A new approach to extracting biofilm from environmental plastics using ultrasound-assisted syringe treatment for isotopic analyses&#160;
Authors: Battulga Batdulam , Atarashi-Andoh Mariko , Koarashi Jun 
Published: March 2022

&lt;p&gt;Plastic debris has been recognized as one of the carriers of hazardous substances in the aquatic ecosystem due to its ubiquitous distribution and potential interaction with pollutants through developed biofilms. Although an increasing number of studies have highlighted that diverse microbial species including biofilm-forming microorganisms are colonized on the plastic surfaces in aquatic environments, biofilm-mediated interactions between plastics and pollutants especially radionuclides remain unclear. In this study, we aimed to extract biofilms from the environmental plastics using a newly developed extraction method and to determine the concentration of radiocesium (&lt;sup&gt;137&lt;/sup&gt;Cs) and stable isotope ratios (&amp;#948;&lt;sup&gt;13&lt;/sup&gt;C and &amp;#948;&lt;sup&gt;15&lt;/sup&gt;N) in the extracted biofilm samples. Visible plastics were collected from the mouths of coastal rivers in Ibaraki prefecture, Japan. Although the plastic concentration along the river mouth was not evaluated in this study, various abundances (96 &amp;#8211; 1868 pieces), sizes (1 &amp;#8211; 50 mm), colors, and morphotypes of plastics were applied to the extraction procedures. After plastic and biofilm separation with ultrasonication, biofilm samples were collected by the two ways: freeze-drying (15.5 &amp;#8211; 44.4 mg); and freeze-drying after syringe treatment (14.5 &amp;#8211; 65.4 mg). The XRD diffractograms of biofilm samples confirmed that biofilms obtained by freeze-drying only were still heterogeneous and the agglomerations of organic substances, mineral particles, and small microplastics (MPs, &lt;1 mm). The results also demonstrated that biofilm extraction was achieved by syringe treatment separating the mineral and small MP particles, resulting in homogenous biofilms from the surface of plastics. Preliminarily results of &lt;sup&gt;137&lt;/sup&gt;Cs activity concentrations in heterogenous (ranging from 0.22 to 0.54 Bq g&lt;sup&gt;&amp;#8722;1&lt;/sup&gt;) and homogenous (0.82 &amp;#177; 0.04 Bq g&lt;sup&gt;&amp;#8722;1&lt;/sup&gt;) biofilm samples revealed that plastics serve as a carrier for &lt;sup&gt;137&lt;/sup&gt;Cs in the coastal river environment mediated by developed biofilms. As a result of the presence of petroleum-derived small MPs, heterogeneous biofilm samples showed a relatively lower &amp;#948;&lt;sup&gt;13&lt;/sup&gt;C value (&amp;#8722;26.03 &amp;#177; 0.34&amp;#8240;; mean &amp;#177; SE) compared with homogenous biofilm samples. A similar trend was observed in the &amp;#948;&lt;sup&gt;15&lt;/sup&gt;N values. Our results suggest that developed biofilms on the plastics might have specific signatures of &amp;#948;&lt;sup&gt;13&lt;/sup&gt;C and &amp;#948;&lt;sup&gt;15&lt;/sup&gt;N depending on the source and pathway of the organic matter. The study contributes to the knowledge of the developed biofilms on environmental plastics and their potential interactions with &lt;sup&gt;137&lt;/sup&gt;Cs in the coastal aquatic environment.&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Keywords: &lt;/strong&gt;environmental plastics, biofilm, coastal river, radiocesium, stable carbon and nitrogen isotopes&lt;/p&gt;