Optimization of Antioxidant Extraction From Moringa oleifera Seeds Using Response Surface Methodology: Phytochemical Analysis and DPPH Assay

Abstract

Moringa oleifera, a medicinal tree native to Africa and Asia, has been utilized for centuries in traditional medicine to help treat a variety of diseases. This paper examines the optimization of the extraction process, characterization, and antioxidant activities of Moringa oleifera seeds. Using central composite design and response surface methodology, the study assessed temperature (25°C–60°C), extraction time (24–72 h), ethanol concentration (40%–80%), and yields (28%–39.4%). Optimal conditions were 42.5°C, 40% ethanol, and 35.5 h, yielding 35.2%. Additionally, physicochemical analysis revealed an acid value of 1.98 mg KOH/g and an iodine value of 83.02 mg/g. These values indicate low free fatty acid content and a high degree of unsaturation. The extracts also exhibited significant antioxidant activities compared to ascorbic acid, with an IC₅₀ of 15.80 mg/mL for the extract and 12.60 mg/mL for ascorbic acid, demonstrating strong reducing power. Fourier transform infrared spectroscopy identified key phytochemical functional groups, such as phenols, flavonoids, steroids, terpenoids, and polyphenols (tannins), all of which are linked to antioxidant activity. Gas chromatography–mass spectrometry analysis identified and quantified various bioactive compounds, including 9,12,15-octadecatrienoic acid, ethyl ester; n-hexadecanoic acid; hexadecanoic acid, ethyl ester; 2-hexadecen-1-ol, 3,7,11,15-tetramethyl-, acetate, [R-[R∗,R∗-(E)]]; 9,12,15-octadecatrienoic acid, ethyl ester, (Z,Z,Z); octadecanoic acid, ethyl ester; and oleic acid 3-(octadecyloxy) propyl ester. Additionally, the p value < 0.0001 indicates high significance (0.01% noise chance), with an adjusted R² at 0.9992 and predicted R² at 0.9986. These compounds all contribute to the antioxidant capacity. These findings suggest that Moringa oleifera seed extracts are potent natural antioxidants, indicating their potential for developing functional health products or natural antioxidant formulations.