**1.1 Plastic debris in the ocean: a global environmental issue of the twenty-first century**

Since 1950, the production and use of plastics has been constantly increased reaching a global production of 280 million tons in 2016 (i.e., as thermoplastics and polyurethanes), with China as the major producer (29%) [1].

Plastics represent a group counting hundreds of different materials derived from fossil sources (e.g., oil and gas) among which the most produced are polypropylene (PP), high- and low-density polyethylene (HDPE, LDPE), polyvinyl chloride (PVC), polyurethane (PUR), polyethylene terephthalate (PET), and polystyrene (PS). Due to their high versatility, durability, low weight, and low cost, plastic materials find applications in almost any market sector, but primarily in packaging (39.9%) and building industries (19.7%) [1].

In recent years, the growing evidence about the massive presence of plastic litter in the ocean, its pressure on the marine environment and wildlife, and its impact on marine-related human activities (such as fishery, shipping, and tourism) has raised lot of attention in the scientific, regulatory, and civil communities (**Figure 1**).

Oceanographic surveys have recorded the presence of plastics in any geographical regions, including remote polar areas, and at any depth, from the sea surface to the seafloor of the oceans (**Figures 2–4**).

The amount of plastic debris in the sea is still unknown due to the large variability of its distribution as regards both spatial and temporal scale, which prevents accurate estimates. However, modeling studies have recently approximated that 5–13 million tons of plastics (i.e., equivalent to 1.5–4% of global plastic production) end up in the oceans every year [2].

The slow degradation rates of plastics under environmental conditions provide additional complexity to this global issue, by contributing to their accumulation in all terrestrial and aquatic environments. It has been estimated that, once in the ocean, the majority of manufactured polymers persist for decades and probably for centuries due to their low degradability (**Figure 5**) [3, 4].

In both terrestrial and marine environments, degradation of petroleum-derived plastics occurs through abiotic and biotic processes (i.e., UV degradation, hydrolysis, and decomposition by microorganisms), leading to their fragmentation into increasingly smaller pieces. Thus, plastic particles dispersed in the environment are commonly divided into three main classes based on their size: macro: >25 mm,

*Elemental Analyzer/Isotope Ratio Mass Spectrometry (EA/IRMS) as a Tool to Characterize... DOI: http://dx.doi.org/10.5772/intechopen.81485* 

meso: 5–25 mm, and microplastics: <5 mm. The smaller-size class, which includes both primary microplastics (i.e., particles produced as such, e.g., plastic pellets, exfoliating cosmetics, or synthetic clothing fibers) and secondary particles (i.e., particles derived from the breakdown of larger plastic debris), is likely to be the most abundant in the ocean today [5].

**Figure 2.**  *Marine litter on the beach (photo by Tomaso Fortibuoni).* 

**Figure 3.**  *Seabird nesting on plastic nets (public domain).* 

**Figure 4.**  *Tangle of fishing nets on a beach (photo by Francesca Ronchi).* 

#### **Figure 5.**

*Estimated decomposition times of different types of garbage dispersed in the marine environment (illustration by Davide Zanella).* 

The concern about the heavy contamination of the marine environment by plastics is related to the potential of plastic debris to cause harm to the inhabiting organisms via different mechanisms. Among the most alarming issues, there is an uptake and a bioaccumulation of plastic debris by marine organisms at almost all levels of the food web and the consequent trophic transfer. Recent studies have reported that micro and nanoplastics can easily be taken up and ingested by marine organisms (i.e., zooplankton, worms, bivalves, crustaceans, demersal and pelagic fishes, seabirds, reptiles, and mammals), resulting in a significant impact on the aquatic wildlife and possibly on human health via seafood consumption [6]. Furthermore, due to the large surface to volume ratio, microplastic fragments can potentially adsorb many kinds of common marine contaminants on their surface, in particular hydrophobic organic substances such as polychlorinated biphenyls, polyaromatic hydrocarbons, and organochlorine pesticides [7, 8]. This can promote their transport in the environment and induce toxic effects following ingestion and desorption (e.g., endocrine disruption, mutation, and cancer). Moreover, another source of concern is the possible release of additives commonly present in plastic formulations (i.e., bisphenol A, phthalates, and flame retardants) [8, 9], and although the leaching rates of these common additives in seawater are poorly known, their potential for toxicity is considered to be very high.

Several actions have currently been undertaking at national and international levels to tackle the contamination of marine environments by plastics. Their main aim is to achieve a general reduction of plastic use (in particular packaging and disposable items), recycling of plastic items at the end of their lifetime, and replacement of the use of plastics with more sustainable materials and biopolymers (e.g., plant-derived polymers [10]), which are more prone to degradation by microorganisms and show a shorter persistence once dispersed in the environment.

