Abstract
Red maple (Acer rubrum), also known as swamp, water or soft maple, is endemic to eastern and central North America and was widely used as traditional medicine by the first peoples. Commercially, its well-known products include maple syrup and high-quality lumber. The potential medicinal benefits of phenolic compounds extracted from the red maple plant, such as glucitol-core containing gallotannins, include antioxidant, and antiglycation effects as well as their importance in cosmetic applications. Plant-derived proteins and peptides are important biomolecules; however to date, there is no published data on the identification of proteins/peptides from red maple leaves. Therefore, the present study focuses on the activity guided purification of proteins from red maple leaves collected in spring and fall seasons. In addition, the focus of this project was in the evaluation of maple leaves employing bottom-up proteomics and De Novo protein profiling by PEAKS Studio-X and Gene Ontology Bioinformatics. The red maple leaves were grounded, defatted in hexane and proteins extracted in 25 mM sodium phosphate buffer pH 6.5. The extracted crude proteins were recovered by precipitation in 80% ammonium sulfate. The first-dimensional chromatography of crude extracted proteins was performed on a gel filtration column (HiLoad 16/600 Superdex200). The separation of crude extract and the partially purified gel filtration fraction was conducted by reversed-phase HPLC. The crude and eluted fractions were analyzed by SDS-PAGE gel electrophoresis. The extract was screened for cytotoxicity activity on Michigan Cancer Foundation-7 breast cancer (MCF-7), M.D. Anderson Metastasis Breast (MDA-MB-231) cancer cell lines and anti-inflammatory activity on murine macrophage (RAW 264.7) cell line from American Type Culture Collection (ATCC). The drug Doxorubicin was used as a positive control whereas untreated cells as a negative control in these experiments Preliminary data revealed that active protein fractions were eluted at two different regions of gel filtration chromatography both in spring and fall leaves. Bottom-up proteomics of crude and active fractions by PEAKS Studio-X and MASCOT bioinformatics database identified over 54 proteins. The Gene Ontology Annotation classified these proteins involved in the biological processing, cellular compartment, and molecular functions.
