Chemosphere 336 (2023) 139186

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Chemosphere
journal homepage: www.elsevier.com/locate/chemosphere

Developing and testing a workflow to identify microplastics using near

infrared hyperspectral imaging
Andrea Faltynkova a, *, Martin Wagner a
a
Department of Biology, Norwegian University of Science and Technology, Høgskoleringen 5, 7491 Trondheim, Norway

H I G H L I G H T S G R A P H I C A L A B S T R A C T

• Open spectral database and chemo­

metric model for routine analysis of
microplastics.
• High performance in identifying micro­
plastics made of four polymer types.
• Hyperspectral imaging accurately char­
acterized size of polystyrene micro­
plastics >1000 μm.
• Photooxidation did not affect precision
of polymer identification.
• Promising technique to increase analysis
throughput for microplastic monitoring.

A R T I C L E I N F O A B S T R A C T

Handling Editor: Michael Bank The analysis of microplastics (MP) is time-consuming which limits our capacity to monitor and mitigate plastic
pollution. Here, near infrared (1000–2500 nm) hyperspectral imaging (NIR-HSI) offers an advantage over other
Keywords: spectroscopic techniques because it can rapidly image large areas relative to other systems. While NIR-HSI can
Imaging spectroscopy successfully detect MP, accuracy and limitations of the method have not been fully explored. In addition, lack of
Method development
open databases and analysis pipelines increases the barrier to use.
Fourier transform infrared spectroscopy
In this work, we developed a spectral database containing preproduction pellets, consumer products and
Raman spectroscopy
Macroplastic marine plastic debris, imaged using a Hyspex SWIR-320me imager. A SIMCA model identified four polymer
Weathering types: polypropylene, polyethylene, polyethylene terephthalate and polystyrene (PP, PE, PET, PS) to identify MP
Chemometrics in hyperspectral images. We determined the accuracy of size estimates for PS MP > 1000 μm using fluorescence
microscopy and tested the impact of photooxidation on detection of plastics by NIR-HSI (PE, PP, PS, PET) and
subsequent prediction by the SIMCA model.
The model performed well across all polymers as shown by high specificity, sensitivity, and accuracy for in­
ternal cross validation (>88%), and sensitivity >80% for external validation. PS MP < 500 μm Feret diameter
were not consistently detected by NIR-HSI when compared with fluorescence microscopy. However, estimates for
Feret diameter were consistent for PS MP > 1000 μm. Analysis by NIR-HSI showed no spectral changes and
SIMCA showed no decreased precision with increased photooxidation across polymer types. Recall varied across
polymer type and photooxidation stage with no clear trends.
This study shows that NIR-HSI is a rapid method which can accurately identify MP of the four most relevant
polymer types, precluding the need to analyze particles one at a time. NIR-HSI can be a key technology for
environmental monitoring of plastic debris where rapid analysis of multiple samples is required.

* Corresponding author.
E-mail address: andrea.faltynkova@ntnu.no (A. Faltynkova).

https://doi.org/10.1016/j.chemosphere.2023.139186
Received 20 January 2023; Received in revised form 7 June 2023; Accepted 9 June 2023
Available online 22 June 2023
0045-6535/© 2023 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
A. Faltynkova and M. Wagner
Abbreviations
ATR attenuated total reflectance
CI carbonyl index
FN false negative
FOV Field of view
FP false positive
FPA focal plane array
FTIR Fourier transform infrared spectroscopy
HI hyperspectral imager
HSI hyperspectral imaging
IR infrared
LOD limit of detection
MP microplastic
1. Introduction
Methodologies for confirming microplastics (MP, 1 nm–5 mm
diameter (CSWRCB)) as synthetic polymers, indicating the polymer type
and describing their size and morphology are critical for characterizing
plastic debris in the environment but also in experimental settings.
Recent research has applied NIR-HSI in parts of the near infrared (NIR)
and short-wave infrared (SWIR) regions as a method to identify MP in
diverse matrices (Faltynkova et al., 2021), including tissues (Piarulli
et al., 2020; Zhang et al., 2019), seawater (Serranti et al., 2018; Shan
et al., 2019; Karlsson et al., 2016), sand (Vidal and Pasquini, 2021) and
soil (Zhao et al., 2018; Paul et al., 2019). NIR-HSI is a spectral imaging
technique distinct from focal plane array (FPA) FTIR as it operates on
principally different instrumentation. FPA-FTIR spectrometers pre­
dominantly measure spectra in the mid-infrared region (4000–400
cm− 1) using an infrared beam and interferometer to generate an inter­
ferogram (Smith, 2011). A Fourier transform is then applied to resolve
wavelengths, thereby producing a spectrum (Smith, 2011). Point mea­
surements are repeated over a given area creating a spectral map on a
pixel-by-pixel basis. By contrast, hyperspectral imagers (HI) can operate
across the electromagnetic spectrum including the ultra-violet, visible
and IR regions (ElMasry and Sun, 2010). For applications in the IR, the
NIR region is often used as it contains absorption bands for various
organic structures. It is a non-destructive technique which can be used in
reflectance or transmission mode, both however relying on diffuse light
reflectance or transmission. Halogen lamps or natural light are therefore
used as an IR source rather than an IR beam (ElMasry and Sun, 2010).
Most NIR-HSI instruments used for MP analysis operate using a line-
scan system (Faltynkova et al., 2021), which is a diffuse reflectance
imaging technique collecting one “line” of pixels at a time. Diffraction
elements and sensor arrays separate wavelengths in space, allowing
multiple spectral measurements to be made simultaneously for the
entire “line” (ElMasry and Sun, 2010). By moving the camera while
imaging, multiple line scans are stitched into a three-dimensional
spectral map, analogous to those produced using FPA-FTIR. The pri­
mary differences between methods are faster acquisition by NIR-HSI and
sampling of a different region of the IR spectrum.
While preliminary work has shown that NIR-HSI can be used to
identify different polymers, further validation is required to assess the
performance of this technique. Limits of detection, false positive and
false negative discovery rates have been explored with model materials
to some extent, but the accuracy of particle count and size estimates has
not been fully explored. In addition, the impact of plastic weathering on
accuracy of the analysis is also unknown. Importantly, very little
reference data is available (Knaeps et al., 2021), and is likely of limited
value across instruments and modes of acquisition. In addition, estab­
lished workflows going from single pixel identification to particle
identification are missing. These factors prevent the uptake of NIR-HSI
2
Chemosphere 336 (2023) 139186
N negative
NIR near infrared
P positive
PC principal component
PCA principal components analysis
PE polyethylene
PET polyethylene terephthalate
PP polypropylene
PS polystyrene
SIMCA soft independent model of class analogy
SWIR short wave infrared
TN true negative
TP true positive
techniques in the scientific community and a broader application in
routine analysis. Further development of this method could enable rapid
analysis of MP in a variety of settings but is perhaps most relevant for
monitoring schemes where high throughput is crucial.
In this study, we developed a spectral database and model to test the
accuracy and limitations of NIR-HSI as a method for the analysis of MP.
The database contains polymer spectra taken from a variety of plastic
sources and is used as training data to develop a chemometric model
which classifies pixels from hyperspectral images. To enable batch
processing of images for routine analysis, the chemometric model is then
embedded in an automated workflow which produces images ready for
particle characterization analysis. Evaluation of this workflow is done
using model MP prepared in the laboratory to assess how the composi­
tion and preprocessing of the spectral database affects model perfor­
mance. To assess the accuracy of the method, fluorescence microscopy
of polystyrene (PS) MP is used to control for particle number and size
estimates. The performance data is reported using sensitivity, speci­
ficity, and accuracy. Finally, we also tested to what extent photooxida­
tion (simulated weathering) of plastics impacts the performance
parameters mentioned above, thus assessing the viability of the analysis
pipeline for realistic environmental MP at different degrees of
weathering.
2. Materials and methods
2.1. Development of an NIR-HSI database
Reference materials for four polymer types: polypropylene (PP),
polyethylene (PE), polystyrene (PS) and polyethylene terephthalate
(PET) were collected to create a database of reflectance spectra. Three
categories of plastic were included: pre-production pellets (PS, Sigma-
Aldrich, 182427; PP, Sigma-Aldrich, 428116; high density PE and
PET, CARAT, Germany, no product num), plastic consumer products and
plastic debris collected from local beaches for all polymer types
(Table 1). Consumer plastics (n = 51) were sourced from multiple
countries (Table S1) as well as local sources (Trondheim, Norway).
Polymer type was identified by recycling codes, but also confirmed by
ATR-FTIR. Marine plastic debris (n = 196) was collected in two beach
locations, Hopavågen, Slettvik (63◦ 35′30.9″ N, 9◦ 31′36.3″ E) and Kors­
vika, Trondheim (63◦ 27′01.2″ N 10◦ 26′04.7″ E). The marine debris was
rinsed with ultrapure water to remove dirt and sand, but no further
cleaning was done. Due to differences in production and occurrence in
the environment, the number of unique samples of marine debris varied
for each polymer type (Table 1). Polymer type for the plastic consumer
products and debris was confirmed by Raman spectroscopy (Table S2).
NIR-HSI spectra were recorded by imaging reference materials with a
Hyspex SWIR-320me hyperspectral imager (hereafter HI) using the
provided macro lens. Two Hyspex halogen lamps which emit strongly in
A. Faltynkova and M. Wagner Chemosphere 336 (2023) 139186

