Graduate Student, Doreen Peters, Finds Microplastic across the Salinity Gradient
We live in a world where plastic is all around us, both where we want it (in inexpensive, lightweight, colorful products) and where we do not want it (in rivers, bays, and coastal areas). Large pieces of plastic are called macroplastics and pieces less than 5 mm in size are known as microplastics. As a graduate student in the Environmental Science and Policy program I had an opportunity to delve into the issue of microplastics in the environment by taking EVPP 581/582, Estuarine and Coastal Ecology/Lab. The coursework covered all aspects of estuarine and coastal ecology from geomorphology and biogeochemistry to plankton, seagrasses, and organisms, as well as human impacts.
Our laboratory experience included getting into the field to collect a variety of types of samples (e.g., nutrients, fish, invertebrates, turbidity, and microplastics), across the salinity gradient from tidal freshwater at the Potomac Science Center (PSC), to brackish water in Edgewater, Maryland, and to salt water in Wachapreague, Virginia. We dodged lightning storms on the PSC trip, but managed to collect some benthic invertebrates from the dock using the Ekman grab sampler. We relied on previously collected samples and data from other trips that had better luck with the weather, including two water samples from Gunston Cove collected by Dr. Dann Sklarew in summer 2018 and destined for microplastics analysis.
On the brackish water trip we were privileged to not only observe Smithsonian Environmental Research Center (SERC) staff perform a periodic fish and macroinvertebrate survey in the Chesapeake Bay, but we also were invited to try our hand at deploying an otter trawl net and measuring/counting the fish that were caught. The first trawl brought in a surprise that confirmed the improper disposal of some human produced objects. It was a challenge to pull the net out of the water with a vehicle tire tangled up in it. The salt water trip was a weekend journey to the Virginia Institute of Marine Science (VIMS) research station. The Captain and our crew of stalwart student researchers led by our skilled and patient professor, Dr. Kim de Mutsert, persevered with our sample collection through a dense and persistent cloud of gnats, as well as rain drops.
The VIMS trip was my maiden voyage to collect water samples for microplastics. Throwing the plankton net and learning to read the mechanical flow meter were all new experiences. The net throw went well, the net went out and I stayed in the boat – a success. The flow meter reading was a little tricky since left handers are not the target audience for flow meters and I ended up with wrong readings, essentially by looking at the backside of the readout. After I made the mental leap of grabbing the meter with my right hand all the subsequent readings were fine. One of the many challenges in studying microplastics is to avoid using equipment containing plastic (to avoid biasing/contaminating the sample with plastic pieces/fibers that may be shed from the equipment). However, there is little choice in nets. The plankton net has nylon (plastic) mesh connected to a plastic cod end with a plastic flow meter counting out rotations that I later used to determine the volume of water filtered through the net. The sample containers I chose are glass with a metal lid that is lined with foil. With each sample location, I gained confidence deploying the net and getting the sample into the container. After three times, I felt like I knew what I was doing. This summer I am pleased to be able to pass on what I learned to the OSCAR students studying microplastics.
The action moved from the field to the newly created GMU microplastics lab, which is set up in the Chemistry Teaching Lab at the PSC. I had stellar support in learning the microplastics laboratory methods since our own Dr. Foster developed the procedure during a sabbatical in 2009 in Washington State. He is a named author on the National Oceanic and Atmospheric Administration (NOAA) Laboratory Methods for the Analysis of Microplastics in the Marine Environment.
The first steps in the method for extracting microplastics from water samples seemed quite benign: 1) sieve the sample, 2) put the sample in the oven to dry, and 3) add reagent and 30% hydrogen peroxide. Then it got a little sketchy with a warning in large bold letters “CAUTION: this solution can boil violently if heated >75°C.” It seemed tame when Dr. Greg Foster performed this step on the first round, but on my first solo try, I was not so sure. This step is designed to oxidize the organic material so that the microplastics stand out under microscopic examination. The goopiest samples (those with globs of zooplankton and bits of vegetation) certainly did bubble, but it was more like cooking pasta. It can boil over if you do not pay attention, so I paid attention and all eight of my samples went well.
