Comparative Phytochemical Analysis and Pharmacological Screening of Different Solvent Extracts of Medicinal Plants

Abstract

Background: Medicinal plants are major sources of bioactive phytochemicals with antioxidant, anti-inflammatory, antidiabetic and antimicrobial activities. Azadirachta indica, Ocimum sanctum and Withania somnifera are widely used in traditional medicine and reported to contain diverse secondary metabolites; however, direct comparative studies of sequential nonpolar to polar solvent extracts across these species are limited. Objective: To compare the phytochemical profiles and in vitro pharmacological activities (antioxidant, anti-inflammatory, antidiabetic and antimicrobial) of hexane, chloroform, ethyl acetate and methanolic extracts of leaves of Azadirachta indica, Ocimum sanctum and Withania somnifera, and to assess correlations between solvent polarity, phytochemical content and biological activity. Methods: Leaves were collected, authenticated and air-dried. Sequential extraction was performed using Soxhlet (hexane, chloroform, ethyl acetate) followed by maceration for methanol to obtain crude extracts; percentage yields were calculated. Preliminary qualitative phytochemical screening (alkaloids, flavonoids, phenolics, tannins, saponins, glycosides, terpenoids, steroids, carbohydrates, proteins) was performed. Quantitative assays included total phenolic content (TPC, Folin–Ciocalteu), total flavonoid content (TFC, aluminium chloride), total alkaloid and total tannin estimations. Antioxidant activity was determined by DPPH, ABTS and FRAP assays (IC50 and equivalent values). Anti-inflammatory activity used the protein denaturation inhibition assay. Antidiabetic potential was assessed via α-amylase and α-glucosidase inhibition assays. Antimicrobial activity against representative Gram-positive (Staphylococcus aureus), Gram-negative (Escherichia coli) and fungal (Candida albicans) strains was evaluated by agar well diffusion and broth microdilution (MIC). Data are presented as mean ± SD (n = 3); statistical analysis used one-way ANOVA with Tukey’s post hoc test (p < 0.05). Results: Percentage yields varied by solvent and species: methanol produced the highest yields overall (Azadirachta indica 12.8 ± 0.4%, Ocimum sanctum 15.2 ± 0.5%, Withania somnifera 10.6 ± 0.3%). Qualitative screening revealed richest diversity in methanolic and ethyl acetate fractions. Quantitatively, methanolic extracts had the highest TPC (A. indica 186.3 ± 4.6 mg GAE/g; O. sanctum 214.9 ± 5.1 mg GAE/g; W. somnifera 162.7 ± 3.9 mg GAE/g) and TFC (A. indica 78.4 ± 2.2 mg QE/g; O. sanctum 112.6 ± 3.0 mg QE/g; W. somnifera 64.1 ± 1.8 mg QE/g). Antioxidant assays showed lowest IC50 values for methanolic extracts in DPPH (O. sanctum methanol 18.4 ± 0.9 µg/mL) and ABTS, with FRAP reflecting higher reducing power in polar extracts. Anti-inflammatory protein denaturation inhibition and antidiabetic (α-amylase and α-glucosidase) assays were strongest for methanolic extracts, particularly O. sanctum and A. indica (inhibition >65% at 250 µg/mL). Antimicrobial testing showed moderate activity for ethyl acetate and methanolic extracts (MICs 125–500 µg/mL), with Gram-positive S. aureus being more susceptible. Statistical analysis indicated significant differences across solvents (p < 0.05) and species. Conclusion: Solvent polarity strongly influences extraction yield, phytochemical composition and in vitro biological activities. Methanolic extracts consistently contained higher phenolic and flavonoid contents and exhibited superior antioxidant, anti-inflammatory and antidiabetic activities. Ocimum sanctum methanolic extract performed best overall, followed by Azadirachta indica. These findings support selection of polar solvents for recovery of bioactive phytochemicals from these species and provide comparative data to guide phytopharmaceutical development...