Perspectives of Aquatic Toxicology/Chapter 3: Micro-plastics: An Emerging Pollutant in an Aquatic Ecosystem

= Chapter 3: Microplastics: An Emerging Pollutant in an Aquatic Ecosystem =

Introduction to Microplastics
If you were to causally glance at your surroundings, chances are you would see a multitude  of products that are made of, or utilize plastics. Plastics are long chains of polymers synthesized from both organic and inorganic materials such as carbon, hydrogen, silicon, oxygen, and chloride which are usually acquired from natural resources such as natural gas, oil, and coal (Ivar do Sul et al., 2013). Plastic is considered to be a fairly recent invention. In 1907, the first plastic Bakelite was synthesized. However, it wasn’t until the 1940s that plastic production began in earnest (Cole et al., 2011). Copious amounts of plastics have been synthesized and released into the environment. One of the major draws of plastic is its durability, which may ultimately lead to its downfall. Tons of plastic pieces find their way each year into various watersheds, and subsequently the oceans. Many studies have demonstrated the hazards of plastics to aquatic wildlife. (Figure 2 & 3) These studies focus on macroplastics: plastics that are easy to see with the naked eye. However, plastics come in a variety of sizes: macroplastics, microplastics, and nanoplastics. There is no standard definition for microplastics, and thus the term “microplastic” is controversial. The term does not refer to the standard micro unit as seen in the International System of Units (1-999 μm). A microplastic can be a particle that has a diameter of < 10 mm, or < 5 mm, or 2-6 mm, or <1 mm (Cole et. al., 2011) (Figure 4). Nor is there a set definition for nanoplastics (plastic particles that are smaller than microplastics) and that absence further complicates matters. Again, the term “nano” in nanoplastic does not refer to the nano size typically seen in a laboratory setting. For this chapter, the term microplastic means any plastic particle that is < 5 mm in diameter but greater than 100 nm.  The term nanoplastic refers to plastics that are between 1-100 nm (Jovanović, 2017).

Types of Microplastics in the Environment
There are two major categories of microplastics: primary and secondary. Primary microplastics are plastics manufactured in the microplastic size range and often used in cosmetics, facial cleansers, facial exfoliants, air blasting media, and even medicine. Occasionally, these microplastics are called micro-beads or micro-exfoliants. Primary microplastics can also include virgin plastic production pellets. Secondary microplastics are microplastic particles that have fragmented from macroplastics (Cole et al., 2011). Common synthetic plastics used today include polypropylene, polyethylene, polystyrene, polyvinyl chloride, polyethylene terephthalate, and as well as others (Ivar do Sul et al., 2013) (Figure 5).

Quantification and Location of Microplastics Worldwide
As previously mentioned, the first major plastic, Bakelite, was produced in 1907; however, the mass production of plastic containing compounds began much later in the 1940s (Cole et al., 2011). Since then, the amount of plastic produced worldwide has continued to increase at a rapid rate. In 2017, it was estimated that over 350 million metric tons of plastic were produced, and since then plastic production has increased even further (PlasticsEurope, 2018). Of the plastic manufactured each year, roughly up to 10% of it will subsequently end up in aquatic environments where it will reside for extended periods of time. Thus, the quantify of plastic accumulating in aquatic environments is continuously increasing. By 2050 it is estimated that the amount of plastic present in the oceans will either be equal to or greater than the total weight of fish in that same environment (Jovanović, 2017). In 2014, van Sebille, et al. estimated the amount of microplastics on the surface of the ocean to be between 15-51 trillion particles, together weighing between 93-236 thousand metric tons.

Microplastic particles have been located worldwide, even in areas that should be devoid of plastics. Additionally, the type of microplastic present in such environments does not substantially differ from region to region: it is possible to find polystyrene microplastics everywhere (Koelmans, et al., 2016). Microplastics are present at various depths in the water column. Typically, microplastics such as polyethylene and polypropylene are very buoyant and may remain on the surface of water. However, it is also possible for other microplastics such as polyvinyl chloride to be less buoyant or for the buoyancy of the particle to decrease or increase due to surface growth of microbial films. These microplastics particles may then be suspended in the water column or be located on, or in the sediment (Ivar do Sul, et al., 2013).

