Name: sampling, sorting, and characterizing microplastics in aquatic environments with high suspended sediment loads and large floating debris
Uploaded: 2018-07-28T14:00:00
Description: Watch this Scientific Journal Video about Sampling, Sorting, and Characterizing Microplastics in Aquatic Environments with High Suspended Sediment Loads and Large Floating Debris at JoVE.com

The project for which this protocol was established was funded by the National Oceanic and Atmospheric Administration (NOAA) Marine Debris Program (# NA16NO29990029). We thank Miles Corcoran at the National Great Rivers Research and Education Center (NGRREC) in Alton, Illinois, for help with site selection and boat operation. Field and lab work was completed with the help of Camille Buckley, Michael Abegg, Josiah Wray, and Rebecca Wagner.

1. Water Sample Collection
<ol>
	<li>Collect water samples and water quality data of interest by boat where the river is well-mixed, ideally at locations where river stage or discharge is known (e.g., United States Geological Survey (USGS) gauging stations).20 To assure that the water is well-mixed, guide the boat using a handheld meter immersed in the river to where conductivity stays relatively constant.</li>
	<li>At the sampling sites, record location coordinates and depth. To find t...

<ol>
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R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Marine+pollution.+Plastic+waste+inputs+from+land+into+the+ocean.">Marine pollution. Plastic waste inputs from land into the ocean.</a> Science. 347 (6223), 768-771 (2015).</li><li>Andrady, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microplastics+in+the+marine+environment.">Microplastics in the marine environment.</a> Marine Pollution Bulletin. 62 (8), 1596-1605 (2011).</li><li>Cole, M., Lindeque, P., Halsband, C., Galloway, T. 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J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microplastic+is+an+abundant+and+distinct+microbial+habitat+in+an+urban+river.">Microplastic is an abundant and distinct microbial habitat in an urban river.</a> Environmental Science &amp; Technology. 48 (20), 11863-11871 (2014).</li><li>Farrell, P., Nelson, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trophic+level+transfer+of+microplastic:+Mytilus+edulis+(L.)+to+Carcinus+maenas+(L.).">Trophic level transfer of microplastic: Mytilus edulis (L.) to Carcinus maenas (L.).</a> Environmental Pollution. 177, 1-3 (2013).</li><li>Rochman, C. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scientific+evidence+supports+a+ban+on+microbeads.">Scientific evidence supports a ban on microbeads.</a> Environmental Science &amp; Technology. 49 (18), 10759-10761 (2015).</li><li>Taylor, M. L., Gwinnett, C., Robinson, L. F., Woodall, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Plastic+microfibre+ingestion+by+deep-sea+organisms.">Plastic microfibre ingestion by deep-sea organisms.</a> Scientific Reports. 6, 33997 (2016).</li><li>Mani, T., Hauk, A., Walter, U., Burkhardt-Holm, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microplastics+profile+along+the+Rhine+River.">Microplastics profile along the Rhine River.</a> Scientific Reports. 5, 17988 (2015).</li><li>Morritt, D., Stefanoudis, P. V., Pearce, D., Crimmen, O. A., Clark, P. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Plastic+in+the+Thames:+a+river+runs+through+it.">Plastic in the Thames: a river runs through it.</a> Marine Pollution Bulletin. 78 (1-2), 196-200 (2014).</li><li>. National Park Servies Available from: <a target="_blank" href="https://www.nps.gov/miss/riverfacts.htm">https://www.nps.gov/miss/riverfacts.htm</a> (2017)</li><li>. United States Census Bureau Available from: <a target="_blank" href="https://www.census.gov/geo/maps-data/data/tiger-data.html">https://www.census.gov/geo/maps-data/data/tiger-data.html</a> (2010)</li><li>. United States Geological Survey (USGS) Available from: <a target="_blank" href="https://waterdata.usgs.gov/nwis/rt">https://waterdata.usgs.gov/nwis/rt</a> (2016)</li><li>Grimes, C. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fishery+Production+and+the+Mississippi+River.">Fishery Production and the Mississippi River.</a> Fisheries. 28 (8), 17-26 (2001).</li><li>Talvitie, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Do+wastewater+treatment+plants+act+as+a+potential+point+source+of+microplastics?+Preliminary+study+in+the+coastal+Gulf+of+Finland,+Baltic+Sea.">Do wastewater treatment plants act as a potential point source of microplastics? Preliminary study in the coastal Gulf of Finland, Baltic Sea.</a> Water Science and Technology. 72 (9), 1495-1504 (2015).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> United States Environmental Protection Agency (USEPA) Method 160.2: Residue, Non-filtereable (Gravimetric, Dried at 103-105C). , (1971).</li><li>Nor, N. H., Obbard, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microplastics+in+Singapore's+coastal+mangrove+ecosystems.">Microplastics in Singapore's coastal mangrove ecosystems.</a> Marine Pollution Bulletin. 79 (1-2), 278-283 (2014).</li><li>Woodall, L. C., Gwinnett, C., Packer, M., Thompson, R. C., Robinson, L. F., Paterson, G. 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Microplastic collection using drift nets is the conventional method in environments like the ocean where both sediment and plastic concentrations are low, thus requiring large sample volumes. However, drift nets are not always practical or safe in rivers with high sediment loads and large floating or submerged debris. Additionally, it is not feasible to use a drift net when attempting to thoroughly capture and quantify microplastic materials, particularly fibers, as most nets used for plastic surveys have mesh sizes &#88...

