Source: UNIVERSITY OF WASHINGTON submitted to NRP
INSTRUMENT ACQUISITION FOR CHARACTERIZATION OF MICROPLASTICS IN SEAFOOD AND ASSESSMENT OF PLASTIC DEGRADATION OF AQUACULTURE GEAR
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
Reporting Frequency
Annual
Accession No.
1024371
Grant No.
2020-70410-32904
Cumulative Award Amt.
$112,754.00
Proposal No.
2020-07804
Multistate No.
(N/A)
Project Start Date
Sep 1, 2020
Project End Date
Aug 31, 2023
Grant Year
2020
Program Code
[EGP]- Equipment Grants Program
Recipient Organization
UNIVERSITY OF WASHINGTON
4333 BROOKLYN AVE NE
SEATTLE,WA 98195
Performing Department
College of the Environment
Non Technical Summary
Plastic pollution is a threat to seafood safety with long term impacts to marine organisms and human health. When plastics enter the ocean, they can break into smaller pieces (microplastics; < 5mm) and contaminate the marine ecosystem, including commercial species intended for human consumption. Identifying what type of microparticles are ingested by marine life is critical to understand the impacts these pollutants may have on organisms and human health. The leading technology to identify, characterize, and quantify microplastics from a variety of sources is Micro-Fourier Transform Infrared (µ-FTIR) spectroscopy. The use and relevance of this technology has increased dramatically in the areas of aquaculture and agriculture to examine synthetic particles from biological and sediment samples. To date, however, this technology is not available at any facility at the University of Washington. This proposal seeks to fill this important gap by requesting funds to purchase a µ-FTIR microscope and spectral library. Keeping up to date with technology is essential to meet and develop food safety standards in a growing industry. The acquisition of a µ-FTIR microscope will increase research and education capabilities across fields and will benefit investigators from multiple departments and institutions involved in food safety, marine pollution, chemistry and material science research. Specifically, the proposed work will establish important ecological baselines for microplastics in shellfish in the Pacific Northwest Region and assess plastic degradation of aquaculture gear. This project will address the USDA-NIFA strategic goal of Providing all Americans Access to a Safe, Nutritious, and Secure Food Supply.
Animal Health Component
50%
Research Effort Categories
Basic
30%
Applied
50%
Developmental
20%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
13308111070100%
Knowledge Area
133 - Pollution Prevention and Mitigation;

Subject Of Investigation
0811 - Shellfish;