Table 1 model training and calibration.

Content of the spectral database. (PE: polyethylene, PET: polyethylene tere­
phthalate, PP: polypropylene, PS: polystyrene). 2.2. Data analysis pipeline
Polymer Source # Unique sources # Spectra

PE Pristine 1 2000
All spectra were preprocessed using first derivative followed by
Consumer products 17 204 standard normal variate. Window size and smoothing type (moving
Marine plastic debris 120 1440 average or Savitsky-Golay) were optimized iteratively with no clear
Sum PE 3644 optimal window size (Fig. S2). However, some form of smoothing,
PET Pristine 1 2000
regardless of window size and filter type, gave better performance
Consumer products 11 132
Marine plastic debris 11 132 metrics than no smoothing. Based on these results, we chose a window
Sum PET 2264 size of seven with a moving average filter to smooth all spectra. The
PP Pristine 1 2000 spectral database was split into a validation set (20%) and a training set
Consumer products 12 144 (80%) using random sampling to build the soft independent model of
Marine plastic debris 62 744
Sum PP 2862
class analogy (SIMCA) using the R package mdatools. SIMCA uses in­
PS Pristine 1 2000 dependent principal component analyses (PCA) to create a supervised
Consumer products 11 207 classification model which assigns membership of an unknown sample
Marine plastic debris 3 36 to one, multiple or none of the classes. The training set was divided by
Sum PS 2275
class, in this case polymer type, thus four different PCA models were
Total spectral database 11045
made. The number of components for each PCA was chosen indepen­
dently by inspecting a scree plot. Components which substantially
NIR (Full, 2023), were used as a light source. To measure the spectral increased cumulative explained variance were kept in the model. PCAs
output of the light source, a reflectance standard was imaged alongside were validated using random repeated cross-validation (5 segments, 5
the samples. The reflectance spectrum obtained from the reflectance repeats). The four PCA models were then combined into the SIMCA
standard is later subtracted from all reflectance spectra. classification model. The decision boundary for the SIMCA models used
The HI is a “line-scan” camera moving perpendicularly to a thin slit here is the statistical significance parameter called alpha (value between
opening behind the fore optic (lens). Multiple slit images are then 0 and 1 representing the probability of false negatives), here set to 0.05.
stitched together to create a cohesive photomosaic. The resulting image Critical limits are calculated using the data driven moments approach.
or datacube has 3 dimensions, two spatial and one spectral. The HI used To classify whole datacubes, the spectrum associated with each pixel
in this study has a fixed line scan size of 1 × 320 pixels while the number was classified by the SIMCA model. The ID assigned to each pixel was
of line scans stitched together can vary. The third, spectral dimension of rasterized to create a mask image which allows localization and size
the datacube contains 256 wavelengths from 962 to 2493 nm (10 estimation of MP. One binary raster image was created per polymer
395–4011 cm− 1) with a spectral resolution of 6 nm. The linear field of type, allowing differentiation between polymers in addition to size and
view (FOV) is 84 mm at a working distance of 0.3 m. Pixel size across-/ shape information. Datacube classification was carried out on a local
along-track at this working distance is 0.22/0.22 mm. We fixed the high performance computing cluster (Själander et al., 2019). Raster
image size to 1800 lines × 320 pixels to acquire images of reference images were exported as.tiff files and analyzed in ImageJ for particle
materials at a working distance of roughly 5 cm, corresponding to pixel counting and size estimation as described by Prata et al. (2019).
size of roughly 53 × 53 μm (scan time ~45 s). The resulting scanned area
was 1.7 × 9.5 cm. 2.3. Internal and external model validation
Reference materials were imaged on glass fiber filters (Whatman GF/
F, 47 mm diameter, WHA1825047) which were taped to a black paper Internal and external validation was carried out to test the perfor­
square. Pre-production pellets (>1 mm diameter) were placed directly mance of the SIMCA model. Random repeated cross-validation (5 seg­
on the filters, while consumer plastics and debris items were cut into ~1 ments, 100 repeats) was used as the internal validation to determine
cm2 pieces and arranged on the filters to avoid overlap. Using the ENVI performance metrics sensitivity, specificity, and accuracy.
software (2022, L3Harris Geospatial Solutions, Inc.), hyperspectral im­ As an external validation set, a subsample of MP from a marine water
ages were visualized in false color to locate plastic particles. For pre- sample (ML3204) collected by The Ocean Cleanup Foundation was
production pellets, subsections of the image containing plastic, imaged using the same approach described for the reference database.
referred to hereafter as regions of interest (ROI), were extracted sys­ The sample originated from a surface trawl, the details of which are
tematically from a variety of pellets across the entire FOV to account for described by Egger et al. (2020). The spectra from these MP were not
variation in surface angle and incident infrared radiation. A total of used in the database indicated in Table 1, and therefore not included in
2000 spectra were taken from each pre-production polymer. For con­ any model training or calibration. We used two size classes from the
sumer plastics and marine debris, ROI were selected from the hyper­ ML3204 sample, 1.5–5 mm and >5 mm. From the size class >5 mm, we
spectral image by systematically sampling twelve blocks of 2 × 2 pixels imaged all particles and confirmed polymer type with ATR-FTIR. For the
over the visible surface of each particle. The ROI spectra were then smaller size class, we subsampled approximately 300 particles for
exported, and all subsequent analysis was conducted using R (version NIR-HSI analysis, also confirming the polymer type with ATR-FTIR for
4.0.2) and RStudio. Packages and code used in the analysis can be found each particle.
in the code repository (https://github.com/afalty/IR-HSI-Microplastics
-analysis-pipeline). 2.4. Accuracy of size estimation
The final database was assembled by averaging the blocks of 4 pixels
from each unique sample (marine debris and consumer products only). We produced fluorescent PS MP as a model material to compare
The number of spectra per particle was therefore reduced from 48 to 12. detection by NIR-HSI and SIMCA with fluorescence microscopy (ground
All spectra in the database were manually inspected for obvious aber­ truth) focusing on MP number and size estimates. To create these par­
rations. Twenty-six spectra were removed due to no signal, sensor ticles, fluorescent PS from neon party cups (partyking.no) was cryo-
saturation and/or clear error (Fig. S1). The remaining spectra were milled and sieved (Retsch woven wire mesh sieve) to produce parti­
normalized by subtracting the reflectance spectrum of a Spectralon cles in size ranges of <45 μm, 45–125 μm, 125–200 μm, 200–300 μm,
reflectance standard. Each spectrum was labelled with its confirmed 300–500 μm, 500–1000 μm and >1000 μm. Particles were rinsed on
polymer ID and origin. The spectra in this database are further used for stacked sieves and washed with a detergent (Tween-20, 1% solution) in