Plastic was first observed in the ocean as early as the 1930s1. Recent estimates of marine plastic debris range from over 243,000 metric tons (MT) of plastic on the ocean&#39;s surface to 4.8-12.7 million MT of plastic entering the ocean from terrestrial sources annually2,3. Early studies on marine plastic debris focused on macroplastics (&#62;5 mm diameter) as they are easily visible and quantifiable. However, it was recently discovered that macroplastics represent &#60;10% of plastic debris, by count, in the ocean, indicating that the overwhelming majority of plastic debris is microplastic (&#60;5 mm diameter)2.
Microplastics are categorized into two groups: primary and secondary microplastics. Primary microplastics consist of plastics that are manufactured at a diameter &#60;5 mm and include nurdles, the raw pellets used to make consumer products, microbeads used as exfoliants in personal care products (e.g., facial wash, body scrub, toothpaste), and abrasives or lubricants in industry. Secondary microplastics are created within the environment as larger plastic debris is fragmented by photolysis, abrasion, and microbial decomposition4,5. Synthetic fibers are also secondary microplastics and are a growing concern. A single garment can release &#62;1,900 fibers per wash in a domestic washing machine6. These microfibers, as well as microbeads from personal care products, are washed down drains and into the sewer system before entering wastewater treatments plants. Murphy (2016) found that a wastewater treatment plant serving a population of 650,000 reduced the microplastic concentration by 98.4% from influent to effluent, yet 65 million microplastics remained in effluent and sludge each day7. Even with high percentages of microplastics being removed during the treatment processes, millions, possibly billions, of microplastics pass through wastewater treatment plants daily and enter surface waters in effluent6,8,9,10,11.
Due to their environmental release, microplastics have been found in the digestive and respiratory tissues of marine organisms across all trophic levels12,13,14,15. Their impact after uptake is variable, with some studies not observing harm, while others demonstrate numerous effects such as physical and chemical tissue damage4,6,14,15. Due to these discoveries, interest in this field has increased over the past five decades. However, only recently have studies begun to quantify plastic debris, particularly microplastics, in freshwater systems, such as rivers and lakes, or assess the effect on organisms dwelling in these habitats12,16,17,18. Rivers are a major source of plastic debris found in the ocean as they receive wastewater effluent and surface water runoff that contain microplastics and macroplastics.
The protocol detailed here can be used to collect microplastic samples where drift nets are not feasible; specifically, in aquatic environments with high concentrations of suspended sediments and large floating debris like the Mississippi River. The Mississippi River watershed is one of the world&#39;s largest and has a population of &#62;90 million people, likely making it one of the largest sources of plastic debris to the ocean19,20. Each year, the Mississippi River discharges an average of 735 km3 of freshwater into the Gulf of Mexico, along with high concentrations of suspended sediments (~60 to &#62;800 mg/L) and large debris13,21. Water samples were collected at two depths (i.e., surface and 0.6-depth) at various locations along the Mississippi River and its tributaries in translucent 1 L low-density polyethylene (PE) containers using a peristaltic pump. In the lab, samples were filtered using nylon mesh sieves and mixed cellulose ester membrane filters simultaneously with a custom-made 63.5 mm (2.5 in) polyvinyl chloride (PVC) cylinder with union joints to insert the sieves and filters22. The inclusion of PVC unions in the filtration device allows for filtration by as many or as few particle size classes as desired. Additionally, it can be used to capture microplastic debris down to sub-micron sizes using membrane filters when studying synthetic fibers. Once filtered, samples were dried and suspected plastics were identified and sorted from the mesh sieves and membrane filters under a stereomicroscope. Suspected plastics were then examined using micro-attenuated total reflectance Fourier transform infrared spectroscopy (micro ATR-FTIR) to eliminate non-synthetic materials or determine polymer type. Considering the size of microplastic particulates and fibers, contamination is commonplace. Sources of contamination include atmospheric deposition, clothing, field and lab equipment, as well as deionized (DI) water sources. Multiple steps are included throughout the protocol to reduce contamination from various sources while conducting all stages of the study.