Models used to estimate the prevalence of microplastics in the ocean often focus on microplastics located on the surface of the water. As such, they capture a brief snapshot of microplastic presence in one specific area of the water column. However, vertical mixing of particles in the water column may occur as a result of wind (Ivar do Sul, et al., 2013.) Additionally, more microplastic particles enter into an aquatic ecosystem after storms, flooding, or other types of extreme weather (Cole, et al., 2011). This makes it difficult to obtain an accurate image of how many pieces of microplastic are present on the ocean surface. Nevertheless, there are areas more or less abundant in the number of microplastic particles compared to other areas. For instance, there are fewer microplastics present in the tropics above 45ᵒN and below 45ᵒS, and the remote coastline off Western Australia. The highest concentrations are found in centers of subtropical gyres located in North Atlantic and North Pacific regions. The highest concentration in subtropical gyres is estimated to be 108 particles km-2. The highest counts of microplastics are located in Mediterranean and North Pacific basins while microplastics present in the North Pacific basin contain the largest total mass (Figure 6). This is likely due to the vast area of the North Pacific basin and the large quantity of plastic waste entering the ocean from the coastlines of the United States and Asia. Ultimately, the majority of microplastics lie in regions of low concentration. Further data is needed to verify these estimates, especially in less analyzed regions such as South America (van Sebille, et al., 2015).

Methods Used to Study Microplastics
Various methods are used to quantify the number of microplastics present in an aquatic environment. These include beach combing, sediment testing, biological sampling, marine observational surveys, and marine trawls. Beach combing involves collecting, identifying, and quantifying all litter items on the shore of a specific coastline region, and If done periodically it can illustrate the debris accumulation during a specific time frame. This process is best suited for quantifying macroplastics and occasionally larger microplastics. For example, it is possible to quantify plastic resin pellets called Mermaid’s Tears while beach combing. Sediment testing involves taking sediment samples from the sediment of an aquatic environment and quantifying the amount of microplastics in benthic regions. Biological sampling involves quantifying the amount of microplastics consumed by aquatic organisms (See Section C for examples). Marine observational sampling is conducted by divers or onboard observers who record the size, location and types of plastic observed in the water. However, the small nature of microplastics allows them to be unobservable and ergo, undetected. Marine trawls constitute one of the more popular methods used to capture, identify and quantify microplastics where fine meshes or nets are dragged behind boats. It is common to use a plankton net in these studies; the size of the net can vary between 0.1 mm to 0.5 mm (Cole et al., 2011; van Sebille et al., 2015).

Microplastics are typically quantified either by sediment testing or marine trawls. It is important to filter out impurities from the samples and to promote the migration of microplastics to various solution surfaces after samples have been collected (Figure 7). Water samples are run through a coarse filter to remove microplastic particles and other larger contaminants. Meanwhile, sediment samples are slurried with saline water to promote the migration of microplastics to the sample’s surface. Additionally, mineral salts may be added to either the water sample or the slurried sample to increase water density, as more microplastics may float to the surface of a sample when water density is greater. Evaporation is then commonly used to concentrate the microplastic particles at the surface of the sample. Subsequently, regions of the samples with microplastics will have been isolated and can be removed for quantification. Following removal, microplastics in surface water samples can be stained with a lipophilic dye such as Nile Red in order to increase ease of visualization. Microbiota that are present in the solution or on the microplastic pieces are not stained with the dye. Sometimes hot dilute mineral acid is used to remove any additional biomass. Finally, the microplastic pieces identified and quantified using various types of microscopy such as Raman spectroscopy, FTIR spectroscopy, electron microscopy, and optical microscopy (Andrady, 2011).

Ingestion of Microplastics
Plastics are ubiquitous: it is not uncommon to find bits and microplastic pieces in and around aquatic wildlife. Haunting images of turtles trapped inside plastic have invaded the media for years. Other images have shown tons of plastics almost spilling out of the guts of aquatic birds or whales. (Figure 8) In fact, at least 44% of all marine bird species have ingested plastics (Andrady, 2011).

Both macroplastics and microplastics have been ingested by a variety of aquatic wildlife at different trophic levels. Microplastics do not discriminate against species (although some species may discriminate against microplastics--e.g. selective filter feeders) and have been found in and around both invertebrates and vertebrates. (Table 1)

Table 1: Examples of species that have ingested microplastics

Ingestion of Microplastics by Invertebrates
Colonies of microscopic organisms can grow microbial biofilms on various microplastics, and these biofilms can accumulate on microplastics in less than a week. As a result, species of invertebrates and algae have been able to grow on the surfaces of these particles (Cole, et.al, 2011). Additionally, insects such as Halobates micans and H. sericeus use plastic pellets as oviposition sites (Majer et al., 2012; Goldstein et al., 2012). For example,  the numbers of eggs of H. sericeus juveniles and adults in the North Pacific Ocean were significantly and positively correlated with the number of microplastics present. Furthermore, 34% of all plastic pellets found in the western Atlantic had eggs attached to them (Barnes, et al., 2009).

