Chronic inflammation and oxidative stress are recognized as significant contributors to the development and progression of various debilitating diseases, including diabetes, cancer, and cardiovascular disorders. The search for natural products with potent anti-inflammatory, antioxidant, and antiglycation properties has gained considerable momentum due to their perceived lower adverse effects compared to synthetic drugs. Annona crassiflora, a native plant of the Brazilian Cerrado, has been traditionally used and has shown promising biological activities in its fruit. Considering the rich biodiversity of the Cerrado and the therapeutic potential associated with Annona crassiflora, this study explored the leaves of this plant as a source of bioactive compounds capable of mitigating oxidative stress and inflammation. The hydroethanolic extract of Annona crassiflora leaves (EHAC) was hypothesized to possess significant therapeutic potential against diseases linked to these processes.
Methods
The study involved the preparation of a hydroethanolic extract from Annona crassiflora leaves. This extract underwent phytochemical characterization using spectrophotometry and HPLC to identify its phenolic and flavonoid components. The in vitro assays conducted to evaluate the bioactivity of the EHAC included assessment of cytotoxicity on fibroblast cells using the MTT assay, anti-inflammatory activity through phagocytosis and macrophage spreading tests, antioxidant potential via lipid peroxidation inhibition and DPPH radical scavenging assays, and antiglycation activity using BSA/glucose and BSA/MGO systems. Statistical analysis was performed using ANOVA and Tukey’s test to determine the significance of the results.
Key Points
•Phytochemical Characterization: Spectrophotometric analysis of the EHAC revealed a high concentration of total phenols (23.08 ± 1.58 g GAE/100 g) and flavonoids (2.27 ± 0.08 g QE/100 g). HPLC analysis further identified the significant presence of the flavonoids rutin and quercetin in the extract. These findings are consistent with previous research on Annona crassiflora.
•Cell Viability: The EHAC showed low cytotoxicity to fibroblast cells, with cell viability remaining above 95% at concentrations up to 800 µg/mL. A significant reduction in cell viability was observed only at the high concentration of 1600 µg/mL (60.46%), and the IC50 was determined to be 2300 µg/mL. This suggests a relative safety of the extract at pharmacologically relevant concentrations.
•Anti-Inflammatory Activity: The EHAC demonstrated significant anti-inflammatory effects by inhibiting both phagocytosis (up to 92.33% at 800 µg/mL) and macrophage spreading (up to 88.55% at 800 µg/mL) in a concentration-dependent manner. At a concentration of 800 µg/mL, the inhibition of these processes was comparable to that of the positive control, dexamethasone. This activity is potentially linked to the presence of quercetin, known for its ability to negatively regulate macrophages and reduce pro-inflammatory cytokines.
•Antioxidant Activity: The EHAC exhibited high antioxidant capacity in both the lipid peroxidation inhibition (up to 99.23% at 800 µg/mL) and DPPH radical scavenging (100% at all tested concentrations) assays. The extract’s activity was comparable to or exceeding that of the positive control, quercetin. The presence of quercetin and rutin, known antioxidants, likely contributes to this activity by neutralizing free radicals and reducing oxidative lesions.
•Antiglycation Activity: The EHAC displayed 100% antiglycation activity at all tested concentrations in both the BSA/glucose and BSA/MGO systems, demonstrating efficacy similar to or greater than the positive control, quercetin. This suggests a strong potential for preventing the formation of advanced glycation end products (AGEs), which are implicated in various diseases, including diabetes mellitus.
This in vitro study provides compelling evidence for the therapeutic potential of the hydroethanolic extract of Annona crassiflora leaves (EHAC). The novelty of this research lies in its comprehensive investigation of the phytochemical profile and the simultaneous evaluation of cytotoxicity, anti-inflammatory, antioxidant, and antiglycation activities of the leaf extract. The findings highlight the presence of significant levels of bioactive compounds, particularly rutin and quercetin, which likely contribute to the observed pharmacological effects. The low cytotoxicity and potent bioactivities of the EHAC suggest its promise as a basis for developing a novel plant-based therapeutic agent for diseases associated with oxidative stress and inflammation. Future implications of this research include further in vivo studies to validate these effects in living organisms and the potential development of a phytotherapeutic product, contributing to the sustainable use and preservation of the biodiversity of the Brazilian Cerrado.
Link to the study: https://www.mdpi.com/2079-9284/12/2/36