The next steps included 4) adding some salt (research has found that table salt contains microplastics, so ideally the salt should be sieved before use) to the sample and pouring it into a density separator then give it time to allow the microplastics to float to the top, 5) drain off the settled solids, sieve the remaining sample and then 6) use the stereo microscope to identify and collect the microplastics – easier said than done. Microplastics are small (< 5 mm) and many things like plant roots, undissolved salt, and vegetative fibers look rather plastic-ish. The trick is developing an eye for microplastics, using the forceps to test uncertain pieces (by pressing on them and if they break they are not microplastics), and trying not to bias the sample by focusing on the more colorful (green, yellow, red, blue) pieces that catch more of your attention than the black, gray, white, and the hardest to spot, clear pieces. I look forward to trying out GMU’s new Fourier transform infrared (FTIR) microscope, which can positively identify types of plastic, to see how accurate I was in identifying the small bits I collected as microplastic.
This experience has been exceptionally valuable to me as I progress towards preparing my dissertation proposal. I feel much better informed in what is doable; what problems may crop up; how much equipment, materials, and boat time cost; and what an issue microplastics may be in the environment – every one of my eight samples collected from across the salinity gradient contained microplastics.
OSCAR Student, Chris Martin, Works on Food Web Analysis
My absolute favorite part of the OSCAR research experience has been putting together the data collected and analyzing it for meaning. In this photograph, I was presenting the culmination of the entire summer’s research project at the Potomac Environmental Research and Education Center (PEREC). It was such an amazing experience to be able to perform my own research, connect the dots of all the data, and be able to say, “This is what I now know about this research that was unknown before”. It was truly an awe-inspiring feeling! (Even if it left me with 20 more questions than I had had before).
And now for a bit of shameless ego stroking. I was particularly proud of the food web I created in this photograph. This specifically filled me with pride as it combined several different avenues of research into a singular cohesive, meaningful piece. I took the results from the stable isotope analysis and combined them with the results of the stomach content analysis to create a food web. This food web displayed input percentages of carbon and nitrogen from external sources into the aquatic ecosystem, calculated the specific trophic levels of each organism, and connected each organism to its consumer. By doing this, we can now see what is coming into the food web from where, and how it is moving through the food web once it has entered. To me, that is pretty darn cool.
OSCAR Student Angelica Frederick Learns About the Fate of Pharmaceuticals
One of my long-term goals has always been to do something for the community. I spent years trying to figure out the best way to give back. Eventually, I realized that science would be a great way to contribute. After my stint in the US Army, I went on to pursue my undergraduate degree in Chemistry. I am projected to graduate from George Mason with my Chemistry B.S with a concentration in Analytical and Environmental Chemistry. I took a class called Aquatic Chemistry with Dr. Foster, and it has inspired to me to learn more about water quality and to pursue research.
When I saw this summer research project “Fate of Pharmaceuticals and Personal Care Products in the Potomac River” funded by OSCAR on a flyer in Exploratory Hall, I knew I wanted to be on the Chemistry team. For the past seven weeks, I have been in the chemistry laboratory at the Potomac Science Center in Woodbridge doing sample preparation. In our chemistry team, we were split into three different groups each with two people. Each group was responsible for prepping specific samples that were collected by the ecology team. The end goal is to be able to identify and quantify the very small amounts of various PPCPs (Pharmaceuticals and Personal Care Products) from the samples. The target area in the Potomac River we are focusing on is Gunston Cove. My laboratory partner, Robert, and I were assigned to do sediment, submerged aquatic vegetation, and clam (Corbicula) samples. Team one was assigned water and plankton, and team three was assigned fish.
Every Monday, the Chemistry and Ecology team has met in a lecture room where we have been given various lectures from our advisor. On other days, the beginning of my day has usually started with a Chemistry team meeting with Dr. Foster and Dr. Huff and the research assistants, Tovga Haji, Tabitha King, and Jonathan Raisigel. It is a way for each group to communicate with each other. Often, the meeting is where we have been brainstorming ideas to solve problems that come up.
After the morning lecture or team meeting, I usually spend the rest of my day in the laboratory. The laboratory is a busy place with many different things going on, which was overwhelming at first. However, with the help of the research assistants and the graduate students, we were able to integrate seemingly over time. They were instrumental in teaching us the extraction methods for each type of sample that they have helped refine over time.