Field Of Science
1070 - Ecology;
Goals / Objectives
This project will address the USDA-NIFA Challenge Area of Global Food Security. Specifically, this project will generate both scientific understanding and proactive solutions for a prevalent, anthropogenic contaminant--microplastics--currently threatening the security and sustainability of the aquatic food production system in the United States.Marine plastic pollution is a serious threat to the aquaculture industry around the globe with long term impacts to coastal environments and the businesses that depend on them.This proposed project will provide information to the largest shellfish industry in the United States to understand what type of microplastics are present and if aquaculture gear is contributing to microplastic pollution. These are critical information gaps of serious concern to the shellfish aquaculture industry, potentially hindering aquaculture development.OBJECTIVES1) Identify the seasonal abundance and types of microplastics in Washington state.2) Assess plastic and rope degradation and examine how aquaculture gear may be contributing to microplastic contamination in locations with different environmental conditions.
Project Methods
Objective 1 - Identify the seasonal abundance and types of microplastics (Years 1-2).Shellfishwill be obtained from ten sites in Washington during summer and winter (N=600 for each species). Collection sites will be located in areas with distinct environmental conditions (i.e. temperature, light) and proximity to metropolitan areas. At each site, we will also collect sediment and water samples (n=5 per sampling period per site). At selected stations, we will also collect samples (shellfish, water and sediments) 1-3 days after storm events, since new research shows that stormwater is the primary source of microplastics in coastal waters.Microplastic extraction - Length of bivalves will be measured with a digital caliper to the nearest mm. Then, the tissue will be removed from the shells, weighed with a digital scale and placed in individual bags in a -20C freezer. To obtain the microplastics from shellfish, we will use a standardized extraction protocol for microplastics in seafood developed by Dehaut et al. (2016). This protocol was found to be superior to other methods because it led to efficient tissue digestion with no significant degradation of the polymers (Dehaut et al. 2016). Tissue will be placed in a glass beaker and digested with 20 ml of potassium hydroxide KOH in an oscillating incubator at 60C for 24 hs. Samples will be covered with aluminum foil to prevent dehydration and airborne contamination. Three samples and a blank extraction control will be processed each time. The resulting digested solution (potentially containing microplastics) will be filtered with 5µm pore size cellulose nitrate membrane using a vacuum pump system. The filter membrane will be placed in a covered Petri dish, left to dry, and observed under a microscope to identify, count and collect the microplastics retained.To obtain microplastics from sediment and water samples we will follow the oil extraction protocol outlined by Crichton et al. (2017). In this method, the oleophilic property of plastics causes them to adhere to the oil which floats to the surface while sediments sink to the bottom. In brief, 400 g of wet sediment will be placed in a clean glass beaker and dried overnight at 60 °C, then weighed. If biological material is present, digestion using KOH will be performed as described above (Dehaut et al. 2016). Microplastics will be floated out of the sediment by adding 800 ml of filtered water and 40 ml of filtered canola oil, then vigorously swirling the solution for > 30 s and allowing the solution to settle. This step will be performed at least two times. The water and oil layer will undergo vacuum filtration, storage and identification as described above. For water samples, we will follow the same protocol with one modification. Raw water samples (5 L) will be filtered through a 25 µm mesh screen. Large debris (> 5 mm) will be carefully removed and the remaining material will be rinsed into a clean glass beaker. The rest of this method will follow the protocol previously described.Given that airborne microplastic contamination is very common (Foekema et al. 2013, Torre et al. 2016), we will adopt a series of measures to prevent it as in Li et al. (2015) and Zhao et al. (2017). To prevent contamination from our own clothes a 100% cotton lab coat will be used during all collections and experimental procedures (Van Cauwenberghe & Janssen 2014).Microplastic identification - After extraction, microplastics will be visually sorted and enumerated under a dissecting microscope. This assessment can help identify different morphotypessuch as fibers (slender and elongated), spheres (round, similar to a ball in shape), film (small and very thin layer of larger plastic debris) and fragments (isolated or incomplete part of larger plastic debris that did not fit any of the previous categories). Once the microplastics have been isolated, we will use micro-Fourier Transformed Infrared Spectroscopy (μ-FTIR) to collect a characteristic IR spectrum of the material. Once the spectra are obtained, they can then be correlated to known IR spectra for identification of the specific materials in the sample. The spectra obtained from each microplastic particle will be compared with a library of different polymers in the OMNIC Picta software (Thermo Fisher Scientific) following Vianello et al. (2013).Objective 2 - Assess microplastic degradation and examine how aquaculture gear may be contributing to microplastic contamination in different environments (Years 1-3).In this study, we will assess the spatial and temporal variability of plastic degradation by deploying plastic strips in different areas where shellfish are commonly raised.For this part of the study will use plastic polymers (strips and ropes) commonly used for aquaculture gear including: high-density polyethylene (PE), polypropylene (PP), polyamide (PA), expanded polystyrene (EPS), acrylonitrile butadiene styrene (ABS) and ultra-high molecular weight polyethylene (UHMwPE). Plastic polymers will be deployed and retrieved after 4-wk, 6-m, 1-y, and 1.5-y. At each deployment site we will have temperature and light sensors to record environmental parameters. This part of the study will be performed in Washington in collaboration with Taylor Shellfish and Baywater Shellfish Farms.The plastic polymers will also be studied to obtain the following information before exposure to the field and after removal from the field: (a) weight: a standard microbalance will be used to look at how much mass of the polymer remains; (b) surface topology: scanning electron microscopy (SEM) will be used to look at how the surface morphology changed; (c) porosity: mercury porosimetry will be used to examine the change in porosity of the polymer beyond just the surface of the sample; (d) hetero-nuclear functional groups: µ-FTIR spectroscopy will be used to study if and how the functional groups reacted in the polymers. As mentioned in objective 1, µ-FTIR is particularly useful for looking at hetero-nuclear functional group vibrations and polar bonds such as those contained in polyamide and styrene-butadiene polymers (SBS). As such, µ-FTIR will be specifically used to investigate if the amide linkages in the polyamide and if the nitrile bonds in SBS are altered during exposure to the environment; (e) homo-nuclear bonds: the other remaining polymers high-density polyethylene (PE), polypropylene (PP), expanded polystyrene (EPS), and ultra-high molecular weight polyethylene (UHMwPE) do not contain hetero-nuclear functional groups. As such, changes in chemical structure of these polymers will be studied using Raman spectroscopy; (f) composition: elemental analysis by combustion analysis will allow for the detection in change in elemental composition of the polymers and (g) mechanical properties: a nanoindenter will be used to measure changes in mechanical properties of the microplastics.Polymer characteristics will be used to calculate the rate of degradation of the six polymers and to determine what degradation mechanism (oxidative, hydrolysis or biodegradation) is the predominant in the different locations. The final form of the six polymers will be compared against the microplastics that have been isolated from the shellfish to verify if aquaculture gear is indeed contributing to microplastic contamination in the different environments.

