## 4 Methods for Detecting of Microplastic Particles
This method involves either recording the fluorescence spectra of the bulk solution or using fluorescence microscopy. The first approach does not directly provide the exact number of particles but rather a general signal proportional to their concentration. The second allows for the assessment of particle quantity, shape, and size. However, the question remains open regarding the sensitivity of this method, the lowest concentration ranges it can detect, as well as its selectivity. The latter issue is likely still relevant despite the fact that the analyte is dispersed in a relatively simple matrix, such as bottled water. This method appears to be suitable for strictly defined, synthetically produced samples with well-known composition, rather than for the analysis of environmental or real-life food samples. A variation in the method described above involves using the fluorescent dye Nile Red. The use of this reagent provides a significant degree of specificity [90]. This method is well recognized for such measurements and is sometimes considered a standard approach [91; 92; 93]. The procedure involves binding the dye to microplastic particles, followed by microscopic observation. However, the detection of less hydrophobic particles, such as PET, may be somewhat limited [94]. The combination of microscopic observations with spectral techniques offers much greater specificity and the ability to identify the material of MPs compared to simple fluorometric techniques mentioned earlier [89; 90; 94]. While \(\upmu\)-FTIR and \(\upmu\)-Raman are more complex and expensive, they do have limitations and do not address all the challenges. Standard FTIR microscopy, for example, struggles with detecting particles smaller than 20 \(\upmu\)m, and the resulting spectra may be inaccurate [95]. This led to the use of more advanced techniques, such as optical photothermal infrared spectroscopy (O-PTIR) and tunable mid-infrared quantum cascade laser (QCL-IR) technology [96]. O-PTIR is a non-invasive technique that uses the photothermal effect. The sample is illuminated with IR radiation, causing thermal expansion of the material and a change in the refractive index, which can then be measured using a narrow visible light beam [97]. Its advantage is its high spatial resolution, around 450 \(\times\) 450 nm [98]. Figure 4: Cumulated microplastic particle size distribution detected in bottled waters and tap water. Each line corresponds to a single water sample. Image from [PERSON] et al. [31], PLOS, CC-BY 4.0. The QCL-IR system includes a set of tunable lasers enabling emission at specific wavelengths, covering the wavelength range from 1800 to 950 cm\({}^{-1}\)[99]. The QCL-IR method is faster and covers a larger area of the sample, but it has a lower resolution compared to O-PTIR. The QCL-IR is not suitable for detecting the smallest MPs, ranging from 1 to 3 \(\upmu\)m [96]. A promising modification of Raman microscopy for MP detection is the use of Surface-Enhanced Raman Spectroscopy (SERS) with a Klarite substrate [100]. The substrate, on which the sample is placed, contains a dense grid of inverted pyramidal cavities made of gold, which focus light into hotspots. This structure enables large electric field enhancement. The prepared surface is then subjected to Raman mapping. The aim of developing this method was to create an uncomplicated but reliable analytical technique for detecting particles smaller than 1 \(\upmu\)m. Research has shown that the application of this method allows for the detection and identification of MPs down to 360 nm. Thermo-analytical techniques, such as pyrolysis gas chromatography-mass spectrometry (py-GC-MS), represent a distinct approach compared to spectral methods [101; 102]. This approach was developed in an attempt to address the limitations of IR and Raman imaging. In this method, polymers undergo thermal decomposition (pyrolysis), followed by separation using gas chromatography. The resulting degradation products are then identified based on mass spectrometry analysis.