My journey in the laboratory began with the samples collected from Gunston Cove. We used a refined mesh screen, which in technical terms was a 500 µm sieve, to separate out the larger particles in the sediment samples. For clams, we start with working with the ecology to team to harvest the clams from their shells, which involved a hammer and lot of arm strength and patience. As someone who had a strong aversion to seafood, this was the most challenging part. We would then take the clams and used a homogenizer to blend the clams to a uniformed consistency.
The submerged aquatic vegetation (SAV) was an adventure because we found that it was impossible to get it to a uniformed consistency using the homogenizer. However, I recalled that for smoothies, I would use the blender to blend fresh kale and celery, which always worked amazingly. So, we took a regular blender that was used for the fish samples and used it to blend our SAV samples. Success!
Once all the samples were homogenized or pre-sieved, as in the case for the sediment samples, we aliquoted out the various samples we needed. For experiments, it is always good to have at least three replicates. Standards were used in the samples to analyze the recoveries. We then performed the extraction using the QuEChERS (“quick, easy, cheap, effective, rugged, and safe”) method, which consisted of adding salt packets into the samples, a lot of shaking, vortexing, centrifuging, and transferring. They were then evaporated using a machine called the “TurboVap” with nitrogen gas. Those evaporated samples were then transferred into a smaller vial filtered, and then transferred into autosamplers.
Mind you, as Robert and I were doing our own extractions with the help of Tovga, the rest of the lab was also in full swing. We had Chau and Sabrina working on the water samples using solid-phase extraction (SPE). If you are interested in seeing the process for setting up the SPE for water samples and other things we have done: click here. And then, for a nice smelly treat, we also had Alex and Lisa homogenizing the fish samples and using the QuEChERS method to extract. And, remember that we also had to share the laboratory with the graduate students who has their own projects. During this time, we also had a faculty member mentor a high schooler who was also working in the lab. Space and resources were limited, and a lot was going on. The one thing you should always think about while working in a laboratory is safety and how you can contribute to the lab. So, on top of washing all glassware and supplies we have used, we also tried to wash other people’s as well. Everyone the lab has been so helpful and really cares about the projects going on and the science, which has really inspired me to do well.
While the process may have seemed mundane, there were a lot of hurdles to go through. One of the main issues my team encountered was the fact that we did not have enough clam sample for our experiment. This is when I was able to explore science out of the laboratory and a classroom! In my quest for more clams, I coordinated with a member of the ecology team, and we both set out to sample for clams. Outfitted in waders, which are waterproof overalls, he taught me were I would have a high probability of finding clams and how to go about it. I would have to say that it was a daunting process at first because I did not expect the bottom of the Potomac River to be so muddy. At one point, I thought I would be stuck in the mud because I was about knee deep into the mud even though we were so close to the shoreline. Eventually, I got the groove of things and had a lot of success finding a lot of clams.
Samples in the autosamplers meant that the sample prep was complete, and our focus then turned to sample analysis. The star of this process is the Potomac Science Center’s very own LC-MS/MS, which we have named “Keiko.” To put it simply, the liquid chromatograph part of Keiko separates out the chemicals in each sample, and their retention time can be used. Keiko’s mass spectrometer is the part that measures the masses within a sample. The data that comes out of Keiko can be used to identify and quantitate different chemicals.
In our case, these are the chemicals that is widely found in pharmaceutical and personal care products. Some of these chemicals you may know with such as ibuprofen, amphetamine, penicillin, etc. Currently, this portion of the project is still in progress. Considering that I have never directly worked with an instrument like Keiko, I am learning a lot of new things such as setting up new methods and learning the software. The learning curve is steep, but Dr. Huff and Michael Gable, who is a graduate student has been helping us immensely.
There were some more hurdles we had to face because a portion of my team’s samples were not showing expected results. Thank to Dr. Foster’s PhD student, Arion Leahigh, we were able to move through this issue and resolve it because she gave us her extra samples from the same sample trip. However, part of research is discovering how to work with the data that comes out.
This was a major lesson for me having only been in a class laboratory setting where experiments are designed to always go smoothly. I learned that the path is not always clear, and you just have to problem solve and move forward with your head high. I am thankful to OSCAR and the PEREC family who have given me this opportunity to grow as a scientist. I am looking forward to analyzing our results and seeing what our data can tell us about our samples. It will be interesting to see how it connects with the ecology team’s project and how it can contribute to society.