#### **1.2 Experimental approaches to assess plastic debris in environmental samples**

 With the growing evidence of the severe impact caused by plastics on the wildlife, the assessment of the presence, behavior, and fate of plastics in the marine environments has become a fundamental research issue, highly advocated to the scope of putting in place more effective policies. However, especially for the smallest particles (i.e.,

#### *Elemental Analyzer/Isotope Ratio Mass Spectrometry (EA/IRMS) as a Tool to Characterize... DOI: http://dx.doi.org/10.5772/intechopen.81485*

 microplastics), their efficient identification to the scope of assessing the plastic load in the environmental compartments (e.g., seawater, sediments, and biota) is a serious challenge for scientists. Many analytical techniques have been used to identify plastic debris in environmental samples, as largely reviewed in the literature [11, 12]. Among the most used approaches, there are electron scanning microscopy coupled with energy dispersive X-ray spectroscopy (SEM-EDS, ESEM-EDS), Raman spectroscopy, Fourier transform infrared spectroscopy (FT-IR) [13], and thermal analysis (pyro-GC/MS). Other analytical methods used to identify plastic materials are near infrared spectroscopy (NIRS), differential scanning calorimetry [14], and UV-VIS spectroscopy [15, 16].

Stable isotope analysis, which is an analytical technique that measures the relative abundance of stable isotopes yielding an isotope ratio that can be used as a research tool, is finding application in a growing number of different research fields and practical case studies. For instance, it is widely used to trace the origin of organic matter in various environments [17, 18], to track fraud in the food industries [19] and to identify microtraces of drugs, flammable liquids, and explosives in forensic cases [20]. This technique has been only rarely applied to assess the presence of microplastics in environmental samples [21]. Its potential for detecting plastic debris in environmental samples relies on different isotopic signatures of carbon in (i) petroleum-derived materials, (ii) C4 plants used in the synthesis of bioplastics, and (iii) marine samples' matrices (e.g., particulate organic matter, plankton, tissues of marine organisms, algae, and marine plants).

#### **1.3 Stable isotope analysis: principles of the method**

The term isotopes (from the Greek iso, same and topos, place) identifies atoms of the same chemical element, that is, the same place in the periodic table of the elements, that has the same atomic number but different atomic mass number. In other words, isotopes are atoms having the same number of protons and electrons (equal chemical properties) and a different number of neutrons (different physical properties). Each element has known isotopic forms, and in total, there are 275 isotopes of the 81 stable elements, in addition to over 800 radioactive isotopes (**Figure 6**).

Isotopes of a single element possess almost identical properties. They are commonly classified as natural or artificial, stable, or unstable. The quantification of the ratio between two isotopes allows to determine if two chemically similar environmental samples have different origins, related to the difference of the original sources. The isotopic distribution characterizing the sources may be influenced by phenomena of a different nature, which in turn may cause significant variations in the final products.

**Figure 6.**  *Stable isotopes have a proton/neutron ratio lower than 1.5.* 

 Depending on the chemical element, variations in the relative mass abundance of its isotopes can be detected through the analysis of stable isotopes. Technological advances in isotope analysis have led to the development of scientific instruments able to measure very small variations in the abundance of stable isotopes with high precision and accuracy (mass spectrometry). Therefore, stable isotope analysis can be applied considering different elements, thus giving nowadays applications in different fields of science.

For a given chemically stable element, its isotopic composition in a sample (R) is equal to the ratio between the abundance of the heavy isotope with respect to the light one (e.g.,13C/12C), and it is expressed as deviation, in parts per thousand, from an international reference standard material (δ‰), according to the equation (Eq. (1)) given below:

$$\text{\{\{\%\}} \, = \, \left[ \{ \text{R}\_{\text{sample}} - \text{R}\_{\text{standard}} \} / \text{R}\_{\text{standard}} \right] \times \text{1000} \tag{1}$$

 where Rsample is the mass ratio of the heavy isotope to the light isotope measured in a sample and Rstandard is the isotopic ratio defined for the standard. The standard reference material that is commonly used for carbon is Vienna Pee Dee Belemnite. Thus, positive δ values indicate that the heavy isotope is enriched in the sample compared to the standard, while negative δ values indicate that the heavy isotope is depleted in the sample.

 The possibility of distinguishing two samples on the basis of their relative abundance of two isotopes bases on the phenomenon of isotopic fractionation, which can be enacted by a wide range of chemical (e.g., nitrification and ammonification), physical (e.g., evaporation and condensation), and biological (e.g., photosynthesis, assimilation, and excretion) processes. In fact, many natural (and anthropic) processes can alter the isotopic signature of a chemical element in a matrix by causing an imbalance of the isotope distribution that leads to a variation of its original isotopic signature [22]. Thus, as the extent of fractionation of many chemical elements have been proved to be sensitive to specific processes/variables, it can be used as a tool to investigate the involved process/variable itself. In general, two mechanisms of isotopic fractionation can be distinguished:

	- heavy isotopes accumulate in oxidized products;
	- • the isotopic fractionation is favored at low temperatures, since at high temperatures, the differences between the isotopes are attenuated;
	- • the process is not relevant in the case of chemical reactions of gaseous substances and biological reactions.
	- the preferential breaking of the bonds formed by light isotopes;
	- • the preferential distribution of light isotopes in products and of the heavy ones in the reagents.

