Quantification of Polybutylene Adipate Terephthalate-based Micro- and Nano-plastics from Soil Using Proton Nuclear Magnetic Resonance Spectroscopy

We developed this method so that we could learn more about the biodegradation of biodegradable plastics and soil. Biodegradable plastic products are designed to degrade completely after they are buried in soil, but we needed a way to measure small plastic fragments that might remain in the soil. Microplastic quantification methods have developed rapidly over the past few years and are not standardized between groups.

Microplastics have been quantified from soil using a variety of different technologies. We think developing methods for NMR spectroscopy is important to diversify access to microplastic quantification. NMR has previously been used to quantify PBAT from plastic products in soil.

We created and shared this protocol in order to adapt this technique into a tool environmental scientists and others can use to rapidly quantify PBAT from numerous soil samples in parallel, allowing for large scale environmental monitoring. Our protocol involves recovering PBAT polymer in a solvent. Because the method isn't focused on isolating individual plastic particles, it isn't limited by the size of micro or nanoplastics.

There are some trade-offs, but this allows us to learn about nanoplastic biodegradation in soil. Now that we can quantify micro and nano-sized plastic in soil, we want to learn more about how nanoplastics interact with soil constituents like organic matter and soil minerals. These interactions might be an important factor, protecting plastics from biodegradation by microbial enzymes.

To begin, collect and prepare the soil before the extraction procedure. In a fume hood, prepare a 100 milliliter 90 to 10 volume by volume mixture of chloroform and methanol for each sample to be extracted. Add 100 grams of dry soil, 100 milliliters of the chloroform methanol solution, and 20 glass beads into a glass extraction jar.

Seal the jar tightly with PTFE-lined lids to prevent solvent vapor from escaping. Place the extraction jar on a shaker table and shake at 200 revolutions per minute for eight hours. After shaking, allow the soil in the extraction jar to settle for at least four hours.

Now, uncap an extraction jar in a fume hood. Using a pipette, load the liquid extract onto a qualitative paper filter with an 11 micrometer pore size, and collect the filtrate in a labeled clean glass jar. Record the volume of the recovered solvent.

Allow the extract in soil to dry completely in the fume hood, which may take up to 24 hours. Then, safely discard the dried soil. Once the extract is dry, cap the sample jar and store it in a cool, dark, dry area for later use.

To begin, obtain dried soil extract containing polybutylene adipate terephthalate or PBAT. Add one milligram of the internal calibrant 1, 4-Dimethoxybenzene into each sample jar containing the dried extract. Resuspend the sample in 500 microliters of deuterated chloroform in a fume hood.

Cap the jar and tap it gently against a hand or bench top approximately 10 times on each side. Then, use a clean pipette tip to transfer the dissolved sample into an NMR tube. Repeat the resuspension step with another 500 microliters of deuterated chloroform, and pull the sample.

Cap the NMR tube and store it for three days before collecting the NMR spectra. The spectrum of shady loam soil spiked with 63 milligrams per kilogram of PBAT showed all five characteristic PBAT peaks clearly resolved. Calibration curves showed strong correlations between the amount of PBAT added and measured in shady loam, Elkhorn sandy loam, and Los Osos loam, with shady loam achieving the highest extraction efficiency at 76%PBAT extraction efficiency from shady loam was higher for microplastics than for nanoplastics.