Re-Discovering Science: A Story By OSCAR Student Alex Marchesani
I’ve always been fascinated with science from a very young age. Tearing through books on biology and trying to memorize as many facts as I could about various other natural phenomenon. As I progressed through my high school career, I found myself enamored with microbiology specifically. Discovering and learning all the various eccentricities and distinctive features of the different classes and species. I now find myself entering my second year here at Mason. I have taken several chemistry and biology courses thus far. Throughout my first two semesters, I began to feel something I had never felt before in regards to science. I felt bored. Not because classes were too easy (CERTAINLY not that). But somehow, the luster was fading. The endless facts and equations were becoming taxing, rather than energizing like years past. I began to fear: “Have I chosen wrong?” “I’ve never had passion for another subject, can I really change?” “What if science isn’t for me?”
I had two basic options for what to do over the summer, continue working at my local country club and wake up at 5:30AM every day to work the grounds of the golf course, or find an internship. Because you are reading this now, I think you can realize which choice I went with and why. As for my internship options, I went with science, the thing I was afraid I was losing passion for. I applied to several opportunities and sent my various cover letters and other requirements. As time passed, I began to think about abandoning everything and saving as much money as possible for the summer and work at the golf course. One day, I received an email inviting me down to a place I had never been before, the PEREC facility. The email stated that my potential mentors were impressed with my credentials and had invited me for an interview. Although I was still unsure what I wanted, I figured it would be good to explore all my options.
Upon getting to PEREC, I found myself taken aback and surprised. I had never visited a research center before, certainly not one as new and fresh and this. My curiosity piqued, I sat and waited for my interview and tour to begin. The tour consisted of informing me about the things we would be doing. Mainly, gathering data from sediment, water, and fish samples and processing them for analysis with a mass spectrometer. A mass spectrometer is an instrument which someone my age rarely gets the chance to work with. Afterwards, I sensed a slight shift in things. I have just recently come to realize what exactly it was. It was a fire inside me. Burning and raring to be a part of something I had never been before. The hunger to consume knowledge and the craving to understand how to make discoveries of my own had been reignited. I quickly accepted the offer to work at PEREC for the summer.
Since then, I have been able to peak behind the curtain so to speak. To see all the goings-on that culminates in publications and conventions and posters. What I observed was not quite what I had expected. The people here are not cold and calculating scientists, though they can be if the need arises, they are warm, jovial, and always willing to assist you. I am, to my knowledge, the youngest person working in my chemistry lab (I’m 19). Thus, I was quite anxious about working here with the least amount of previous experience, and the fact that I’m not a chemistry major like my colleagues (which I get teased for constantly). Despite a few awkward moments and lots of uncertainty in the beginning weeks, I have since developed friendships with my peers that I normally never would in this situation. I am a very introverted and reserved person, it takes a lot for me to engage others in conversation. However, because so much of the lab work was over my head, at least at first, it forced me to ask people for help, something I’m not very good at. In doing so, I started to feel a sense of “belongingness” that I have never felt towards a work environment before. I feel like part of a team of genuine friends here in the PEREC chemistry lab. The kinds of friend who will help each other with preparing samples of fish (which smelled awful) with no complaint, and then turn around and invite you to trivia night or to a barbecue for the weekend.
Not only do I feel comfortable at PEREC, I also am extremely proud of the work we do and the things we hope to accomplish. I suppose after about two pages, I should probably inform you all what it is we actually do here. My group, as part of a program run through the OSCAR office, is focusing on processing and analyzing fish samples for the purpose of validating the mathematical model we are using to construct a food web of the Potomac River. The other two parts of our project deal with water and sediment samples. The data from these samples will be input into our mathematical model (called Kabam believe it or not) and will deliver us with estimations about what levels of chemicals may be present in many organisms. The kinds of chemicals we are interested in are called micropollutants. Specifically, we are looking into pharmaceuticals and personal-care products (PPCP’s). Many of these compounds have been found in concentrations of just a few parts-per-million. While that may not sound like much, PPCP’s include things like estrogenic compounds and antibiotics. These sorts of chemicals can have major effects on ecosystems even in such small amounts. While the exact effects of many PPCP’s, as well as the sources where they originate, are not fully understood, our work here at PEREC is helping make steps towards forming more comprehensive knowledge about the Potomac and aquatic ecosystems overall.