<table border="0" cellpadding="0" cellspacing="0" style="width:1076px;" width="1075">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">1L Cubitainer Containers, Low-Density Polyethylene</td>
			<td style="width:127px;">VWR</td>
			<td style="width:119px;">89094-140</td>
			<td style="width:612px;">Containers used to collect and store samples.</td>
		</tr>
		<tr height="126">
			<td height="126" style="height:126px;width:219px;">2-1/2&#34; Clear Schedule 40 Rigid PVC Pipe</td>
			<td style="width:127px;">United States Plastic Corporation</td>
			<td style="width:119px;">34138</td>
			<td style="width:612px;">The PVC pipe used to make the device comes as an 2.43 m pipe. The pipe was then cut to the desired lengths for each section seperated by union joints. Section lengths were decided by predicting smaller pore sizes would clogg the device quicker. Longer sections were placed above the smaller pore sizes to collect and hold water to prevent needing to disassemble the device to change a filter while a sample remained in the device. For one filtration device one 18 in, one 12 in, and two 6 in peices are needed.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">2-1/2&#34; PVC SCH 40 Socket Union&#160;</td>
			<td style="width:127px;">Supply House</td>
			<td style="width:119px;">457-025</td>
			<td style="width:612px;">Union joints were glued to PVC pipe to house nylon sieves and mixed cellulose membranes.</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:219px;">Nylon 6 Woven Mesh Sheet, Opaque Off-White, 12&#34; Width, 12&#34; Length, 500 microns Mesh Size, 38% Open Area (Pack of 5)</td>
			<td style="width:127px;">Small Parts via Amazon</td>
			<td style="width:119px;">CMN-0500-C/5PK-05</td>
			<td style="width:612px;">Mesh sheets were cut into circles to match the diameter of the outer diameter of the PVC pipe. The edges were glued to esure no fraying would occur. The glue &#39;s diamter should not extend into the inner diameter of the PVC so that it will not be affected during filtration.&#160;</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:219px;">Nylon 6 Woven Mesh Sheet, Opaque White, 12&#34; Width, 12&#34; Length, 100 microns Mesh Size, 44% Open Area (Pack of 5)</td>
			<td style="width:127px;">Small Parts via Amazon</td>
			<td style="width:119px;">B0043D1TB4</td>
			<td style="width:612px;">Mesh sheets were cut into circles to match the diameter of the outer diameter of the PVC pipe. The edges were glued to esure no fraying would occur. The glue &#39;s diamter should not extend into the inner diameter of the PVC so that it will not be affected during filtration.&#160;</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:219px;">Nylon 6 Woven Mesh Sheet, Opaque White, 12&#34; Width, 12&#34; Length, 50 microns Mesh Size, 37% Open Area (Pack of 5)</td>
			<td style="width:127px;">Small Parts via Amazon</td>
			<td style="width:119px;">B0043D1SGA</td>
			<td style="width:612px;">Mesh sheets were cut into circles to match the diameter of the outer diameter of the PVC pipe. The edges were glued to esure no fraying would occur. The glue &#39;s diamter should not extend into the inner diameter of the PVC so that it will not be affected during filtration.&#160;</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:219px;">Mixed Cellulose Ester Membrane, 0.45um, 142mm, 25/pk</td>
			<td style="width:127px;">VWR</td>
			<td style="width:119px;">10034-914</td>
			<td style="width:612px;">Mixed cellulose membrane filter with 0.45 um was used as the last filter. A large diameter was used to allow the filter to be folded into a cone to increase surface area of the filter to prevent clogging.&#160;</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:219px;">Metal Mesh Basket Tea Leaves Strainer Teapot Filter 76mm Dia 3pcs</td>
			<td style="width:127px;">Uxcell via Amazon</td>
