Phytochemical Analysis, Cytotoxicity, and Antitrypanosomal, Antioxidant, and Anti-Inflammatory Activities of Clausena anisata Fruit, Azadirachta indica Leaf, and Stem Bark Extracts

Human African trypanosomiasis (HAT) is an infectious disease which kills humans and animals as a result of hematological distortions, oxidative stress, tissue and neuroinflammations. This study reports on the differences in cytotoxicity, antitrypanosomal, antioxidant, and anti-inflammatory activities of ethanol extracts from Clausena anisata fruit (CFE), Azadirachta indica (neem) leaf (NLE), and stem bark (NSBE), medicinal plants used to treat HAT in its endemic communities. The extracts remarkably inhibited Trypanosoma brucei brucei (GUTat 3.1) parasite in vitro with CFE recording the highest effect with an IC₅₀ of 0.0055 (0.0955) μg/mL. The IC₅₀ of the standard Coptis japonicum was 0.5957 (0.0693) μg/mL. Also, the antitrypanosomal activity of NLE was 123.34% higher than that of NSBE. The percentage number of wells containing viable T. b. brucei parasites was very significantly (p < 0.001) reduced for all the extracts after 48 h of incubation. Furthermore, the extracts did not show cytotoxicity against the liver (HepG2) cells (CC₅₀s > 100 μg/mL and SI = 13.12–32,025.45). NSBE contained the highest quantity of phenolic compounds and flavonoids and also produced the highest antioxidant and anti-inflammatory activity in the DPPH free radical scavenging assay (IC₅₀ = 4.99 ± 0.018) and protein denaturation assay (IC₅₀ = 0.1805 ± 0.0002 μg/mL). In addition, phytochemical analysis showed that NLE contained the highest number of classes of phytochemical constituents (seven) among the extracts. These results indicate that CFE, NLE, and NSBE possessed significant antitrypanosomal activity as a result of their antioxidant and anti-inflammatory actions. However, a different mechanism was also involved in the antitrypanosomal activity of CFE and NLE, since their antitrypanosomal activity is greater than NSBE which demonstrated the highest antioxidant and anti-inflammatory activities. Due to the remarkable antitrypanosomal action of CFE, its constituents are being isolated for possible development into novel antitrypanosomal agents.