Laboratory experiments have been used often to highlight how invertebrates ingest microplastic particles. In 2004, the first experiment to test microplastic ingestion by barnacles (Semibalanus balanoides), amphipods (Orchestia gammarellus), and lugworms (Arenicola marina) was performed, and demonstrated that these three species could ingest microplastics (Thompson et al., 2004). Other studies have tested various mollusks, crustaceans, annelids, and echinoderms. For example, one of the more commonly studied mussels in regard to microplastic ingestion is Mytilus edulis. This species ingests and accumulates microplastics within 12 hours of exposure. Interestingly, not only were microplastic pieces found in the intestines, they were also present in the gills and hemolymph of the mussels (Browne et al., 2008; von Moos et al., 2012). Additionally, microplastics found in these mussels were later transferred to a higher trophic level: Carcinus maenus. As a result, microplastic particles have been found in the hemolymph and various other tissues of these crabs (Farrell and Nelson, 2012). Thus far, studies of microplastic ingestion by benthic crustaceans have been limited. However, 83% of sampled lobsters in the Clyde Sea had microplastic particles in their stomachs. Additionally, lobsters were shown to ingest microplastics within 24 hours of being exposed (Murray and Cowie, 2011). Microplastic ingestion has even been identified in humboldt squids (Dosidicus gigas), and sea cucumbers (Holothuria) have been known to preferentially feed on nylon and PVC fragments over sediment grains (Braid et al., 2012; Graham and Thompson, 2009). Finally, many types of zooplankton have also been studied for the possibility of microplastic ingestion, and It was confirmed that many taxa do indeed ingest the particles (Cole et al., 2013). The possibility of ingestion of microplastics by various types of invertebrates seems endless.

Ingestion of Microplastics by Vertebrates
Microplastic particles have been discovered in the guts of various vertebrates since 1972. Some of the first particles detected in larvae and juvenile fish were found in Psuedopleuronects flounder, while some of the first particles detected in adult fish were found in Morone america and Protonus evolans (Carpenter et al., 1972; Hoss and Settle, 1990). The potential for possible ingestion of microplastics became concerning when a study (Boerger et al., 2010) found synthetic fragments in 35% of planktivorous fish located in the North Pacific Central Gyre. A study (Lusher et al., 2013) that evaluated demersal and pelagic fish in the coastal waters around the United Kingdom found further evidence of microplastic ingestion among various species, with 36% of the species tested having ingested fibers and microplastic fragments. Additionally, mesopelagic fish located in the North Pacific were found to have ingested microplastic particles and fibers (Davison and Asch, 2011). To further illustrate the enormity of the problem, 40% of lantern fish tested in the Marianna Islands--a region that contains less microplastic particles--were found to have ingested microplastics, further illustrating how ubiquitous microplastics are in terms of ingestation (Van Noord, 2013).

Microplastic ingestion by vertebrates is not limited to the marine environment. Studies have found these particles in fish living in estuaries as well. For example, a study conducted in the western South Atlantic Ocean reported that mojarras (Gerreidae), estuarine drums (Sciaenidae), and catfishes (Ariidae) all ingested microplastics. All three species feed on or just below the surface of the sediment, and the most popular ingested microplastic was blue nylon thread. They most likely ingested the microplastic particles accidentally through normal suction feeding, eating contaminated prey, and/or by actively feeding on plastics with biofilm (Ramos et al., 2012; Dantas et al., 2012; Possatto et al., 2011). Microplastics have been found ingested by fish in all areas of the world and at all levels of the water column.

