Trends in Analytical Chemistry 111 (2019) 229e238

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Trends in Analytical Chemistry

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Data preprocessing & evaluation used in the microplastics

identification process: A critical review & practical guide
Gerrit Renner a, b, **, Alexander Nellessen a, Alexander Schwiers a, Mike Wenzel a,
Torsten C. Schmidt b, Jürgen Schram a, *
a
Instrumental and Environmental Analytics, Niederrhein University of Applied Sciences, Frankenring 20, D-47798 Krefeld, Germany
b €tsstr. 5, D-45141
Instrumental Analytical Chemistry and Centre for Water and Environmental Research (ZWU), University of Duisburg-Essen, Universita
Essen, Germany

a r t i c l e i n f o a b s t r a c t

Article history: Characterising microplastics based on spectroscopic measurements is one key step of many studies that
Available online 15 December 2018 analyse the fate of microplastics in the environment. Over the years, many potential sources of error were
identified, which can be seen by the implementation of anti-contamination protocols, measuring labo-
Keywords: ratory blanks or using less aggressive chemicals for sample purification. However, the identification
Microplastics identification process itself in the meaning of a traceable and transparent documentation is hard to find in many
Preprocessing
research studies. Unfortunately, this can make it difficult to estimate if the presented results are
Library searching
representative and reproducible, as the evaluation process during microplastics identification depends
FTIR
Raman
dramatically on the performed data treatment. To increase the awareness of this often neglected topic,
this article reviews suitable data preprocessing and evaluation methods for microplastics identification
based on spectroscopic measurements and will recommend a practical workflow or check list that can
easily be used for further research studies.
© 2018 Elsevier B.V. All rights reserved.

1. Introduction However, the preprocessing and evaluation processes of mea-

The analytical process to identify microplastics in environ-
mental samples is hampered by numerous pitfalls that can affect
accuracy or informative value of a study [1e4]. The use of aggres-
sive chemicals during sample preparation [5,6], ignoring an anti
airborne fibres contamination protocol [7,8], identifying micro-
plastics in the micrometre range by visual inspection only [2,9] or
disregarding influences of aging effects and contamination by (bio)
organic matter [10e13] are some examples of the don'ts in this
analytical field. Fortunately, awareness for these impediments is
growing [14] and most research studies consider many analytical
aspects to ensure that the presented results are accurate and
representative. With respect to this, individual steps are almost
always described in a very detailed and transparent way.
* Corresponding author. Instrumental and Environmental Analytics, Niederrhein
University of Applied Sciences, Frankenring 20, D-47798 Krefeld, Germany.
** Corresponding author. Instrumental and Environmental Analytics, Niederrhein
University of Applied Sciences, Frankenring 20, D-47798 Krefeld, Germany.
E-mail addresses: grenner87@gmail.com (G. Renner), schram@hsnr.de
(J. Schram).
surement data are often insufficiently taken into account, and un-
fortunately, these steps are very important and critical aspects,
especially for techniques that deal with chemical imaging or a high
sample throughput. Modern analytical methods provide enormous
amounts of data that cannot be evaluated manually by experts but
have to be evaluated automatically by chemometric concepts.
Therefore, it is even more important to spend some time in
developing suitable and practicable data treatment strategies, and
additionally, to document those in the same detailed way as other
experimental steps.
The major part of microplastics identification is performed by
spectroscopic methods like infrared or Raman spectroscopy
[4,14e16]. These techniques are often connected with library
searching to evaluate measured spectra, however, the framework
parameters of this step are described significantly less detailed than
other steps within the analytical process. Even the choice of the
searching algorithm for hit quality index (HQI) calculation and the
interpretation of HQIs themselves are crucial, as there exist big and
relevant differences in their determination approaches [11,17]. In an
extensive literature investigation, one could observe that only 25%
of all studies mentioned the used evaluation concept, but