3
A. Faltynkova and M. Wagner
ultrapure water (ELGA Purelab) followed by three rounds of rinsing with
ultrapure water. 0.5–1 g of each size fraction was collected into a 50 mL
glass vial and suspended in 10–20 mL ultrapure water. The volume of
ultrapure water was adjusted as needed to achieve an appropriate par­
ticle concentration to hinder particle aggregation and overlap at the
later filtering stage. In triplicate, 1 mL of the diluted MP suspension was
filtered onto a 25 mm GF/F glass fiber filter for size classes ≤500 μm. For
the two largest size classes, aliquots of 3 mL were used. Blanks were
prepared in triplicate alongside the samples to control for contamina­
tion. After filtration, filters were stored covered in watch glasses until
imaging analysis.
After air drying in the loosely covered watch glasses, filters were
imaged on a Zeiss EMS3/SyCoP3 microscope using a PlanNeoFluar
Z1.0x objective, 7× magnification with filters 559–585 and 600–609 nm
(reflector: 63 HE Red Fluorescent Protein). An entire 25 mm filter could
be visualized in one image at this magnification. Microscopy images
were analyzed using the same particle analysis pipeline as for the raster
images generated by the SIMCA model, the only exception being that the
microscopy images were converted to 2-bit depth and a threshold was
manually chosen before particle analysis. The same samples were then
imaged by NIR-HSI (as described in 2.1) to directly compare results.
Particle size estimates were compared using a paired t-test by matching
size estimates for individual particles between fluorescence microscopy
and NIR-HSI. For reasons of practicality, a cut-off of ~1000 μm was
chosen to carry out individual particle comparisons.
Throughout the procedures described above, various quality assur­
ance measures were implemented to reduce contamination. Cotton lab
coats were worn, and synthetic clothing avoided when carrying out lab
work. Gloves were worn on occasion when necessary for chemical
handling. Glassware was used instead of plastic where possible and all
glassware was washed three times in ultrapure water before use. Ul­
trapure water was used exclusively to wash equipment or make solu­
tions. Filtration was carried out in a laminar flow hood, and all samples
were covered under transportation.
2.5. Assessing impact of photooxidation on detection of plastic by NIR-
HSI
We exposed plastic consumer products to UVA/B to simulate a nat­
ural weathering process. These UVA/B exposed plastics were then
analyzed by NIR-HSI to assess if simulated weathering affects the
spectral signal, which in turn could decrease model performance. Ex­
amples of changes which could impact the spectral signal include but are
not limited to the presence/absence of broad peaks due to incorporation
of oxygen through photooxidation, changes to the surface properties
which impact diffuse spectral reflection, or changes to the signal to noise
ratio. Novel items not used in the database were collected and cut into
~1 × 1 cm squares and varied in thickness from 0.25 to 1.5 mm. Four
polymers, corresponding to the polymers in the database (PP, PE, PS,
PET), were tested using three different products of each polymer type
(hereafter 1, 2 and 3). The three unique samples of each polymer type
were used to account for variation in additives and chemical composi­
tion which may affect photooxidation processes. A glass Petri dish
containing all twelve samples (four polymer types, three products of
each) was made for each time point along with one Petri dish used as a
dark control. The control samples were stored in the dark at room
temperature until analysis. All samples began exposure on day 0 and one
dish was removed every 14 d according to the sampling schedule
(Table 2). After removal, exposed samples were also stored in the dark at
room temperature until analysis.
For UVA/B exposure, we used an irradiation chamber (BS-02,
Opsytec Dr. Gröbel) set to maximum irradiance in both channels giving a
UVA irradiance of 4 mW/cm2 and UVB irradiance of 2 mW/cm2 in the
250–400 nm range (see Fig. S3 for spectral output). Based on calcula­
tions by Sait et al. (2021), we calculated that one day of exposure in the
chamber (combined UV irradiance of 6 mW/cm2) corresponds to
4
Chemosphere 336 (2023) 139186
approximately 8.76 days of average European solar irradiance (see
supplementary calculations S1). The UVA/B exposure was carried out
over eight weeks with sample removal after 2, 4, 6 and 8 weeks. The
estimated natural weathering times for these exposures vary from 0 to
1.5 years (Table 2).
We used the carbonyl index (CI) over the course of the UVA/B
exposure to monitor photooxidation. ATR-FTIR measurements were
made with a Bruker Alpha Platinum ATR-FTIR instrument. Spectra were
measured in triplicate and averaged, followed by CI calculation using
the specified area under band (SAUB) method as per Almond et al.
(2020). Weathered samples were then imaged by NIR-HSI as described
above. The hyperspectral images were analyzed using the SIMCA model
to determine the number and size of MP detected for each size fraction.
Prior to analysis, pixel coordinates containing the weathered plastic
samples were delineated based on false color images generated by the
hyperspectral imager. The x, y coordinates of all pixels containing
plastics were used as a ground truth to compare with the model output.
3. Results and discussion
Initial work to optimize the model included model validation with (i)
internal cross validation, and (ii) an environmental sample not used in
the database (described in Table 1) used as an external validation set. All
performance metrics (sensitivity, specificity and accuracy for internal
validation and sensitivity for external validation) were >80%. It should
be noted that no PET or PS samples were in the external validation set,
limiting the scope of these results to PP and PE. In addition to these
metrics, size comparisons between fluorescence microscopy and NIR-
HSI were not significantly different for PS MP > 1000 μm, showing
that NIR-HSI was accurately characterizing PS MP > 1000 μm. Finally,
surface photooxidation did not have a consistent impact on detection of
plastic by NIR-HSI and SIMCA.
3.1. Internal validation of the SIMCA model
Inspection of the scores plots for each PCA showed some subgrouping
and general trends. For PE, pre-production pellets are clustered together
at one end of the first principal component (PC), with consumer prod­
ucts predominantly on the opposite end of the first PC (Fig. 1A). This
variation could be driven by general surface reflectance properties or
chemical additives which differ between pre-production pellets and
plastic materials that have been modified for use in products. Marine
debris items span the entire first PC indicating that there is more vari­
ation in the samples collected from the environment, and that this
variation encompasses the characteristics of both pre-production pellets
and plastic consumer products. PE samples were abundant among the
marine debris, providing a larger and more heterogeneous spectral
database. This is also true for PP (Fig. 1B), which was the class with the
second most abundant marine debris items in the database. PS (Fig. 1C)
and PET (Fig. 1D) are less abundant in the environment (Kannankai
et al., 2022), so there are fewer marine debris samples for these classes.
Less heterogeneity and fewer individual particles are reflected in the
PCA as both PET and PS scores plots contain distinct subclusters. Sub­
clusters are groups of multiple spectra taken from the same particle
(Fig. S4). We can presume that adding more unique samples to these
Table 2
UVA/B exposure scheme with equivalent natural weathering time. Esti­
mates are based on mean European solar irradiance.
Sample Time in chamber Estimated natural weathering time
0 0d none
1 14 d 17.6 weeks/4.4 months
2 28 d 35.1 weeks/8.7 months
3 42 d 52.7 weeks/13.2 months/1.1 years
4 56 d 70.3 weeks/17.6 months/1.5 years
A. Faltynkova and M. Wagner Chemosphere 336 (2023) 139186

