Lake Kallavesi is the biggest lake situated in eastern Finland, encompassing the city of Kuopio.
It is the tenth largest lake in Finland with an area of 478.1 km2, containing several sub-basins and diversified islands (Uurasjärvi et al., 2020; Johansson et al., 2019; Partanen and Hellsten, 2005). The average and maximum depth of Lake Kallavesi are 9.7 m and 75 m, respectively (Uurasjärvi et al., 2020). The lake is located in the boreal vegetation zone. The annual average temperature of this region is +3 °C while the average temperature of the coldest month, January is –10 °C and the warmest month, July is +17 °C. Around 644 mm/year of precipitation is recorded, out of which half is in the form of snow. For around 6 months from November to April, the lake remains covered with ice (Johansson et al., 2019). Lake Kallavesi is the source of drinking water for Kuopio city (Uurasjärvi et al., 2020). Samples were taken from Kallavesi lake under the Kallansillat Bridge (Fig. 1).
Figure 1. Study site (National Land Survey of Finland) 3.2 Sampling and sample processing
The sediment trap method was used for trapping samples from lake Kallavesi. On 11 November 2016, a Sediment trap was placed to the bottom of the Lake from the boat. Boat traffic and ice cover interferences were avoided, and sample representativeness was ensured by choosing maximum depth for the placement of a sediment trap. Winter 2016-2017 samples were taken on 3rd June 2017 in two collector tubes and then these collector tubes were fixed again to the sediment trap which was installed back to the bottom of the lake for summer samples. Summer 2017 samples were collected on 21st October 2017. A sample representing winter 2017-2018 was taken on 26th May 2018 and a sediment trap was slowly placed to the bottom of the lake for summer samples. Summer 2018 samples were collected on 6 Oct 2018 (Saarni et al., 2021).
Sediment properties were determined by using a sample from one collector tube while microplastic analysis was done by using the sample from another collector tube. Hydrogen peroxide (H2O2) was used for the decomposition of organic matter (from microplastic surfaces) and then the heavy-liquid density separation method was used for separating microplastics from clastic sediments. Lithium heteropolytungstate was diluted to density of 2.0 g/cm3 and used as
a heavy liquid solution for the density separation method (Saarni et al., 2021). The isolated microplastics and remaining particles were filtered on standard 12-15 µm pore size general purpose filters (Munktell Ahlstrom, size 90 mm, grade 1003) for further inspection.
3.3 Visual inspection of microplastics
The visual method was used for the selection of microplastics. Microplastics were visually isolated from filters with a pair of micro tweezers under Zeiss Stemi 508 stereomicroscope with 6.3 - 50 × magnification. Particles were photographed with a stereo microscope camera (Axiocam ERc 5s camera). Particles were picked according to the criteria set by Noren (2007):
fibre particles having an equal thickness, uniformly coloured, non-natural, and transparent particles. Picked particles were transferred to 15 ml centrifuge tubes with the help of micro tweezers and then vacuum filtered through a silver membrane filter having a pore size of 5µm and diameter of 25mm. Filtration was carried out under a vacuum hood and filtration funnel and centrifuge tubes were rinsed with deionized water. Filters were dried at room temperature in lidded glass Petri dishes, Next, dried filters were attached to glass microscope slides with the help of double-sided tape for microplastic identification and analysis.
3.4 Identification and analysis of microplastics
Microplastic identification was conducted using an imaging Fourier-transform infrared spectroscopy (FTIR). Calibration and background check of imaging FTIR was done before by putting a glass microscope slide under the imaging FTIR for analysis. Imaging FTIR utilized for the analysis of particles consists of Agilent Cary 670 spectrometer and Cary 620 microscope incorporated with focal plane array (128×128 FPA) detector. Measurements were taken in a reflection mode with a pixel size of 5.5 µm, a spectral resolution of 8 cm-1, and a spectral range of 3800-750 cm-1. Total scans taken with FTIR were 4.
siMPLe software which is produced by Aalborg University, Denmark, and Alfred Wegner institute, Germany, (Primpke et al., 2020) was used for the analysis of spectral data obtained from FTIR. This software provided information about particle size, mass, and plastic types by calculating Pearson’s correlation between samples and reference spectra. Reference spectral library consists of both open source and in-house spectra of prevalent plastic types as well as natural proteins and cellulose polymers. Common plastic types used for the reference database were polyamide (PA), polyethylene (PE), polyethylene terephthalate (PET), polystyrene (PS), polypropylene (PP), polyvinyl chloride (PVC), polyurethane (PU), acrylonitrile butadiene styrene (ABS), polyacrylonitrile (PAN), and polymethyl methacrylate (PMMA).
Raw spectra, first derivative, and second derivative were used for the calculation of correlations.
siMPLe software determined the major dimensions of the particles by calculating the longest distance between the pixels of identified plastic particles. Correlation thresholds were also set for the determination of plastics.
3.5 Data Reporting
Morphological characteristics of particles were evaluated from the data provided by siMPLe software. siMPLe calculates the major and minor dimensions (µm) of the particles and this information was used for finding the shapes of particles. Shapes were estimated by dividing the major and minor dimensions of the particles (Eq. 1). If the dimension ratio of the particle was greater than 5 then it was considered as fibre.
Particle dimension ratio = 𝑀𝑎𝑗𝑜𝑟 𝑑𝑖𝑚𝑒𝑛𝑠𝑖𝑜𝑛 (µm)
𝑀𝑖𝑛𝑜𝑟 𝑑𝑖𝑚𝑒𝑛𝑠𝑖𝑜𝑛 (µm) (1)
The concentration of microplastics in sediments was calculated by using the number of plastic particles provided by siMPLe and the dry weight of samples (Eq. 2). The concentrations of particles were presented in items/g.
MP concentration = 𝑃𝑎𝑟𝑡𝑖𝑐𝑙𝑒 𝑛𝑢𝑚𝑏𝑒𝑟
𝐷𝑟𝑦 𝑤𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 (𝑔) (2) 3.6 Contamination control
Laboratory contamination was assessed by the control sample. Open petri dish containing clean filter was exposed to laboratory air for thirty minutes during laboratory work. Afterward, particles were picked under stereomicroscope and microplastic contamination was analyzed by FTIR in the same way as real samples. Moreover, samples were kept in closed centrifuge tubes to avoid air contamination and filtration was carried out under fume hood.