https://doi.org/10.1016/j.trac.2018.12.004
0165-9936/© 2018 Elsevier B.V. All rights reserved.
230 G. Renner et al. / Trends in Analytical Chemistry 111 (2019) 229e238
unfortunately, a more detailed description can be found in only 2%
of them [14]. Therefore, a very low level of awareness for data
evaluation can be deduced, and to that end, this article will present
and discuss common data preprocessing and evaluation concepts
for microplastics identification based on spectroscopic techniques.
Moreover, suitable and accurate data evaluation methodologies are
presented comprehensively, with the overarching aim to increase
the awareness for data evaluation during microplastics investiga-
tion. To that end, this article may help to choose a data pre-
processing for microplastics identification that fits the purpose.
2. Manual spectra inspection: the gold standard
Before focussing on the chemometrics of spectra evaluation to
investigate microplastics, a reliable reference method has to be
worked out. This offers the possibility of a validation, which is a
necessary step during data evaluation. Thereby, manual spectra
inspection has two main tasks. On the one hand, data quality can be
rated, and using this feedback, measurement parameters can be
adjusted (trouble shooting). On the other hand, spectra can be
evaluated based on the researcher's expertise. In case of micro-
plastics, the latter is supported by the fact that most plastics in the
environment are packaging materials like polyethylene, poly-
propylene or polystyrene [15,18,19]. This narrows down the
assignment choices to a manageable extent. In practice, manual
spectra inspection is subdivided into two steps: looking at the
characteristic shape of the whole spectrum, and characterising in-
dividual vibrational bands. However, high level of expert knowl-
edge is required, and as this method is very time consuming, it is
not useful for high sample throughput analysis [11,17]. Neverthe-
less, it is the best choice for validating other microplastics identi-
fication concepts.
3. Data preprocessing: how to treat spectra?
After measuring a vibrational spectrum of microplastics, in
many cases, the resulting raw data should not be evaluated directly,
due to many disturbances that can interfere with the analyte sig-
nals [12,20,21]. In principal, a recorded spectrum can be described
mathematically as shown in Eq. (1).
Equation 1. Vibrational Spectrum of Microplastics
Ið~nÞ ¼ Microplastics þ Matrix þ Baseline þ Noise
where Ið~nÞ is the measured signal intensity at each wavenumber ~n.
For microplastics identification, similarity of sample and consid-
ered references data are estimated and ranked. A numerical
concept for this estimation is called Hit Quality Index (HQI), which is
discussed in Section 4. However, at this point, it becomes clear that
the success of identifying microplastics is connected to the extent
of suppressing Matrix, Baseline and Noise effects. As far as
possible, optimising measurement conditions should be the first
option. In this context, purify samples physically, chemically or
biologically to get rid of matrices [5,22e25], adjust measurement
geometry to reduce baseline effects [26,27] and optimise spectral
resolution, number of co-added scans and aperture to increase
signal-to-noise ratio [28e30].
However, in many micro spectroscopic approaches like FTIRe or
Raman microscopy based imaging, spectra quality improvements
by adjusting measurement conditions are rather limited [31].
Moreover, different measurement modes such as attenuated total
reflection (ATR), diffuse reflectance (DRIFTS), transmission or
reflectance influence the measured spectra significantly, which
decreases universality of reference spectra libraries. Furthermore,
reflectance or scattering take place on the sample surfaces. Bio
films and other condensed impurities or contaminants may
particularly heavily overlay these spectra, which hamper successful
library searching.
In all these cases, spectra processing becomes very important.
All mentioned disturbances can also be suppressed numerically. In
real analytical systems, both approaches, optimising ‘hardware’ by
choosing best measurement parameters and ‘software’ by prepar-
ing measurement data with suitable methods are equally essential
for correct microplastics identification. At this point, it is important
to realise that data preprocessing can impair microplastics identi-
fication, if it is done improperly. Therefore, it is necessary to opti-
mize and validate all data preprocessing steps. A typical concept for
this is monitoring a defined target value, e.g., mean squared error
(MSE) of raw and processed data [32].