Fig. 1. PCA scores plots showing marine debris (red), consumer products (blue) and pre-production pellets (green). The PCAs contain all spectra in the
database described in Table 1, separated by polymer type. A) PE (polyethylene) B) PP (polypropylene) C) PS (polystyrene) and D) PET (polyethylene terephthalate).
(For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

classes would help reduce the impact of distinct particles, as seen with
PP and PE, creating a more representative database.
The numbers of PCs for the PCA models were 5, 4, 5 and 3 for PE,
PET, PS and PP, respectively. Including more components in the PCA
may lead to overfitting, however since each class is a heterogeneous mix
of materials from various sources, more PCs may be required to explain
the variation in each class (De Maesschalck et al., 1999). We therefore
included PCs adding only marginally to cumulative explanation of
variance.
There is considerable overlap between the three categories of source
materials (pellets, consumer products and marine debris) which sup­
ports the assumption that all plastics of the same polymer type can be
classified correctly despite differences in color, surface morphology or
degree of weathering. This can also be confirmed by a PCA including all
four polymer types (Fig. S5).
Individual PCA models were then combined into the SIMCA to create
a supervised classification model. Using 20% of the database as an in­
ternal validation set, the SIMCA model shows an overall high perfor­
mance, that is, the model correctly predicts the polymer type of the
samples in the validation set in >93% of cases (Fig. 2). Since the SIMCA
model operates on a binary decision boundary for each class, spectra
which do not belong in any of the polymer classes are labelled as “none”.
This is a great advantage of the SIMCA model as it precludes the need to
collect spectra for different non-plastic materials or media. In general,
plastics were slightly more often misclassified as not being plastic than
being another polymer type (see Fig. 3).
To further assess the model quality, the metrics of sensitivity (true
positive rate), specificity (true negative rate), and accuracy were
assessed using random repeated cross validation. (Table 3). For each
iteration, a random 80% subset of the database was used as a training set
to build the model and the remaining 20% used for validation. Average
values for all polymers indicated that overall performance was high

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A. Faltynkova and M. Wagner Chemosphere 336 (2023) 139186

