Uncovering the Potential of Indonesian Rubus fraxinifolius Fruit: Optimization of Green Extraction with Simplex-Lattice-Design and Phytochemical Analysis

Abstract

Unlocking the full potential of Rubus fraxinifolius (R. fraxinifolius) fruit as a source of high-value ingredients necessitates efficient and green extraction protocols. This study employed ultrasound-assisted extraction (UAE) coupled with a simplex lattice design (SLD) to systematically optimize the ethanol-water solvent for maximizing the phenolic (TPC), flavonoid (TFC), and antioxidant capacities. The optimum R. fraxinifolius extract (ORFE) was subsequently profiled using gas chromatography-mass spectrometry (GC-MS). Our models revealed key complexity: with TPC maximizing in absolute ethanol, while TFC and antioxidant capacity peaked in distinct hydroethanolic mixtures. Numerical optimization identified an overall optimal composition of 92.2% ethanol and 7.8% water, which was experimentally validated to yield high TPC (106.2 mg gallic acid equivalent /g), TFC (30.114 mg quercetin equivalent/g), and potent 2,2-diphenyl-1-picrylhydrazyl scavenging capacity (91.2% inhibition). GC-MS analysis of ORFE revealed a unique chemical signature, identifying key semi-volatile contributors including anethole, γ-tocopherol, and a substantial alkaloid fraction. We conclude that the extract's potent bioactivity reflected the concerted action of the substantial non-volatile polyphenolic load, evidenced by the high TPC and TFC results, and the specific semi-volatile bioactive compounds identified by GC-MS. This work establishes a scientifically validated, green protocol for producing a chemically characterized, high-potency antioxidant ingredient from R. fraxinifolius for nutraceutical applications.