Assessing the Lost Fraction: Diversity, Abundance, and Mass of Microplastics (1-300 µm) in Aquatic Systems

Summary

This protocol describes a methodology to be applied to aquatic samples collected on filters to detect, identify, and quantify micron-sized (1-300 µm) MPs. Raman microspectroscopy can identify the polymeric chemical structure of MP particles and quantify their abundances in terms of the number of particles and their mass in water samples.

Abstract

The protocol presented here enables the quantification of microplastics (MPs) as small as ~1 µm in diameter, accurate identification of polymer types, and estimation of particle volume, critically allowing for the calculation of MP mass. Representative results from samples collected in the Great South Bay (GSB), NY, showed that particles within the 1-6 µm equivalent spherical diameter (ESD) range were the most abundant, with approximately 75% of particles measuring less than 5 µm. Notably, the pre-sieving step failed to yield any particles larger than 60 µm, suggesting that large MPs were rare at the coastal sites sampled. Prior to filtration, a chemical oxidation step was used to remove organic debris, which facilitated the filtration of larger water volumes (>1 L) from discrete bottle-collected samples and reduced filter clogging. While this approach significantly improved filtration efficiency, aspects of the methodology still require refinement to reduce the total time required for sample preparation and data analysis. Raman microspectroscopy and associated data processing remain time-intensive, particularly for accurately analyzing particles smaller than 10 µm in complex environmental matrices. Ongoing efforts are focused on minimizing analytical uncertainties, optimizing the trade-off between particle counting accuracy and processing time, and reducing artifacts in the detection workflow.

Introduction

Microplastics (MPs) are a highly heterogeneous group of particles that vary widely in size, shape, color, density, chemical composition, and other physical properties, making their identification and quantification particularly challenging¹. A variety of analytical methods have been employed in MP surveys, from visual techniques such as light microscopy² to more advanced chemical approaches like pyrolysis-gas chromatography³. Light microscopy is a commonly used method for characterizing larger MPs (typically 0.5-5 mm) due to its simplicity, speed, and cost-effectiveness¹. I....

Protocol

This protocol has been modified from the original version⁹. The protocol is a guidance for the analysis of aquatic samples (drinking water, river, lake, coastal, or open ocean) to extract, detect, and quantify MPs (<300 µm) on filters. The reagents and equipment used are listed in the Table of Materials.

1. Quality control

1. Ensure to filter all reagents through a 0.22 µm membrane filter before use.
2. Ensure the personnel wear cotton lab coats (no synthetic materials) and wash their hands before and between sample handling.
3. Rinse the filtration equipment w....

Results

Here, results obtained using the most recent version of the methodology are presented. The method has been modified from its original application^9,14. Samples used in this article are part of a project to assess MP pollution in three main coastal water bodies of Long Island, NY: Great South Bay, the Peconic Estuary, and Shinnecock Bay. The results presented here serve as a proof-of-concept for our improved method rather than a systematic assessment of water quali.......

Discussion

This methodology enables the quantification of microplastic (MP) particles as small as 1 µm from water samples collected on filters, allowing for detailed measurements of particle abundance, chemical composition, size, and mass. However, it is essential to recognize and address sources of uncertainty inherent in this approach, particularly when analyzing particles smaller than 10 µm in complex environmental matrices.

The current lack of standardized procedures for determi.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Aluminum foil	any commercial brand
Analytical stainless-steel sieves	Retsch GmbH, Germany	DIN 4188
Anodisc Filters	Whatman
Filter holder	FisherBrand-Millipore Sigma	XX1012542
Glass Bottles	FisherBrand-Millipore Sigma	02-912-313	Any glass container works for the sampling process
Glassware funnel	FisherBrand-Millipore Sigma	XX1012514
Hidrogen Peroxide 30 %	Innovating Sciences	7722-84-1
inVia confocal Raman microspectrophotometer	Renishaw
Laminar Flowhood	Labconco or any other laminar hood fro laboratoy use
Metal clamp	FisherBrand-Millipore Sigma	XX1012503
Metal forceps	FisherBrand-Millipore Sigma	13-820-061
Microscope slides	Thermo Fisher	https://www.thermofisher.com/
Nitrile gloves	Kimtech	55080-54
Nitrocelulose membrane fliters 0.22	GVS filter technology	1214898
Radial heat Oven	Lab Line Instruments	3609
Vacuum Pump	Welch	25468-01 A
WiRE 5.2	Renishaw