Progress 09/01/20 to 03/07/23

Outputs
Target Audience:1) Formal classroom instruction and practicum experience (upper level undergraduate and graduate students) in the School of Aquatic and Fishery Sciences (SAFS) at University of Washington (UW) 2) Curriculum Development for UW interns 3) Seattle Aquarium: Outreach, Education and Research 4) Plastic workshop for Maritime High school students Changes/Problems:The main difficulty we had wasthe instrumentation and technical support. The iN10 from ThermoFisher that we obtained with this grant did not function properly and we required technical assistance from Fisher several times. Things got worsewhen the University established the COVID vaccination requirement rule. The Fisher technician that provides support to our equipment did not want to get vaccinated and could not come to campus to fix the machine. Also, some instrumentparts were not available due to COVID closures. Our machine continued having several issues and now it has been replaced by ThermoFisher. We lost research time due to these issues. What opportunities for training and professional development has the project provided?I have met with graduate students that are interested in using the iN10micro-FTIR microscope. Upper-level undergraduates and graduate students have used iN10micro-FTIR better understand the origin, sources and transport of plastic and the effects of plastic pollution on marine organisms.Students also received training to operate the micro-FTIR and examine plastic content in water samples. I hosted ~45 students from Maritime High School to show them research on microplastics. The event took place on the UW campus at the School of Aquatic and Fishery Sciences in our dry lab, FSH 236, and our wet lab in Portage Bay Building, PBB 186. We had activities led by members of my team in each lab. The students were split into two groups to avoid overcrowding in the labs. For most of the activities, students actively participate and interacted and asked questions with the activity leaders. Activities included: - How to use micro-FTIR to identify microplastics - Filtering environmental water samples - Picking microplastics from a filter under a microscope - Feeding sea anemones microplastics - Microplastics adhesion by corals experiment - Identifying adhered microplastics on corals under a microscope - Tour of the wet lab experimental systems How have the results been disseminated to communities of interest?My research team and I have disseminated the results of our research in different formats including: classes, publications, student workshops and conferences by graduate students and principal investigator. 2022 International Coral Reef Symposium, Germany 2023 School of Aquatic and Fishery Sciences Graduate Student Symposium, Seattle, Washington, USA What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? We continue our partnership with the Seattle Aquarium to identify the seasonal abundance and types of microplastics in Washington state. We filter samples, develop protocols and have started to analyze filters with water samples. We have created manuals to support mentoring and student training. We performed experiments to assess microplastic degradation and have started to create spectral libraries of degraded plastics.

Publications

  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Phan S, Padilla-Gami�o JL, Luscombe CK (2022) The effect of weathering environments on microplastic chemical identification with Raman and IR spectroscopy. Part I. Polyethylene and Polypropylene. Polymer Testing