Although not commonly thought of as aquatic vertebrates, shore birds have also been known to ingest microplastics. In fact, the ingestion of plastics by birds has been studied for the past four decades, although many of these studies did not distinguish between microplastics and macroplastics. Microplastic pellets have been found in the guts of migratory petrels--shearwaters and prions--in the Atlantic and south-western Indian Ocean among others since the 1980s. Plastic pellets and other plastic fragments were found in 80% of two Fulmarus glacialis colonies in the Canadian Arctic (Provencher et al., 2009). Additionally, the Fulmarus glacialis species has been monitored in both the North Sea and the Netherlands for the past 30 years. During this time, it was discovered that while the number of plastic pellets decreased by 50% in 20 years, the number of plastic fragments tripled. Interestingly, juvenile Fulmarus glacialis ingested more plastic than the adults. It was also noted that there was an increased presence of ingested microplastics in areas near highly industrialized areas specializing in either shipping or fishing (Van Franeker et al., 2011; Kühn and van Franeker, 2012). Furthermore, around 90% of all fulmars sampled in the North Pacific Ocean had ingested microplastics (Avery-Gomm et al., 2012). Microplastics were found ingested by birds located in Iceland as well; but overall, fewer had ingested the particles. Thus, it has been hypothesized that birds further north are less likely to be contaminated by microplastics because there are fewer microplastics available on the surface of the water in these regions (Van Franeker et al., 2011; Kühn and van Franeker, 2012). This hypothesis was also proposed by a study done on the ingestion of microplastics by Uria lomvia in Nunavut, Canada in which 11% of the birds investigated had ingested microplastics; however, the authors hypothesized that the low number may be due to these birds feeding below the water surface and are therefore less likely to ingest floating plastics (Provencher et al., 2010). It is interesting to note that birds never exposed to marine environments may ingest microplastics as well. For example, 83.5% of fledgling cory shearwaters (Calonectris diomedea) were found to have ingested microplastics. These chicks were exposed to pieces of microplastic by eating the regurgitated food from their parents (Rodríguez et al., 2012). Other bird species known to ingest microplastics include Larus glaucescens, P. nigripes, Phoebastria immutabilis, among others (Lindborg et al., 2012; Gray et al., 2012; Avery-Gomm et al., 2013).

Research related to ingestion of microplastics by marine mammals is limited; however, it has been shown by analyzing their scat that marine mammals do indeed ingest microplastics. For example, fur seal scat (A. gazelle and Arctocephalus tropicalis) collected on Macquaire Island contained plastic fragments and pellets that were 2-5 mm in size. These plastics were believed to have come from the animal’s prey that ingested microplastics present in or on the water (Eriksson and Burton, 2003). Future studies will likely highlight the presence of microplastics found in aquatic mammal intestines.

Degradation of Microplastics
Macroplastics eventually become microplastics, and microplastic pieces eventually degrade into nanoplastics. It is estimated that it will take approximately 320 years for a 1 mm sized microplastic to degrade into a nanoplastic within a marine environment (Koelmans et.al, 2015). Degradation is the chemical process by which the average molecular weight of a polymer is reduced. Plastic degradation to any size will eventually weaken the material and the particles may become so weak they fall apart into a powdery substance subsequently degraded further. Plastic is said to have mineralized when all carbon present in the polymer is converted into CO2: mineralization is the goal of degradation. The process of degradation can fall into one of five major categories: thermal degradation, photodegradation, thermooxidative degradation, biodegradation, and hydrolysis.

Different types of Degradation
Thermal degradation is the degradation that occurs via high temperatures. This is not considered a mechanism by which microplastics degrade in the environment. Photodegradation is the process by which light, usually sunlight, degrades a material. UV-B is the primary component in photodegradation. It is also considered to be one of the fastest methods of degradation for microplastics, working best for particles that are exposed to air or lying on a beach; however, the degradation rate decreases when the microplastic particles enter the water. Additionally, photodegradation is often the precursor to other types of degradation, and it is often followed by thermooxidative degradation. Thermooxidative degradation is the process by which particles are slowly broken down by oxidation in moderate temperatures--a process which can continue to degrade the microplastic particle as long as oxygen surrounds the microplastic. The quantity of oxygen present decreases in an aquatic environment, and a lack of oxygen and colder temperature will often decrease the degradation rate of a microplastic. Additionally, the growth of organisms on a microplastic can decrease the rate of degradation. This process is different from biodegradation. Biodegradation is the process by which living organisms degrade microplastics and it is typically carried out by microscopic organisms; however, these organisms are rare in nature. Finally, hydrolysis is the process by which water is used to degrade a compound; it is one of the slowest in the marine environment and rarely occurs. Once again, photodegradation and thermooxidative degradation are the most common ways in which microplastic particles are degraded into smaller pieces and eventually into nanoplastics (Andrady, 2011).