OSCAR Students Collect Fish Data
My name is Brian Kim and I am a member of the OSCAR fish team at the Potomac Science Center (PSC). Over the course of this summer, I have been going out to Gunston Cove and Hunting Creek located within the Potomac River to collect fish species. My 
research focuses on assessing the diet items of the fish species and determining if there is a significant difference between the two locations as well as any differences in submerged aquatic vegetation (SAV) and non-aquatic submerged vegetation (NSAV) areas within each location. The most enjoyable part of this experience has been the process of the actual fish collection. There have been two separate trips for each location where we would use a series of seine nets, fyke nets and trawls to catch fish from as little as 10 mm to as large as 530 mm. The trawls were the most thrilling method because we usually caught the largest specimens during each trial. As each trawl was hauled onto the boat, there was a rush of excitement as to see which specimens we caught and how many. Once we had recorded all required data on each fish, the samples were taken back to the Potomac Science Center to begin the process of answering the question “What exactly has each fish species been eating?”
The majority of my time at the PSC has been spent dissecting the stomachs from the fish samples and opening them up to see what was inside. Each stomach from the fish species differed in some way both internally and externally. The most exciting stomach contents that I have found so far came from a yellow perch that had eaten five juvenile fish within the last day it was caught. I was able to identify the five juveniles as three white perch and two spottail shiners. There have been various different prey items in 
the stomachs such as amphipods, chironomids, gastropods and aquatic insects that must now be analyzed. Using the stomach contents from each fish species, I will be able to determine which prey items are the most impactful as well as how location, habitat and fish species differ in diets. My time at the PSC and with OSCAR has been the most exciting summer during my undergraduate years and I hope that in the future I will be able to return and continue to work on related projects.
Micropollutant Research by Current OSCAR Student, Sabrina
My name is Sabrina. I am a current student in OSCAR program for summer 2018. My work is about investigation and fate of emerging contaminants in Gunston Cove of Potomac river in Alexandria. We extract micropollutants from water, sediment and fish samples and use liquid chromatography-mass spectrum (LC-MS/MS) method to analyze the extracts. We solid phase extract the micropollutants from water samples and use QuEChERS to extract them from sediments and fish and then run the extracts in the LC-Ms/MS instrument and we analyze the results. After that, we apply the KABAM model to predict the bioaccumulation of chemicals in organisms’ tissues. 
More interestingly, the work is collaborative, and this gives me a good opportunity to interact with people with different background and be involved in group work. This research is the best experience in my academic pathway because I feel that I absorb lot of information related to my field and I am surrounded by a huge, friendly and experienced team working with me in the lab. Moreover, this research involves lot of data analysis and use a lot of literature resources where I learn more about my research and related topics and I develop skills in data analysis and time management. I learn from every single step of the process, strengthen my experience in lab work, interact with people with high experience and learn to work under pressure, which I can apply in my daily life as well. I, also, should admit that this research is a guide for me to pursuit my graduate program in the same field of study.
Outreach Events During Summer Impact Research for Undergraduates
In addition to doing our research, we have been able to do some local outreach events, teaching kids about the water and the invertebrates in it!
The second outreach event was at Occoquan Regional park for their grand opening! This day was all about turbidity (how much dirt is in the water, and how clear it is) and also what kind of zooplankton are in the water! There was a microscope w a daphnia (a type of zooplankton commonly found in the Potomac River), and jars with different turbidity levels to show the difference!
OSCAR video: “What’s in Our Water?”
George Mason (https://www2.gmu.edu/) undergraduate research, led by principal investigators Amy Fowler and Kim de Mutsert, who are researchers at the Potomac Environmental Research and Education Center (Perec.gmu.edu). In the summer of 2017, this team project looked at the effects of micropollutants on the Potomac River watershed. Students and faculty discuss some of the results of the study and the future of this work. Projects were funded by the Students as Scholars at Mason (https://oscar.gmu.edu/) as well as the Patriot Green Fund (https://green.gmu.edu/patriot-green/) , and the videos were produced by graduate student, Chelsea Gray, thanks to the Virginia Sea Grant (https://vaseagrant.org/).