3.2. External model validation with environmental plastics

As an external validation set, we used a manta trawl sample collected

Fig. 2. Confusion matrix for internal validation of the SIMCA model
showing the true and predicted class for an internal validation set (n ¼
2209). “None” represents the classification in none of the four polymer classes.
(>0.88, Table 3). Sensitivity was slightly lower than specificity indi­
cating that false negatives (type II errors) are more frequent than false
positives (type I errors).
This internal model validation agrees with previous research which
shows that spectral signatures for PP, PE, PET and PS in the SWIR are
characteristic enough to achieve separation by supervised analysis
(Serranti et al., 2018; Shan et al., 2019; Karlsson et al., 2016; Vidal and
Pasquini, 2021; Ai et al., 2022). PE was the primary contributor to
decrease in sensitivity (i.e., PE spectra being classified as “none” or
another polymer type). Since the accuracy for PE is high (0.971), the loss
of sensitivity can be attributed primarily to false negatives for PE spectra
rather than confusion of PE spectra with another polymer type. The PE
group contained the most unique training samples, and therefore the
most variation. This could indicate that some PE MP had unique fea­
tures, either due to unique peaks in the spectra or noisy spectra from
dark MP. Further optimization such as adjusting the alpha value could
help reduce false negatives but may also lead to more false positives.
Interestingly, performance for the PET and PS classes is not relatively
poorer than PE or PP despite their databases containing fewer spectra.
This could indicate that PET and PS have more distinct spectra, a logical
conclusion given the presence of heteroatoms, functional groups and
aromatic structures absent in PP and PE. Another explanation could be
that there were fewer black or dark particles in these classes as
compared to PP and PE, thus a lower false negative rate.
from the Great Pacific Garbage Patch. We used 359 particles from the
sample and calculated sensitivity based on identification of particles
rather than pixels, since particle identification is the main goal of the
analysis. Particles were automatically counted using ImageJ and an
additional manual control was performed to ensure small fragments or
partial detection leading to segmentation did not inflate the particle
count. Performance for the external validation was slightly lower, but
still >80% (Table 4). Since only PE and PP MP were present in the
external validation sample, so no performance parameters could be re­
ported for PS or PET MP. PE gives a better indication of the external
performance results, as we analyzed a total of 330 PE MP whereas there
were only 29 PP MP.
To further inspect sources of error, results from the NIR-HSI analysis
and SIMCA model were overlayed on an RGB representation of the
hyperspectral image (Fig. 3). The images show that false negatives
occurred predominantly with black MP, as has been reported in other
studies (Zhu et al., 2020; Huang et al., 2021). We observed this trend for
both size classes where 73% and 90% of the false negatives were black
MP for the >1.0 mm size class and >0.5 cm class respectively. Other
false negatives arose due to colonization of the surface of MP by calci­
fying organisms or very thin films which did not produce a strong
enough signal. These phenomena also contributed to some MP being
partially detected, leading to segmentation errors. Changes to the clas­
sification model, such as adding multiple thresholds or scoring based on
adjacent pixels, could reduce these errors. These strategies are used in
the common analysis software siMPle (Primpke et al., 2020a). Adapting
this model to use a similar framework could also help to harmonize
results between NIR-HSI and FPA-FTIR as FPA-FTIR is not suitable for
analysis of larger MP (≥300 μm) in transmission mode. Reflection mode
can be used but produces poorer spectra relative to ATR-FTIR which is
the more common method for MP > 300 μm.
Only one particle we imaged gave a signal for both PE and PP. ATR-
FTIR analysis confirmed the particle to be PE, so this particle was
counted as a false positive for PP (Fig. 4, center of image). We were
unable to establish the reason for this false positive as multiple ATR-
FTIR spectra were collected and classified as PE (>80% match),
without any obvious aberrations or unique features. Confusion could be
the result of PP and PE overlapping in the PCA space as they have most
similar spectra of the four polymer types tested (Fig. S5). Despite these
errors, sensitivity remained >80% and was as high as 93% for the largest
group of PE particles, indicating that the performance is good for envi­
ronmental MP.
One key learning that emerged from development of the model was

Table 3
Average performance metrics for cross validation of the SIMCA model ± standard deviation. Random repeated cross validation was done using 5 segments
and 100 repeats. Score of 1.0 indicates no errors. TP = true positive, TN = true negative, FP = false positive, FN = false negative, P = positive, N = negative.
Polymer Sensitivity TP/TP + FN Specificity TN/FP + TN Accuracy (TP + TN/P + N)

PE 0.89 ± 0.00 1.00 ± 0.00 0.97 ± 0.00

PET 0.97 ± 0.00 1.00 ± 0.00 0.99 ± 0.00
PP 0.98 ± 0.00 1.00 ± 0.00 0.99 ± 0.00
PS 0.97 ± 0.00 1.00 ± 0.00 0.99 ± 0.00
All 0.95 ± 0.00 1.00 ± 0.00 0.99 ± 0.00

Table 4
Performance data for model prediction based on the external validation set with microplastics collected from the marine environment and imaged using
NIR-HSI. Model prediction using SIMCA. Data is reported based on number of particles.
Size class Total particles (Ground truth) Total particles (HSI detection) Sensitivity TP/TP + FN

PE PP PE PP PE PP
>0.5 cm 45 5 36 6 0.8 1
>1.0 mm 285 24 265 20 0.93 0.83

6
A. Faltynkova and M. Wagner
Fig. 3. Overlay of prediction results of the SIMCA classification model onto an RGB representation of the hyperspectral image using microplastics >0.5 cm
from an environmental sample. False negatives for entire particles occurred primarily with black MP. Some false negative pixels can be seen creating holes within
particles, and only one particle shows pixels identified as a mix of polypropylene (PP) and polyethylene (PE).
the importance of the composition of the database on external validation
performance. Internal cross validation tests gave very good results
regardless of the composition of the database, that is, including or
excluding consumer products and weathered samples made little impact
(Table S3). However, external validation on weathered samples from
outside the database showed almost no detection when only the pre-
production pellets were used to train the model. Performance
improved with the addition of consumer products, and further improved
by including marine debris (Fig. S6). Similar findings by Vidal and
Pasquini (2021) underscore the importance of including weath­
ered/environmental materials when creating training datasets for su­
pervised classification.
To the best of our knowledge, all studies using NIR-HSI have
collected their own training data by imaging model plastic materials,
either pristine or from the environment. This is a time-consuming step
and demands that many unique plastic samples are collected for a range
of polymer types. Especially for samples coming from the environment,
this requires collecting considerable amounts of plastic to ensure plastics
of many different polymer types are represented. This is a somewhat
inevitable step, as transferring calibrations from one instrument to
another is a well-recognized challenge. Here, combining training data
from various instruments can be a solution (Fearn, 2001) that can be
tested once more open datasets become available. Further research with
a focus on calibration transfer could facilitate identification of spectra on
online platforms such as OpenSpecy (Cowger et al., 2021) where data­
sets are crowd sourced. Ultimately, this could reduce the work required
to use NIR-HSI for high throughput analysis. For this reason, we have
made our training database, model and analysis pipeline available on
Github (https://github.com/afalty/IR-HSI-Microplastics-analysis-pipeli
ne).
3.3. Particle detection and size estimation
We created a “ground truth” for size data by analyzing model MP
using fluorescence microscopy. Adjusting the threshold manually when
7
Chemosphere 336 (2023) 139186
creating binary images allowed us to correct for artefacts of fluorescence
microscopy (e.g., blurred edges, holes in particles) ensuring that our
ground truth was as accurate as possible. In addition, the fluorescence
microscopy images have a pixel size of 6.49 × 6.49 μm, which is smaller
than the upper limit of the smallest model MP (45 μm). Especially when
measuring particles ≫45 μm Feret diameter, we assume the error
associated with these size estimates to be negligible. We are therefore
confident that the size estimates from fluorescence microscopy are a
suitable ground truth to evaluate the accuracy of the size estimates made
by NIR-HSI.
When comparing MP size distributions between fluorescence mi­
croscopy and NIR-HSI, there were clear deviations in particle size esti­
mates for the smallest size classes (<500 μm). NIR-HSI analysis did not
detect any MP in size classes <300 μm. In the 300–500 μm size class,
only 60% of particles (66 of 110) were detected. Notably, the distribu­
tion for this size class is heavily skewed towards smaller particles
(Fig. 4A). This indicates that only a few pixels were detected per particle,
and that NIR-HSI with the SIMCA model underestimates the actual size
of MP 300–500 μm. This was confirmed visually by comparing the
SIMCA results with the fluorescence microscopy for PS (Fig. 4B). The MP
size distributions for the two largest size classes (500–1000 μm and
>1000 μm) were more comparable. Size distributions between fluores­
cence microscopy and NIR-HSI begin to align above 500 μm, indicating
that NIR-HSI can detect and characterize entire particles. The spatial
limit of detection (LOD) for this instrumental setup and prediction with
SIMCA was thus taken to be 500 μm, which is within the range of other
LOD reported (14.8–1000 μm) for instruments used to analyze MP
(Faltynkova et al., 2021).
The spatial resolution of the HI is the largest determinant of the LOD
as it determines how large an area is covered by one pixel. For the im­
aging parameters used in this experiment, one pixel covered 53 × 53 μm.
Theoretically, it could be possible to detect (but not characterize) a MP
particle of 50 μm Feret diameter. However, the particle would need to be
centered perfectly within one pixel and reflect a strong enough signal to
be detected. Meeting these criteria is unlikely, resulting in small MP
A. Faltynkova and M. Wagner Chemosphere 336 (2023) 139186

