Sampling and Identification of Microplastics in Groundwater

Protocol for the Sampling and Identification of Microplastics from Groundwater. Preparing the borehole for sampling. Open the borehole and remove any samplers if present.

Measure the groundwater level using the water level meter. Record the GPS coordinates of the borehole, the date of sampling, water level, and other sampling details on the data sheet. Setting up the sampling equipment.

Assemble and carefully lower the submersible pump into the well to the desired depth. Set up the filtering system without the filter support screens and filters, and place it horizontally. Close the main valve and open the bypass valve.

Connect the water supply hose. Cleaning the borehole. Start the submersible pump and let the water flow through the bypass to clean the borehole.

Cleaning the filtering system before sampling. Open the valves of the sampling branches and the main valve of the filtering system. Afterwards, close the bypass valve.

Inserting the filters. First, open the bypass valve and then close the main valve. Open the filter chamber and check that the filter chambers are clean.

Rinse with ultrapure water if necessary. Insert the filter support screen. Rinse the filter with the desired pore size with ultrapure water and place it on the filter support screen.

Close the filter chamber. Repeat the process for all branches. Several filters with decreasing pore sizes can be inserted sequentially for cascade filtration.

Sample collection. Read and record the water meter or reset it to zero. Open the main valve, close the bypass valve, and mark the start time of the sampling.

Monitor the pressure gauge during sampling to ensure the pressure does not exceed four bars to avoid damaging the equipment and minimizing microplastic fragmentation. Stop sampling when the planned volume of water is filtered or when the filters start to clog, that can be observed by the increased pressure or significantly decreased flow. To stop sampling, first open the bypass valve and then close the main valve and the valves on the filtering branches.

Turn off the pump. Record the time and final readings from the water meter. Collecting the filters.

Rinse a Petri dish with ultrapure water. Open the filter chamber and carefully transfer the filter maintained in a horizontal position into a clean Petri dish. Seal the Petri dish with a sealing film and label it with the sample name and date of sampling.

Repeat the procedure for all filter chambers. After the sample collection, disassemble the system. Always rinse the system with fresh water and dry it before storage.

Separation of microplastics from samples. Remove the sealing film and open the Petri dish. Transfer the Petri dish under the stereo microscope with at least 30 times magnification and search for potentially plastic particles, focusing on characteristics such as color, shape, and other visible features.

With the transfer of each particle that seems to be plastic, take a photo and measure its size. The following properties have to be determined for each microplastic particle:size, shape, color, and chemical composition. Keep in mind that some microplastics will be easily identifiable by their color and shape, while others may be more challenging.

Chemical identification of microplastics. The chemical analysis of potential microplastic can be done for large microplastics by ATR-FTIR, and for small microplastics by micro-FTIR. Alternative methods, such as Raman, are also possible ATR-FTIR spectroscopy.

Before beginning the analysis, thoroughly clean the ATR crystal and sample pressor using alcohol and a lint-free cloth. Configure the measurement settings, typically 16 scans with a wave number ranging from 4, 000 to 450 reciprocal centimeter and resolution 4 reciprocal centimeter. Then collect the background spectrum.

Place the particles one by one onto the ATR crystal, apply pressure and initiate the measurements. Match the acquired infrared spectra with those in reference libraries to confirm the particles as microplastic. Typically, a 70%correlation with library spectra is considered sufficient for positive identification.

Export the obtained data for further analysis and reporting. Micro-FTIR spectroscopy. Ensure all relevant parts of the instruments, such as the stage, are cleaned with alcohol and a lint-free cloth prior to analysis.

If sampling has not been done directly on a surface suitable for the selected measurement mode, such as ATR or reflection, place the potential plastic particles on an appropriate reflective surface, such as gold or aluminum-coated membranes or microscope slides. Select the measurement settings for the session, including the number of scans, spectral range, resolution, and session name. Locate your sample and capture a mosaic image of the area where all particles are situated.

For reflection measurements, first measure the background before measuring the infrared spectra of potential plastic particles. Identify and mark the points where infrared spectra of the selected particles will be measured. If necessary, select multiple points on each particle.

After selecting all the points, start the measurement. It's also feasible to obtain infrared images of larger areas of interest, particularly if the sample is collected directly on a suitable infrared measurement surface. Compare the collected infrared spectra with those from reference libraries.

Choose a threshold value for establishing the presence of microplastics. Typically, a 70%match is sufficient for positive identification. Export the obtained data for further analysis and reporting.

Representative results. The main result of this protocol is the database with all isolated microplastic particles, each characterized by shape, size, color, and material composition. The concentration of microplastic per sample and sampling location can be calculated.Conclusion.

This comprehensive protocol ensures accurate identification of microplastics for research or monitoring purposes, considering the recommendations from the annex of Commission Delegated Decision EU 2024/1441.