			<td style="width:119px;">a15071600ux0260</td>
			<td style="width:612px;">The mesh basket used to provide extra support for the membrane filter to prevent tearing when pressure was applied by a vacuum pump.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">1/2&#34; PVC Barbed Insert Male Adapter</td>
			<td style="width:127px;">Supply House</td>
			<td style="width:119px;">1436-005</td>
			<td style="width:612px;">A vacuum adapter was added to allow vacuum filtration in the case of slow filtration due to high sediment concentration.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">1/2 in. O.D. x 3/8 in. I.D. x 10 ft. PVC Clear Vinyl Tube</td>
			<td style="width:127px;">Home Depot</td>
			<td style="width:119px;">702229</td>
			<td style="width:612px;">Tubing used to connect the vacuum pump to the filtration device.</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:219px;">YSI Professional Plus Multiparameter Instrument with Quatro Cable</td>
			<td style="width:127px;">YSI</td>
			<td>6050000</td>
			<td style="width:612px;">Handheld meter used to measure additional water quality parameters parameters (e.g., turbidity, temperature, conductivity, pH, and dissolved oxygen (DO)).</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:219px;">2100P Portable Turbidimeter</td>
			<td style="width:127px;">Hach</td>
			<td style="width:119px;">4650000</td>
			<td style="width:612px;">Handheld meter used to measure turbidity.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">FEP-lined PE tubing</td>
			<td style="width:127px;">Geotech</td>
			<td style="width:119px;">87050529</td>
			<td style="width:612px;">Tubing used with perestaltic pump to collect water samples from desired depths.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">Geopump Peristaltic Pump Series II</td>
			<td style="width:127px;">Geotech</td>
			<td style="width:119px;">91350123</td>
			<td style="width:612px;">Pump used to collected water samples.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">MeiJi Techno EMZ-8TR Microscope</td>
			<td style="width:127px;">Microscope.com</td>
			<td style="width:119px;">EMZ8TR-PLS2</td>
			<td style="width:612px;">Microscope used analyze mesh sieves and membrane filters to quanitfy suspect microsplastics.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">Nicolet iS10 FTIR Spectrometer</td>
			<td style="width:127px;">Thermo Electron North America</td>
			<td style="width:119px;">912A0607</td>
			<td style="width:612px;">FTIR used to analyze suspect microplastics.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">Nicolet iN5 FTIR microscope</td>
			<td style="width:127px;">Thermo Electron North America</td>
			<td style="width:119px;">912A0895</td>
			<td style="width:612px;">FTIR microscope used to analyze suspect microplastics.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">Germanium (Ge) ATR</td>
			<td style="width:127px;">Thermo Electron North America</td>
			<td style="width:119px;">869-174400</td>
			<td style="width:612px;">Geranium ATR accessory used along with the Nicolet iN5 FTIR microscope to analyze suspect microplastic.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">Aluminum EZ-Spot Micro Mounts (Pkg of 5)</td>
			<td style="width:127px;">Thermo Electron North America</td>
			<td style="width:119px;">0042-545</td>
			<td style="width:612px;">Microscope slides used along with the Nicolet iN5 FTIR microscope to analyze suspect microplastic.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:219px;">Aluminum Coated Glass Sample Slides</td>
			<td style="width:127px;">Thermo Electron North America</td>
			<td style="width:119px;">0042-544</td>
			<td style="width:612px;">Microscope slides used along with the Nicolet iN5 FTIR microscope to analyze suspect microplastic.</td>
		</tr>
	</tbody>
</table>

