| Dry leaf of Mate was water distilled, which was then extracted with dichloromethane using a liquid-liquid continuous apparatus. In addition, Mate leaf was extracted with hexane and methanol sequentially by a soxhlet extractor. The methanol extract showed strong antioxidative activity in a thiobarbituric acid (TBA) assay (90.24 +/- 0.55%). The methanol extract was further fractionated into seven fractions using column chromatography with elution of water/methanol mixtures and acetone. After removal of solvents, dried samples from all seven fractions were tested with a TBA assay, malonaldehyde/gas chromatography (MA/GC) assay, 1,1,-dipheny1-2-picrylhydrazyl (DPPH) free radical assay and hexanal/hexanoic acid assay. The fraction eluted with a methanol/water (60/40) solution exhibited the most potent antioxidative activity. This fraction inhibited the formation of malonaldehyde in the TBA assay (97.19 +/- 1.75%) and the MA/GC assay (79.59 +/- 3.71%) at the level of 1.0 mg/mL, and had a free radical scavenging ability (86.4 +/- 0.10%) at the level of 0.125 mg/mL. The constituents of the 60% methanol fraction were isolated and identified by high performance liquid chromatography (HPLC) and liquid chromatography/mass spectrometry (LC/MS). The active compounds found in this fraction were 3,4-dicaffeoylquinic acid, 3,5-dicaffeoylquinic acid, 4,5-dicaffeoylquinic acid, rutin and luteolin diglucoside. |