Creating a receiver operating characteristics (ROC) is also a
suitable method to compare different parameter setups and to
decide which of the investigated settings is most useful [33]. The
main idea of ROC is an analysis of true positive and false positive
rates of microplastics identification [11,33e36]. Concretely, it has to
be checked how many microplastic particles are identified correctly
in comparison to the amount of incorrectly identified microplastics,
which can be observed in Fig. 1 [11]. Especially for multivariate
algorithms that are often too complex to investigate single
parameter dependencies, ROC becomes a powerful optimization
tool [37e39].
Considering all these aspects, a workflow of identifying micro-
plastics can be outlined as follows:
a) Recording vibrational spectra of microplastics (optimised
measurement conditions).
b) Reduce noise intensities, if necessary.
c) Perform baseline correction, if necessary.
d) Normalise data.
e) Compare data to a suitable spectra library.
f) Identify microplastics (validate identification workflow).
3.1. How to increase signal-to-noise ratio?
A common way to increase signaletoenoise ratio is smoothing,
and different concepts are implemented in most spectrometer
softwares. In this context, moving average and SavitzkyeGolay
(1)
Fig. 1. Comparison of different library searching approaches for identifying 300
environmental microplastics. Manual inspection was used as reference method to
validate all algorithms. Considering these results, only the new developed mIDENT and
Pearson Correlation of derivative spectra are recommendable. (1) mIDENT, (2) Absolute
Distance, (3) Euclidean Distance, (4) Dot Product, (5) Pearson Correlation, (6) Euclidean
Distance 1st Derivative, (7) Pearson Correlation 1st Derivative, (8) Euclidean Distance
2nd Derivative, (9) Pearson Correlation 2nd Derivative [11].
 G. Renner et al. / Trends in Analytical Chemistry 111 (2019) 229e238
smoothing [40] are well established and both are useful techniques
to increase results certainty [11,41e43]. However, the used
smoothing span is an important parameter and should be defined
carefully. Thereby, too low values do not show the desired effect,
while too high values lead to ‘over smoothing’, which impairs the
identification results [20,44]. Fig. 2 underlines this phenomenon
and shows the similarities of polyamide microplastics to polyamide
and chitin references. In this case, one can observe an optimal
window for the span parameter for different sample spectra qual-
ities (s/n).
In order to define the optimal smoothing span, Vivo -Truyols et al.
[45] worked out an automatable concept for SavitzkyeGolay
smoothing. They hypothesised that a measured spectrum consists
of the true signal and noise ðIexp: ¼ bI þ εÞ. If the smoothing window
size is set correctly, the smoothed spectrum should be similar to the
true signal (Ismooth zbI). For optimisation, they monitored the re-
sidual noise of the measured and smoothed spectrum. For optimal
smoothing span, the noise ‘color’ of the residual noise should be
equal to the noise ‘color’ of the characteristic spectrometer noise,
which is given by Eq. (2).
Equation 2. Concept for Automatable SavitzkyeGolay Parameter
Optimisation
P 2 P
1 I exp:  I smooth ! 1 ðεi  εi1 Þ2
1 P 2 ¼1 P 2
2 ε 2 ε
|fflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflffl{zfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflffl} |fflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflffl{zfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflffl}
noise of the residues spectrometer noise
More sophisticated smoothing concepts like wavelet filters [46]
or multivariate denoising [47] should be suitable for microplastics
identification as well, but they do not play any significant role yet,
as their implementation is very complex and there is no significant
improvement compared to simpler smoothing algorithms like
moving average or SavitzkyeGolay smoothing.
Fig. 2. Influences of signal-to-noise ratio (s/n) on similarity (HQI) of polyamide
microplastics to polyamide reference (x axis) and chitin reference (y axis). Each square
represents an individual HQI pair of a polyamide sample to both references after
discrete smoothing. All basis spectra (aed) were smoothed using moving average with
different spans from 3 to 61 cm1. While HQI can be increased by optimal smoothing
(aec), result accuracy for almost noiseless spectra (d) is decreased by smoothing.
Yellow B indicate optimal smoothing conditions for each spectrum (a/27 cm1,
b/23 cm1, c/13 cm1, d/0 cm1).
231
Unfortunately, smoothing can change numerical properties of
signals, which also affects spectra evaluation. This phenomenon is