Equilibrium Partitioning Equations
It is helpful to discuss the concept of equilibrium partitioning before discussing the relative toxicity of each microplastic. Chemicals often partition in either organic matter or water, as discussed in previous chapters. Additionally, toxicants can have sorption to the plastic particles themselves. However, microplastics do not follow typical equilibrium partitioning concepts: they do not have a log Kow. Instead, specialized equilibrium factors for microplastics must be used to determine how a toxicant may partition. There are many different factors that affect the overall kinetics of the equilibrium partitioning process such as the equilibrium between water-biota, water-sediment, and water-plastic. The equilibrium partitioning equation used to determine if a toxicant will sorb to a piece of microplastic (KPL) is defined as:

KPL = CPL/Cw

where CPL (μg/kg) is the concentration of the toxicant in plastic and CW is the concentration of the toxicant in water. If KPL > CPL/CW, the toxicant can desorb from plastic to water, but if KPL <CPL/CW, the toxicant will sorb from water to plastic (Koelmans, et al., 2016). Transfer of chemicals to plastic can also be defined more precisely as:

dCPL/ dt= k1CW − k2CPL

in which k1 and k2 are first-order rate constants related to the thickness of the undisturbed boundary layer surrounding the microplastic and the ability of a chemical to diffuse in an aqueous environment, in which t= time, and in which CPL /CW are the same values as previously discussed. This equation differs from the previous equation by factoring the unbound layer that surrounds the microplastic. Both equations describe whether a compound is more likely to sorb from water to the microplastic or from the microplastic to water. Thus, microplastics can serve as a way to clean up toxicants from water or a possible manner by which toxicants enter the water. This concept of equilibrium portioning also occurs inside an organism. (Figure 9). The process of toxicant movement from one source to another will continue until equilibrium is reached. Bioaccumulation is the ability of a chemical to continuously accumulate inside an organism. Koelmans, et al. modelled the bioaccumulation of hydrophobic compounds from an environment containing plastics to an organism as follows:

dCB,t/dt = kdermCW + IR(SFOODaFOODCFOOD + SPLCPLR,t) − klossCB,t

where Kderm CW is equal to the dermal (including gills) uptake of a chemical from the water, the second term reflects the uptake of the chemical from the diet and the exchange of that chemical with plastic particles, and the final term refers to the overall loss of the chemical as a result of elimination and egestion. More specifically, Kderm (L x kg x d-1) and kloss (d-1) are first order rate constants while IR (g x g-1 x d-1) is the mass of food ingested per organism dry weight and per time. Additionally, aFOOD refers to the absorption efficiency from the diet while SFOOD and SPL refer to the mass fractions of plastic and food in ingested material. Finally, CFOOD refers to the contaminant concentration that occurs from food to the organisms itself. Once again, this equation would highlight the ability of a compound to bioaccumulate inside an organism. Koelmans, et al. (2013a, b, 2014b) assumes that the degradation of microplastics will not occur during their journey through the gastrointestinal tract. Thus, the ability for the compound to transfer from a plastic particle to the gut is dependent on the concentration of microplastic and biota lipids; the kinetics of this are as follows:

CPLR,t = [(k1GCPL − k2GCL,t) / k1G + (MPL/ML)k2G] * (1 − e − [k1G + (MPL/ML)k2G]GRT

where k1G and K2G are first-order rate constants which describe the transport of the compound between plastics and lipids inside the gastrointestinal tract. Next, GRT is equal to the gut residence time while CPL and CL,t refer to the chemical concentrations in the ingested plastic and the biota lipids at the moment when the microplastic is first ingested. Finally, MPL and ML refer to the mass of the plastic and lipid within the organism. If the term in the numerator of this equation is positive, then the chemical will transfer from the plastic to the biota lipid. When the term is negative, cleanup of the compound from the lipid will occur, and the compound will sorb to the microplastic (Koelmans, 2015).

To summarize, a chemical compound has the potential to sorb from water to the microplastic and from the lipids of an organism to a microplastic. This is known as cleaning up. Additionally, a chemical compound may sorb from the microplastic to the water or from a microplastic to the gastrointestinal tract of an organism. The tendency of a chemical to act in such a manner can be defined according to the previously mentioned equations.

Microplastic Induced Toxicity
Microplastics, as a whole, are not considered intrinsically toxic to aquatic life. However, it is possible for chemicals added to the plastic upon synthesis to later leach out of the plastic upon degradation. Leaching is the process by which a chemical leaves the microplastic and enters the aquatic environment. Additionally, microplastics have the ability to adsorb various chemicals already in an aquatic environment. Once adsorbed, it is possible for these chemicals to subsequently desorb from the microplastics. If a microplastic with an adsorbed chemical is ingested, it is possible for the chemical to desorb inside the aquatic organism and subsequently, harm the aquatic organism. It is additionally possible that the effects of desorption by contaminated particles may bio-magnify up the food chain.