Fig. 4. Size estimation of microplastics by the NIR-HSI and SIMCA model compared to fluorescent microscopy. A) Comparison of size distributions for the
three largest size classes. Smaller size classes had no data for NIR-HSI as there was no detection. B) image analysis comparing the ImageJ thresholding of fluorescence
microscopy images (pink) and the results of the SIMCA model (blue). Both sets of images were analyzed using ImageJ to produce data on particle number and size.
(For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 5. Correlation of size estimates for MP between fluorescence microscopy (x-axis) and NIR-HSI (y-axis). Red lines show a linear model fit to the size
correlation and corresponding equation showing slopes deviating slightly from 1 (y = x, grey line). This indicates that the size accuracy is consistent over the size
range from 1000 to 7000 μm. A) Estimated area of MP based on fluorescence microscopy and corresponding estimate from NIR-HSI for matched particles (n = 22). B)
Estimated Feret diameter of the matched particles. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of
this article.)

8
A. Faltynkova and M. Wagner
being only partially covered by any given pixel. The signal from the MP
is therefore mixed with the filter background or other materials present
in the same pixel. Once MP are large enough to reflect a signal in mul­
tiple adjacent pixels, detection becomes more likely, and quantification
becomes possible. As shown here with particles larger than 500 μm, if
pixel size is substantially smaller than MP size, good detection and
quantification can be achieved. This principle should apply for MP
smaller than 500 μm if the pixel size decreases in accordance. This is
demonstrated by Zhu et al. (2020) who were able to detect MP down to
100 μm (pixel size 14.8 μm) by using an extension tube together with the
hyperspectral imager. The LOD and LOQ is therefore limited – at this
stage – by instruments which are generally made to image larger objects
(>1 cm).
When comparing dimension estimates per particle (n = 22), MP size
estimates between NIR-HSI and fluorescence microscopy were well
correlated over the tested size range (Fig. 5). Size estimates were nor­
mally distributed in both groups, as tested with a Shapiro-Wilk test
(SIMCA p-value: 0.526, fluorescence p-value: 0.967) allowing us to
apply a paired t-test. There were no significant differences between es­
timates for both Feret diameter (mean Δ = − 42.3 μm, confidence in­
terval: 337.6, 253.0, p = 0.768) and area (mean Δ = 390276 μm2,
confidence interval: 95989, 876540, p = 0.11) when comparing NIR-HSI
and fluorescence microscopy. This demonstrates that NIR-HSI can be
used not only to detect particles and determine their polymer type, but
also to accurately characterize PS MP > 1000 μm using common size
metrics like Feret diameter and area. Characterization of morphological
parameters by NIR-HSI has been shown by Serranti et al. (2018), how­
ever quality of these estimates was based on overall pixel-wise pre­
dictions by partial least squares discriminant analysis (PLS-DA) as
opposed to measuring the same morphological parameters with another
imaging technique.
3.4. Quality control
The quality control includes assessment of fluorescence microscopy
imaging as a “ground truth” and sources of error, followed by evaluation
of blanks to detect contamination.
The PS MP size distributions (based on fluorescence microscopy es­
timates) were generally larger than the desired size range based on the
sieving limits (Fig. 4A). This could be due to particle morphology with
long, thin particles passing through sieves lengthwise, as has been
documented for fibers (Schönlau et al., 2020). Another source of error
was aggregation of particles in the correct size range, which caused
multiple particles to be counted as a single particle (Fig. S7). This is a
known source of error which can impede accurate quantification of MP
(von der Esch et al., 2020; Kühn et al., 2018). Use of detergents or
organic solvents, as well as reduction of particle concentration can
reduce aggregation (von der Esch et al., 2020; Kühn et al., 2018).
However, aggregation was primarily problematic for size fractions
<125 μm. Since no signal was detected with NIR-HSI for PS MP < 200
μm, even at high MP concentrations, we chose not to further optimize
the MP concentration for size classes below 125 μm.
Despite lower MP concentrations for larger size classes, some PS MP
> 300 μm were still too close to other PS MP on the filter such that they
were counted as one particle. When comparing particles directly be­
tween the two analyses, this resulted in no data for some particles, and
erroneous overestimates for other particles. This source of error was not
of interest in this study, and thus missing or aggregated particles were
removed for the t-test and size correlation for particles >1000 μm.
Examination of the blanks under higher magnification (100×)
showed minimal contamination of MP from the laboratory. Particle
count for blank samples arose primarily from noise at the edges of the
filter where no solution was filtered (Fig. S8A). To eliminate this source
of noise from the samples with PS MP, microscope images were cropped
to the exposed filter area which was used for filtration of particles in
solution. In addition, a lower size cut-off of 100 μm2 was used when
9
Chemosphere 336 (2023) 139186
analyzing particle size and number for size ranges <45 μm, 45–125 μm
and 125–200 μm. This effectively reduced false positives due to high
signal/noise ratio in images where the fluorescence signal was relatively
low (i.e., small PS MP with weak fluorescence). In addition, the number
of particles detected in the blank samples (n = 5, see Figs. S8B–F) was