sampling, sorting, and characterizing microplastics in aquatic environments with high suspended sediment loads and large floating debris

The ubiquitous presence of plastic debris in the ocean is widely recognized by the public, scientific communities, and government agencies. However, only recently have microplastics in freshwater systems, such as rivers and lakes, been quantified. Microplastic sampling at the surface usually consists of deploying drift nets behind either a stationary or moving boat, which limits the sampling to environments with low levels of suspended sediments and floating or submerged debris. Previous studies that employed drift nets to collect microplastic debris typically used nets with &#8805;300 &#181;m mesh size, allowing plastic debris (particles and fibers) below this size to pass through the net and elude quantification. The protocol detailed here enables: 1) sample collection in environments with high suspended loads and floating or submerged debris and 2) the capture and quantification of microplastic particles and fibers &lt;300 &#181;m. Water samples were collected using a peristaltic pump in low-density polyethylene (PE) containers to be stored before filtering and analysis in the lab. Filtration was done with a custom-made microplastic filtration device containing detachable union joints that housed nylon mesh sieves and mixed cellulose ester membrane filters. Mesh sieves and membrane filters were examined with a stereomicroscope to quantify and separate microplastic particulates and fibers. These materials were then examined using a micro-attenuated total reflectance Fourier transform infrared spectrometer (micro ATR-FTIR) to determine microplastic polymer type. Recovery was measured by spiking samples using blue PE particulates and green nylon fibers; percent recovery was determined to be 100% for particulates and 92% for fibers. This protocol will guide similar studies on microplastics in high velocity rivers with high concentrations of sediment. With simple modifications to the peristaltic pump and filtration device, users can collect and analyze various sample volumes and particulate sizes.

To validate the recovery rates of this protocol, three samples (V1-V3) from Oso Bay, Corpus Christi, Texas (adjacent to the Texas A&#38;M University Corpus Christi Campus), were spiked with 10 blue PE particulates (ranging from 50-100 &#181;m in diameter) and 50 green nylon fibers of various lengths (Figure 3). Sample TSS was calculated (Section 2) and then the samples were filtered using the methods outlined in Sections 3-5. The blue PE...

Watch this Scientific Journal Video about Sampling, Sorting, and Characterizing Microplastics in Aquatic Environments with High Suspended Sediment Loads and Large Floating Debris at JoVE.com

Sampling, Sorting, and Characterizing Microplastics in Aquatic Environments with High Suspended Sediment Loads and Large Floating Debris (Video) | JoVE

Most microplastic research to date has occurred in marine systems where suspended solid levels are relatively low. Focus is now shifting to freshwater systems, which may feature high sediment loads and floating debris. This protocol addresses collecting and analyzing microplastic samples from aquatic environments that contain high suspended solid loads.

Sampling, Sorting, and Characterizing Microplastics in Aquatic Environments with High Suspended Sediment Loads and Large Floating Debris

The ubiquitous presence of plastic debris in the ocean is widely recognized by the public, scientific communities, and government agencies. However, only ...

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Environment

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