A Nile Red staining method for microplastic identification and quantification

Abstract

Microplastics are classically visually identified with a microscope. Microscopic identification has low reliability especially in small (e.g. <200μm) transparent or white particles. FT-IR equipped with a microscope has been frequently applied to spectroscopic identification of micro-scale polymers including qualitative confirmation of polymer types. It is a little hard to detect microplastics less than 50μm in size. We aim to develop a subsidiary method to facilitate identification of synthetic polymers using a fluorescent dye.Nile Red (NR), a fluorescent dye, which was well known to dye neutral lipid in cell and tissue sample is used in this study. It is strongly fluorescent only in the presence of a hydrophobic environment. Applicability of NR for identification and quantification of microplastics was tested and staining condition was optimized in this study. Micro-sized polyethylene (PE), polypropylene (PP) and expanded polystyrene (EPS) were used as model plastics. Stained polymer particles were recognized better in green fluorescence (Ex. 534-558 and Em. 515-565) than red (Ex. 534-558 and Em. >590). The 50mg/L solution showed higher fluorescence than 5 mg/L solution. Fluorescence intensity was in the order of EPS > PP > PE. The developed NR staining method was successfully applied to identify and quantify the fragmented polymer particles in a laboratory accelerated mechanical abrasion study. Other polymeied to spectroscopic identification of micro-scale polymers including qualitative confirmation of polymer types. It is a little hard to detect microplastics less than 50μm in size. We aim to develop a subsidiary method to facilitate identification of synthetic polymers using a fluorescent dye.Nile Red (NR), a fluorescent dye, which was well known to dye neutral lipid in cell and tissue sample is used in this study. It is strongly fluorescent only in the presence of a hydrophobic environment. Applicability of NR for identification and quantification of microplastics was tested and staining condition was optimized in this study. Micro-sized polyethylene (PE), polypropylene (PP) and expanded polystyrene (EPS) were used as model plastics. Stained polymer particles were recognized better in green fluorescence (Ex. 534-558 and Em. 515-565) than red (Ex. 534-558 and Em. >590). The 50mg/L solution showed higher fluorescence than 5 mg/L solution. Fluorescence intensity was in the order of EPS > PP > PE. The developed NR staining method was successfully applied to identify and quantify the fragmented polymer particles in a laboratory accelerated mechanical abrasion study. Other polyme