insignificant compared with model MP samples. No corrections were
therefore made to the overall particle counts for model MP samples.
3.5. Impact of plastic photooxidation on model performance
CI and visual inspection were used to track photooxidation caused by
UVA/B exposure. Trends in CI over the course of the exposure were
generally similar within each polymer type, and all polymers showed
increased CI (Fig. S9) or visual signs of photooxidation throughout the
experiment. For further discussion of photooxidation see the supple­
mentary information (S2).
Evaluation of NIR spectra throughout the photooxidation experiment
do not show any apparent changes (Fig. 6). Differences in amplitude can
be seen in the light-source corrected spectra, however these differences
are corrected by the preprocessing steps described in 2.2. Visually
inspecting the uncorrected spectra did not reveal any distinguishable
changes in major peaks. It stands to reason that oxidation processes
could be detected in NIR spectra as bonds such as C–O, O–H, C=O and
COOH do typically absorb strongly in the NIR. However NIR spectra are
difficult to interpret qualitatively due to the presence of weak overtone
bands (Bart, 2006).
A PCA was used to further assess changes in NIR-HSI spectra due to
photooxidation. It is clear from the scores plot that the polymer type is a
stronger driver of spectral characteristics than the photooxidation stage
(Fig. S5). There are no apparent trends by photooxidation stage, neither
within nor between polymer types. Nonetheless, no visual changes in
spectra and no trends in the PCA indicated that the photooxidation
protocol used in this study did not induce any characteristic changes in
the NIR spectra of the plastics tested. This contrasts with the FTIR results
where oxidation was detected for the weathered PP and PS samples.
Depth of penetration of the NIR radiation is likely a contributing
factor for this difference, as ATR-FTIR analysis (mid-IR, 400–4000
cm− 1) is constrained to the first few μm of the surface (Nagai et al.,
2005) capturing only the oxidized outer layer. In contrast, NIR radiation
(12 820–4000 cm− 1) penetrates to a depth of 1–4 mm in reflectance
mode (Bart, 2006). For samples of this size (~1 cm2, ~1 mm thickness),
the oxidized outer layer is likely a small fraction of the bulk material due
to oxygen diffusion limitation (Delprat and Gardette, 1993). Shyichuk
and White (2000) performed a similar experiment using PS exposed to
0.4 mW/cm2 UV radiation (compared to 6 mW/cm2 used here) for up to
6 weeks and determined that oxidation occurred primarily in the upper
0.5 mm of the surface. Due to the thickness of the samples used in this
study and the photooxidation protocol used, it is likely that pristine
material on the inside of the particle was contributing to the spectral
signature enabling detection by NIR-HSI. NIR-HSI could therefore be
used to detect spectra of weathered particles despite surface-layer
photooxidation. This reduces the possibility of using NIR spectra for
qualitative determination of surface weathering processes, however,
may offer advantages for analysis of larger MP where surface weathering
produces poor ATR-FTIR spectra. It should be noted the polymer iden­
tification by ATR-FTIR analysis was not hindered by the photooxidation
protocol used here. Since the extent of degradation is also dependent on
weathering conditions and not just thickness, more intense photooxi­
dation protocols which also include other important weathering pro­
cesses (i.e., mechanical, and biological degradation) could be used to
systematically determine characteristics of weathered NIR spectra.
Considering the importance of the bulk material as compared with
surface properties, this also demonstrates that thickness is an important
parameter that controls identification. Vidal and Pasquini (2021)
detected plastics with a thickness of 80 μm, but not 50 μm using
NIR-HSI. Thickness is connected to signal strength, so instruments with
A. Faltynkova and M. Wagner Chemosphere 336 (2023) 139186

Fig. 6. Impact of photooxidation on the preprocessed NIR-spectra of four polymers. The days indicate time of exposure in a UVA/B chamber. Spectra shown are
an average of 6 pixels taken from one of the three examples (sample 2) of each plastic and preprocessed using standard normal variate (SNV), 1st derivative and
smoothing. A: PE, B: PS, C: PP, D: PET.

more sensitive sensors will likely perform better in detecting very thin
MP.
Similar to photooxidation processes, other sources of interference
could be overcome by using NIR-HSI as found by Vidal and Pasquini
(2021) who identified MP on sand despite adherence of sand particles to
MP – the bulk material provided a strong enough signal to overcome
interference from sand particles. This could indicate that interference
from adsorbed organic matter or biofilms could also be negligible when
using NIR-HSI, however this is yet to be tested.
To confirm the performance of the model was also unaffected, per­
formance metrics of precision and recall were checked for each sample
over the time-course experiment. Photooxidation did not affect preci­
sion, which reflects the false positive rate (Fig. 7). Recall, which reflects
the false negative rate, varied more over the course of the experiment.
There was no clear, consistent trend in recall variation with respect to
time or polymer type. In some cases, a single sample within the polymer

Fig. 7. Impact of plastic photooxidation on the performance of NIR-HSI and the SIMCA model. In the binary images, the detection of three different examples
of each polymer type is included in the overall performance score. The binary images use white pixels to indicate a positively identified pixel.

