAE condition 1, respectively, with significant differences (\(p<0.05\)). In these cases, it was possible to determine the IC\({}_{50}\) values of 253.68 (MAE condition 1) and 95.32 \(\upmu\)g/mL (UAE condition 1). Figure 1: Effects of _C. pepo_ extracts obtained by UAE and MAE exposure on the viability of Caco-2 cells at different concentrations (0.1–1000 \(\upmu\)g/mL), measured by MTT assay. Values are expressed as mean \(\pm\) standard deviation (\(n=3\)). Different letters represent significant differences between concentrations of the same sample (\(p<0.05\)), according to [PERSON]’s HSD test. Figure 2: Effects of _C. pepo_ extracts obtained by UAE and MAE exposure on the viability of NSC-34 cells at different concentrations (0.1–1000 \(\upmu\)g/mL), as measured by the MTT assay. Values are expressed as mean \(\pm\) standard deviation (\(n=3\)). Different letters indicate significant differences between concentrations of the same sample (\(p<0.05\)), according to [PERSON]’s HSD test. Oppositely, MAE and UAE condition 2 showed an increasing trend with increased concentrations. For MAE condition 2, the results varied from 54.17% (0.1 \(\upmu\)g/mL) to 82.97% (1000 \(\upmu\)g/mL). In the case of UAE condition 2 extracts, the highest cell viability (98.21%) was found for the highest concentration tested (1000 \(\upmu\)g/mL), following in descending order 100, 1, 10 and 0.1 \(\upmu\)g/mL. The inferior cells' viability observed after exposure to MAE condition 1 and UAE condition 1 extracts may be due to the presence in higher amounts of the antiproliferative compounds neochlorogenic acid and catfaric acid, which is not observed in MAE condition 2 and UAE condition 2 extracts.