Types of Toxicants
There are a variety of toxicants associated with the aquatic toxicity of microplastics. Toxicants are human-made chemicals that cause a deleterious or harmful response in an organism. The size and type of microplastic ingested by an aquatic organism may result in an adverse physical response. Thus, the properties of a microplastic can elicit a physical adverse response. Additionally, toxicants found in the aquatic environment can potentially adsorb to the microplastic. For example, hydrophobic organic compounds can adsorb to microplastics. Hydrophobic organic compounds (HOCs) are organic compounds that are more soluble in organic solutions than water. Some hydrophobic organic compounds are polybrominated diethyl ethers, polycyclic aromatic hydrocarbons, and polychlorobiphenyls. These HOCs have been found on microplastics synthesized from polystyrene, polyethylene, polyvinylchloride, and polyoxymethylene (Koelmans, 2015). Additionally, persistent organic pollutants are potential toxicants and a major concern. Persistent organic pollutants are lipophilic chemicals that prefer to concentrate on hydrophobic regions of microplastics. Some POPs include polychlorinated biphenyls, organochlorine pesticides such as DDT and DDE, and polyaromatic hydrocarbons (Cole, et al., 2011). There some chemical overlaps classified as HOCs and POPs. Finally, additives are considered possible toxicants. Additives are compounds added to plastic while it is being synthesized. Some examples of additives include flame retardants, plasticizers such as di-ethylhexyl phthalate and bisphenol A, antioxidants, and photostabilizers (Kwon, et al., 2017).

Target Sites for Toxicity
It is important to mention target toxicity sites before discussing possible toxicological effects of compounds or the physical ramifications of ingesting microplastics. Organisms are known to ingest toxicants, as mentioned in previous chapters. Thus, a major toxicity site is the gastrointestinal tract. Aquatic organisms such as fish swallow microplastics by mistaking the particles for natural prey, or swallow microplastics unintentionally by feeding on prey that either have microplastic particles inside or adhered to their surface. As previously mentioned in section C of this chapter, many organisms studied have contained microplastic particles in their gastrointestinal tract. In fish there is no correlation between the number of microparticles residing in the gut and its mass,  length, and/or the species trophic level within the food web (Güven et al., 2017); therefore, microplastics do not exhibit biomagnification. It is only the fish’s location or type of habitat that shows a difference in the number of ingested microplastic particles. For example, pelagic fish ingest more microplastic particles than benthic fish. Overall, the average number of particles found in fish is typically less than 2-3 microplastics (Güven et al., 2017). It is possible for these particles to enter into the circulatory system following ingestion, but this process is extremely rare except for microplastics that measure less than 150 μm. Small amounts of ingested microplastics may also be translocated to the liver, but typically are extremely small: less than 5 μm (Jovanović, 2017).

Adverse Physical Reactions
As previously mentioned, the ingestion of microplastics can lead physically adverse responses. Ingested microplastic particles can physically block the digestive organs and interfere with the organism’s feeding process. For example, ichthyoplankton in later developmental stages can ingest microplastics, and once swallowed the particles may lead to starvation or satiation, the latter resulting in reduced growth rate, reduced ability to avoid predators, and reduced general fitness. Additionally, fish that feed on microplastic containing zooplankton require over twice as much time to consume food when compared to those that feed on microplastic-free zooplankton. Other studies on gobies have indicated that microplastic ingestion leads to decreased predatory performance.

Microplastic particles are often irregular in shape and can thus harm the intestinal lining of organisms such as fish. For example, microplastic ingestion has shown a widening of a lamina propria, vacuolation of enterocytes, increase of goblet cells, hyperplasia of goblet cells, loss of the regular serosa structure, detachment of the mucosal epithelium from the lamina propria, and a shortening and swelling of the villi. It is also possible for microplastics to cause neurological effects and metabolic changes such as the upregulation of fatty acids (Jovanović, 2017). Additionally, the presence of microplastic particles on beaches can raise  beach sand temperature, affecting the ratio of male to female organisms for species that have sex determined by temperature (Ivar do Sul, et al., 2014). As a result, the population of these species may decrease across subsequent generations. Indeed, the physical presence of microplastic can have adverse effect on organisms.