10
A. Faltynkova and M. Wagner
groups was the source of decreased performance (PS-1), implying that
weathering could be negatively affecting model performance for specific
products.
In other cases, changes in recall could be more directly linked to
problems with detection as opposed to prediction by the SIMCA model.
For example, morphology played a large role in detection by NIR-HSI for
PP-2. The initially flat piece of plastic was deformed after photooxida­
tion causing the edges of the piece to curl upwards. The depth of field for
the HI is limited at higher spatial resolution, and thus some regions
(primarily edges) of the particle were out of focus during imaging.
Similar issues have been reported for Raman imaging with larger, 3-
dimensional particles or the simultaneous detection of small and large
particles in different vertical planes (Sobhani et al., 2019; Anger et al.,
2018). The problem of warping/deformation could be less relevant for
environmental MP as other forms of degradation (e.g., mechanical
stress) will affect morphology through fragmentation and abrasion, thus
breaking off or smoothing protrusions. In any case, the size of the MP
relative to the depth of field will play a critical role, suggesting that there
is an upper size limit for optimal performance for any given instrument.
While this presents an analytical limitation, it does not indicate that
chemical photooxidation leads to changes in chemical structure that
produce unrecognizable spectral signatures.
Other factors contributing to poor performance were entirely inde­
pendent of photooxidation, as seen in sample PE-1. NIR-HSI did not
reliably detect PE-1 at any stage, showing that false negatives can arise
from inherent properties of the plastic. Properties that hinder identifi­
cation could include the presence of additives which lead to a significant
change in the spectral signature. However, a more likely explanation in
this case is a low signal to noise ratio. Spectral reflectance will vary due
to surface characteristics and color of the plastic. The spectra from
sample PE-1 (not detected) contained more noise irrespective of
photooxidation stage (Fig. S10F). This was unexpected as noisy spectra
are typical for dark plastics due to high IR absorbance, however PE-1
was white in color. Inspection of the raw spectra show a reduced
signal as compared with the other PE samples, which might indicate that
the surface is simply less reflective. Surface characteristics like rough­
ness or even coatings could contribute to low signal to noise ratios. In
this case, we can see that some signal is still present in the NIR spectra
(Fig. S10C), but the SIMCA model did not classify this low signal as PE.
Interestingly, FTIR spectra of PE-1 also differ from the other two PE
samples. FTIR spectra show a broad signal between 750 and 500 cm− 1
absent in PE-2 and PE-3 (Fig. S11), which could indicate an additional
component present on the surface of the plastic. Including more diverse
plastics in the database would capture more variation in spectral sig­
natures which arise from these properties, however in the case of PE-1
the NIR spectra only showed reduced signal rather than a unique
peak. In this case, signal reduction caused either by surface properties,
coatings or additives was strong enough to “mask” or reduce the poly­
mer chain signal to the point where the model failed.
The false negative for this sample is likely due to a combination of
poor detection (NIR-HSI) and false prediction (SIMCA). Adjusting the
alpha value could increase the likelihood that poorer spectra are
correctly predicted by the model, but it also raises the risk of increasing
false positives. Averaging pixels could also help to smooth noisy spectra
without losing important information, but this would reduce spatial
resolution. Further optimization of the imaging set-up to increase signal,
combined with optimizing the predictive model (or trying other models)
could improve performance for challenging samples.
4. Outlook
Current analysis methods for MP > 500 μm include visual identifi­
cation and ATR-FTIR (Ivleva, 2021; Rocha-Santos and Duarte, 2015;
Roscher et al., 2021). Visual identification is highly accessible to re­
searchers, but accuracy is low compared to methods which allow
chemical analysis (Lenz et al., 2015; Song et al., 2015), and
11
Chemosphere 336 (2023) 139186
determination of polymer type is not possible. Particles must also be
identified individually, increasing time demand. This is also true for
ATR-FTIR which requires handling of particles for manual analysis and
often uses visual sorting as a preliminary step (Ivleva, 2021). Since most
MP sample preparations end with a filtration step, using a glass fiber
filter and imaging the filter directly with NIR-HSI eliminates the need to
handle individual particles. NIR-HSI could replace ATR-FTIR as a high
throughput method for analysis of larger MP providing polymer type
and size characteristics simultaneously for many particles and removing
the visual pre-sorting process. Data resulting from this pipeline is anal­
ogous to data output from siMPle (Primpke et al., 2020a) making
comparisons and harmonization easier. This makes analysis by NIR-HSI
comparable to imaging methods like FPA-FTIR which are commonly
used for analysis of MP down to 10 μm (Ivleva, 2021; Rocha-Santos and
Duarte, 2015; Primpke et al., 2021). Harmonization of data reporting
between smaller (<1000 μm) and larger (>1000 μm) size fractions
would increase comparability of results and reduce researcher bias for
size fractions of MP which are often analyzed separately.
As previously discussed, smaller MP (<500 μm) could potentially be
quantified by NIR-HSI if optical systems with higher spatial resolution
are used, and if enough signal is generated by thin particles. Higher
spatial resolution would likely increase analysis time to some extent,
requiring mosaicking of multiple hyperspectral images to analyze one
sample. However, since the image acquisition time is on the order of
seconds, increasing analysis time to a few minutes still offers a time
advantage over FPA-FTIR which requires 4 h to analyze an area of 14 ×
14 mm (Primpke et al., 2020b). Due to time and resource requirements
for analysis, imaging methods will often analyze a subsection of the
filter, extrapolating results to the entire filter area. Some techniques for
improving the representativeness of subsampling have been proposed
(De Frond et al., 2023) but subsampling will always introduce uncer­
tainty possibly leading to over/under-estimation (Brandt et al., 2021;
Schymanski et al., 2021; Thaysen et al., 2020). Errors based on extrap­
olation could be eliminated by using a faster method such as NIR-HSI
which allows analysis of the entire filter. Faster analysis would also
make replication and higher frequency sampling campaigns feasible,
further reducing uncertainties about temporal and spatial distribution of
MP in the environment (Kershaw et al., 2019).
This study contributes to a growing body of research which dem­
onstrates successful and rapid identification of MP using NIR-HSI
(Piarulli et al., 2020; Zhang et al., 2019; Serranti et al., 2018, 2019;
Shan et al., 2019; Karlsson et al., 2016; Vidal and Pasquini, 2021; Zhao
et al., 2018; Paul et al., 2019; Knaeps et al., 2021; Zhu et al., 2020;
Huang et al., 2021; Schönlau et al., 2020). Here, we present evidence
that NIR-HSI can not only reliably identify the polymer type, but also
provide accurate size estimates for MP > 1000 μm. No differences were
detected in NIR-HSI spectra after photooxidation, in contrast to FTIR
spectra where PP and PS showed clear indication of photooxidation. Due
to sample thickness and penetration depth of NIR-HSI, surface photo­
oxidation did not impact identification of different polymer types. This
is especially relevant for analysis of environmental samples, but also to
identify MP in studies where more realistic materials are being used to
elucidate MP toxicity, transport, interactions with biofilms and in­
teractions with environmental pollutants. NIR-HSI is therefore a prom­
ising technique for MP otherwise analyzed on a particle-by-particle
basis.
Author contributions
AF: Conceptualization, experimental work, data analysis & compu­
tational modelling, writing, reviewing, and editing of the manuscript.
MW: Methodology, supervision, writing, reviewing, and editing of
manuscript.
A. Faltynkova and M. Wagner
Declaration of competing interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Data availability
Data will be made available on request.
Acknowledgements
This work was supported by internal funding from the Norwegian
University of Science and Technology. The authors would like to
acknowledge Geir Johnsen for assistance in reviewing the manuscript,
The Ocean Cleanup for donating a microplastic sample and PlasticsEu­
rope donating plastic materials for inclusion in the database.
Appendix A. Supplementary data
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.chemosphere.2023.139186.
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