Progress 09/01/21 to 08/31/22

Outputs
Target Audience:1) Formal classroom instruction and practicum experience (upper level undergraduate and graduate students) in the School of Aquatic and Fishery Sciences (SAFS) at University of Washington (UW) 2) Curriculum Development for UW interns 3) Seattle Aquarium: Outreach, Education and Research Changes/Problems:The main difficulty we have isthe instrumentation and technical support. The iN10 from ThermoFisher that we obtained with this grant has not been functioning properly and we have required technical assistance from Fisher several times. Things got worsewhen the University established the COVID vaccination requirement rule. The Fisher technician that provides support to our equipment did not want to get vaccinated and could not come to campus to fix the machine. Also, some instrumentparts were not available due to COVID closures. We lost research time due to these issues. What opportunities for training and professional development has the project provided?I have met with graduate students that are interested in using the iN10micro-FTIR microscope. I also taught a seminar class: FSH 511 Marine Plastic Pollution.In this class, students learned about the origin, sources and transport of plastic and the effects of plastic pollution on marine organisms.As a part of the class students learned about the micro-FTIR instrumentation and technological applications to examine plastics in the ocean. Students also received training to operate the micro-FTIR and examine plastic content in water samples. How have the results been disseminated to communities of interest?My research team and I have participated in microplastic oriented conferences: 2021, Nov 2, Pacific Northwest Microplastics Workshop, Seattle Aquarium. 2021, Oct 28,Pacific Northwest Consortium on Plastics teleconference, Oct 28.SOLUTIONS to plastic pollution: from Zero Waste Washington and the Seattle Aquarium on progress in Washington State and on federal legislation from the honorable Senator Merkley (D-OR). Organized by Oregon State University. What do you plan to do during the next reporting period to accomplish the goals?1) Refining protocols and continuing the development of spectral libraries. 2) We will continue extracting and analyzing microplastic samples of bivalves from different areas in Washington state. 3) Wewill develop a website interface to share our protocols with the community. 4) We will disseminate our results and protocols with communities of interest. 5) We will publish our manuscript on plastic degradation

Impacts
What was accomplished under these goals? We have started our partnership with the Seattle Aquarium to identify the seasonal abundance and types of microplastics in Washington state. We have filtered samples, developed protocols and have started to analyze filters with water samples. We have developed protocols and manuals to support mentoring and student training. We performed experiments to assess microplastic degradation and have started to create spectral libraries of degraded plastics.

Publications


    Progress 09/01/20 to 08/31/21

    Outputs
    Target Audience: Nothing Reported Changes/Problems:The main difficulty we have isthe instrumentation and technical support. The iN10 from ThermoFisher that we obtained with this grant has not been functioning properly and we have required technical assistance from Fisher several times. Things got worsewhen the University established the COVID vaccination requirement rule. The Fisher technician that provides support to our equipment did not want to get vaccinanted and could not come to campus to fix the machine. Also, some instrument parts were not available due to COVID closures. We lost research time due to these issues. What opportunities for training and professional development has the project provided?I have met with graduate students that are interested in using the iN10micro-FTIR microscope. I also taught a seminar class: FSH 511 Marine Plastic Pollution.In this class, students learned about the origin, sources and transport of plastic and the effects of plastic pollution on marine organisms. As a part of the class students learned about the micro-FTIR instrumentation and technological applications to examine plastics in the ocean. Students also received training to operate the micro-FTIR and examine plastic content in water samples. How have the results been disseminated to communities of interest?My research team and I have participated in microplastic oriented conferences: 2021, Nov 2, Pacific Northwest Microplastics Workshop, Seattle Aquarium. 2021, Oct 28, Pacific Northwest Consortium on Plastics teleconference, Oct 28.SOLUTIONS to plastic pollution: from Zero Waste Washington and the Seattle Aquarium on progress in Washington State and on federal legislation from the honorable Senator Merkley (D-OR). Organized by Oregon State University. What do you plan to do during the next reporting period to accomplish the goals?1) Refining protocols and continuing the development of spectral libraries. 2) We will continue extracting and analyzing microplastic samples of bivalves from different areas in Washington state. 3) Wewill develop a website interface to share our protocols with the community. 4) We will disseminate our results and protocols with communities of interest. 5) We will publish our manuscript on plastic degradation

    Impacts
    What was accomplished under these goals? We have started our partnership with the Seattl Aquarium to identify the seasonal abundance and types of microplastics in Washington state. We have filtered samples, developed protocols and have started to analyze filters with water samples. We have developed protocls and manuals to support mentoring and student training. We performed experiments to assess microplastic degradation and have started to create spectral libraries of degraded plastics.

    Publications

    • Type: Journal Articles Status: Other Year Published: 2022 Citation: In preparation: The effect of weathering environments on chemical identification with Raman and IR spectroscopy: Part I: Polyethylene and Polypropylene