Hydrophobic Organic Compounds
Microplastics, themselves, are not intrinsically toxic, as previously mentioned. Most of the concern surrounding microplastic presence in the ocean reflects the fact that toxicants may adsorb to plastic and then desorb inside an organism, or leach from microplastic into the environment and/or an organism and cause a deleterious response. HOCs as previously mentioned comprise a set of compounds that may adsorb to plastics (Figure 10). It has been shown that HOCs on microplastics may indeed be a source of HOCs in an aquatic environment; however, whether HOCs on microplastics cause a more deleterious response than other HOCs found in the environment is more controversial. Some studies argue that the affinity of HOCs for plastics is very high. On average, the half-life for a HOC to desorb to a microplastic particle is around 2 years. However, the smaller the microplastic, the shorter the half-life of the HOC. Nevertheless, a different picture begins to emerge when one factors in the ratio of microplastics to organic matter or other carrier media for HOCs Some of these other carrier media types have equilibrium partition coefficients that are equal to that of microplastic particles. Thus, the potential toxicity from HOCs absorbed to microplastics can only occur if the concentration of HOC on the microplastic is greater than the concentration of HOCs present in the water or in an organism. Nevertheless, the amount of microplastic particles in an aquatic environment such as the ocean is too small to cause a meaningful redistribution of HOCs from the ocean to microplastic particles themselves. For example, when comparing the transfer of HOCs to microplastics or organic black carbon, it was determined that the partition coefficient for black carbon is twice as high as the partition coefficient of microplastics. Thus, it is twice as likely that a HOC would sorb to organic black carbon than to sorb to a piece of microplastic. Usually, HOCs are present in the water. Therefore, the most likely route of exposure for an organism is by ingesting contaminated water rather than ingesting contaminated microplastics (Koelmans, et al., 2016).

Persistent Organic Pollutants
Persistent organic pollutants also have the potential to adsorb to pieces of microplastic, causing concern that once ingested these POPs may desorb from the microplastic and cause harm to the organism (Figure 11). POPs have very large partitioning coefficients in favor of microplastics; therefore, microplastics have a tendency to clean up POPs from aqueous solutions, but the amount of macroplastics present in the environment is quite small compared to that of POPs. Hence, not all POPs can be removed via the presence of microplastics.

A study by Tueten, et al. indicated that POPs have a very slow desorption process off of microplastics. Although it is possible for POPs to already be present in a microplastic particle and leach out of the particle, the number of particles for which this could occur is negligible compared to the amount of POPs that enter the aquatic environment from other sources. Nevertheless, microplastics tend to have a cleaning up effect on POP. If a microplastic particle is ingested inside an organism whose gut contains substantial levels of POPs, the microplastic will serve as a sink for the POPs and actually decrease the level of POPs inside the organism (Andrady, 2011; Koelmans, 2015). Whether toxicants such as HOCs and POPs that can adsorb to microplastic particles and then later desorb are toxic is in direct correlation to the amount of that toxicant present in the environment. A bioaccumulation of the compound inside an organism might occur if the microplastic contains the only source of the chemical. Nevertheless, it is extremely unlikely that microplastics would contain the only source of these toxicants. Therefore, the possibility of bioaccumulation via a microplastic vector decreases as the chemical gradient shifts towards the adsorption of these toxicants by microplastics. As a result, microplastics are more likely to serve as cleaners than contributors to the bioaccumulation of HOCs and POPs (Koelmans, 2015; Koelmans, et al., 2016).

Additives
Finally, a major concern is that additives might act as toxicants to aquatic organisms. Since these chemicals may leach out of plastic, there may be more chemicals present in the plastic during the beginning stages of leaching than in the water or organism. Once the process of leaching has commenced, it could continue until a chemical equilibrium gradient has been established with the microplastic and its surrounding solution. Some laboratory studies have indicated that leaching of an additive from microplastic can occur inside an organism. Other model studies indicate that the number of additives that leach out of a microplastic in an organism would be negligible.

To further complicate matters, compounds considered to be additives have a widespread range of characteristics. Some of them are very hydrophobic while others are more hydrophilic. Thus, the probability of an additive causing a toxicological effect varies from additive to additive. For example, additives with long alkyl groups are not likely to bioaccumulate, but those such as benzotriazole UV stabilizers may do so. The ability for additives to cause a deleterious effect on aquatic organisms has been documented, but it is unclear whether the effects were due to additives that came directly from microplastics or from other sources (Kwon, et al., 2017).

Problems Regarding Microplastic Research and Future Directions
The field of microplastic research that has investigated microplastics’ potential toxicity to aquatic environments is relatively new. Thus, there are numerous problems to consider when designing an experiment or reading papers on the topic. First and foremost, there is not a standard definition for microplastics. The definition changes according to the researcher who attempts to redefine it, making it difficult to fully grasp the topic. Additionally, there are no standardized techniques on how to measure, study and analyze microplastics, or their effects on the aquatic environment. Plankton nets are commonly used, but they fail to collect some of the smaller microplastics. Furthermore, current data regarding the toxicity of microplastics or possible toxicants adsorbed to them is controversial. Some experiments demonstrate that the presence of microplastics and their toxicants such as HOCs, POPs, and additives do cause a deleterious response. Other experiments and models indicate there is little to no effect caused by microplastic particles when compared to the presence of the same chemicals in water. Furthermore, the problem of microplastic research is exacerbated by the fact that it is not clear what the actual environmental concentration (AEC) of microplastics is, and that the AEC is highly variable both on a macroscale (geographically) and on a microscale (water surface, water column, and sediment).