*Elemental Analyzer/Isotope Ratio Mass Spectrometry (EA/IRMS) as a Tool to Characterize... DOI: http://dx.doi.org/10.5772/intechopen.81485* 

Given a chemical substance AB characterized by the presence of a certain isotopic distribution of element X, we can calculate the fractionation factor by dividing the ratio of the number of isotopes X in product A with the ratio of the number of isotopes X in product B (Eqs. (2) and (3)).

$$\begin{array}{rcl} \mathfrak{\alpha}\_{AB} &=& \frac{R\_A}{R\_B} = & \mathbf{1} + \left[ \frac{(\boldsymbol{\delta}\_A - \boldsymbol{\delta}\_B)}{1000} \right] \end{array} \tag{2}$$

where

$$R\_{\perp} = \frac{X\_h[\text{atoms of the heavier isotope (pure)}]}{X\_l[\text{atoms of the lighter isotopes}] \,\text{atm} \,\text{atm} \,\text{(abaudant)}} \tag{3}$$

 However, the fractionation factor (α) is normally replaced by the isotopic enrichment factor (ε), which is defined as (α − 1) × 1000.

#### **1.4 Carbon isotope ratio as a tool in environmental studies**

Carbon and nitrogen isotope analysis is used to investigate the trophic web and the matter flows among the main components of an ecosystem (e.g., organic matter, producers, primary and secondary consumers); it can be used to understand chemical and biological processes occurring at both ecosystem and organism levels. Stable isotope analysis can also be a useful tool for assessing the origin of water, atmospheric, and soil pollution.

The two main carbon reserves in nature are represented by organic and inorganic carbon, which are characterized by different isotopic fingerprints due to the different processes in which they are involved (**Figure 7**). The inorganic carbon (carbonate) is involved in the exchange equilibrium among (i) atmospheric carbon dioxide, (ii) dissolved bicarbonate, and (iii) solid carbonate. The exchange reactions among these three forms lead to an enrichment of the heavy isotope in the

**Figure 7.**  *Isotopic fingerprint of naturally occurring carbon.* 

 solid carbonate form (δ13C equal to 0‰). In contrast, the kinetic reactions which mainly involve the organic carbon (i.e., photosynthetic process) determine a concentration of the lightest isotope in the synthesized organic material (δ13C equal to about −25‰) [17].

 The fractionation of organic carbon is mainly linked to the specific photosynthetic pathway featuring each plant. The terrestrial plants, classified as C3 and C4, can follow two different photosynthetic pathways. Both types synthesize organic matter characterized by δ13C values more negative than that of carbon dioxide (~−7‰), because during the photosynthesis, the produced organic substance accumulates the light isotope compared to the heavy one. The C3 plants, typical of temperate climates, produce the 3-phosphoglyceric acid, a compound with three carbon atoms (Calvin cycle) with an average value of δ13C of about −26.5‰. The C4 plants generate oxaloacetate, a compound with four carbon atoms (Hatch-Slack cycle) characterized by a value of δ13C around −12.5‰.

The chemical composition of animal tissues is related to the food sources they assimilate, and therefore, it reflects the isotopic composition of the diet [23, 24]. The enrichment between primary producers and consumers (herbivores) has been estimated to be approximately +5‰, whereas at the successive trophic levels, the enrichment is less marked (+1‰) [25]. Thus, the isotopic value detected in the tissues of an organism can be potentially used as an indicator of its trophic position. However, since the variation of the δ13C values due to trophic passages is relatively modest, δ13C is mainly used to trace the primary carbon source used [26].

 Through the analysis of the stable carbon isotopes, it is also possible to differentiate terrestrial and marine trophic webs. The "marine" carbon derives from the dissolved inorganic carbon (dissolved bicarbonate) characterized by an isotopic value equal to about 0‰, while the "terrestrial" carbon derives from the atmospheric carbon dioxide which has a lower δ13C value (approximately −7‰). This difference is maintained at every trophic level both in the marine and terrestrial trophic chain (**Figure 8**).

#### **Figure 8.**

*Variations of δ13 carbon and δ15 nitrogen (‰) isotopes in different organisms of the terrestrial and marine food chain.* 

*Elemental Analyzer/Isotope Ratio Mass Spectrometry (EA/IRMS) as a Tool to Characterize... DOI: http://dx.doi.org/10.5772/intechopen.81485* 