One of the difficult aspects of any aquatic toxicity tests is selecting the type of test to perform. A variety of tests can be performed in microplastic research such as laboratory tests, model systems, and field tests. However, each of these tests has its positive and negative attributes. Often, the doses used in laboratory tests do not reflect actual or estimated environmental concentrations and far exceed worst-case scenarios. These tests also focus on only one species and ignore intricate aspects of the ecosystem. Furthermore, modeling studies often fail to be evaluated with actual data; thus, they may not reflect what is actually occurring in the environment. Finally, field-based studies are difficult to perform. They require considerable time, money, and preparation. When preformed, it is often difficult to extrapolate data regarding the compound due to the presence of many confounding factors. These tests are also unable to be replicated in the exact manner.

In the future it will be prudent for the field of microplastic research in the aquatic environment to settle on a proper definition of microplastic and establish standardized techniques. Additionally, more studies should investigate the fate of microplastics in larger marine animals and in freshwater organisms. Additional data is needed to illustrate how HOCs, POPS, and additives interact with microplastics both outside and inside aquatic organisms for toxicological purposes. This will require the development of a highly thought out screening process  that will allow multiple types of microplastics and possible toxicants to be evaluated simultaneously. It is impossible to test all combinations of microplastics with all HOCs, POPs, and additives in a lab. Thus, steps need to be taken to streamline the process.

Conclusion
The field of microplastic research in aquatic toxicology is a relatively new field. Microplastics are located worldwide in all types of aquatic environments. It has been well documented that microplastics can be ingested by both aquatic invertebrates and aquatic vertebrates. Additionally, microplastics degrade at a very slow rate through photodegradation and thermooxidative degradation. Although microplastics are not considered intrinsically toxic themselves, it is possible for chemicals to adsorb to particles and then leach off the microplastics once inside an organism. This is highly unlikely for HOCs and POPs, but more data is needed for additives. Finally, as with all new fields, this field has some shortcomings such as a lack of standardized definitions and techniques. It is expected that more solid definitions and protocols will emerge in the future.

Glossary
1.    Additives: Compounds added to plastic while it is being synthesized.

2.    Beach combing: Collecting, identifying, and quantifying all litter items on the shore of a specific coastline region.

3.    Biodegradation: Process by which living organisms degrade microplastics or other chemicals.

4.    Degradation: Chemical process by which the average molecular weight of a polymer is reduced.

5.    Hydrolysis: Process by which water is used to degrade a compound.

6.    Hydrophobic organic compounds (HOCs):  Organic compounds that are more soluble in organic solutions than in water.

7.    Leaching: Within the context of this chapter, a process by which a chemical leaves the microplastic and enters the aquatic environment.

8.    Macroplastics : Plastics that are easy to see with the naked eye and are > 5 mm in diameter.

9.    Marine observational sampling: Involves divers or onboard observers who record the size, location, and types of plastics in the water.

10.  Marine trawls: Use of fine meshes or nets to collect microplastic particles or marine organisms.

11.  Microplastic: Plastic particle that is < 5 mm in size but greater than 100 nm.

12.  Nanoplastic: Plastics that are between 1-100 nm.

13.  Persistent organic pollutants (POPs): Lipophilic chemicals that prefer to concentrate on hydrophobic regions of microplastics.

14.  Photodegradation: Process by which light, usually sunlight, degrades a material.

15.  Physical Adverse Response: Within the context of this chapter, a negative physiological response to a microplastic particle.

16.  Plastics: Long chains of polymers that are synthesized from both organic and inorganic materials such as carbon, hydrogen, silicon, oxygen, and chloride; usually acquired from natural resources such as natural gas, oil and coal.

17.  Primary microplastics: Plastics manufactured in the microplastic size range.

18.  Secondary microplastics: Microplastic particles that have fragmented from macroplastics.

19.  Thermal degradation: Degradation that occurs via high temperatures.

20.  Thermooxidative degradation: Process by which particles slowly break down through oxidation in moderate temperatures.

21.  Toxicant: Human-made chemicals that cause a deleterious or harmful response to an organism.