the more observable, the more the applied smoothing algorithm
considers neighboured data points. In this context, unweighted
approaches such as moving average lowers signal intensities,
broaden signal shapes and changes peak positions (in case of
asymmetric signal shapes). Therefore, these effects should be
looked at carefully and considered in order to choose a suitable and
practical smoothing algorithm.
3.2. How to compensate baseline effects and atmospheric
interferences?
Baseline effects can heavily impede the microplastics identifi-
cation process, which underlines the importance of baseline
correction before calculating HQI, as shown in Fig. 3. The origin of
noneconstant baselines is very complex and can be caused by
several sources, e.g., optical properties of samples are inhomoge-
neous, sample geometries are noneplanar or fluorescence phe-
nomena occur in Raman measurements. The latter heavily
decreases signaletoenoise ratio next to nonelinear baseline ef-
fects, which mostly limits success of numerical treatments.
Therefore, the best way to compensate these effects is optimising
hardware parameters of the measurement setup. However, there
(2) are also well established computational approaches for baseline
treatments.
SavitzkyeGolay differentiation offers an easy and effective so-
lution. This technique based on SavitzkyeGolay smoothing, but
instead substitute a raw data point with a centre point of poly-
nomial fit of a moving smoothing window, the nth derivative of this
Fig. 3. FTIR spectra of five polyamide microplastics before and after baseline correction
with airPLS, SavitzkyeGolay differentiation and numerical differentiation. It can be
obtained that without baseline correction, similarity of microplastics of the same
polymer type can differ due to variance of particle geometry. For calculating similarity
average, all spectra were correlated to each other by Pearson's r2. The numerical dif-
ferentiation cannot be recommended for microplastics identification, as signal-to-
noise ratio decreases dramatically.
232 G. Renner et al. / Trends in Analytical Chemistry 111 (2019) 229e238
polynomial is used. The main advantage of this technique is a good
automatability, which makes this method very useful for chemical
imaging approaches. The parameters that have to be adjusted for
SavitzkyeGolay differentiation are span and polynomial order. For
first and second order differentiation a span between 9 cm1 and
21 cm1 can be recommended, while a polynomial order of 2 or 3
seems to be suitable (based on our empirical observations).
Performing a conventional numerical differentiation for base-
line correction can not be recommended, as signaletoenoise ratio
decreases, which can be seen in Fig. 3 and indirectly in Fig. 2.
Next to its use for smoothing to reduce noise, moving average
can also be applied to define a baseline [48]. In contrast to the
former derivative approach presented, the moving average concept
leads to a normal baseline corrected spectrum. Hereby, a highly
over smoothed vibrational spectrum is estimated, which defines
the baseline. While sharp vibrational bands will disappear in the
moving average data, broad banded signals influence baseline
characteristics most. However, this can be a disadvantage, if broad
vibrational bands are focussed on, e.g. the characterisation of wood
polymer composites (WPC) or characterisation of bio films growing
on microplastics surface.
Another common baseline correction method is based on
empirical user input, whereby several data points are selected and
used as junctions for a polynomial baseline. The necessity of an
individual treatment makes this approach subjective, time
consuming and less automatable, especially if baselines vary a lot.
Therefore, this concept is very limited and cannot be generally
recommended. However, if the number of measurements is low,
such empirical approaches are also suitable but have to be per-
formed carefully.
A bit more complex but also very suitable is the concept of a
rolling-circle filter (RCF), which was presented by Brandt et al. [32].
In essence, a circle or an ellipsoid ‘rolls' over the measured vibra-
tional spectrum (transmission mode). Due to its diameter, which is
the parameter for this baseline correction algorithm, the circle
cannot follow sharp vibrational bands but ‘over rolls’ them. Broad
and slow spectral changes can be followed, which finally defines
the baseline.
Another mentionable baseline correction concept is asymmetric
least squares by Boelens et al. [49]. Hereby, the main idea is a
compromise of two different target values. On the one hand a
discrete measured data point Ii should be close to its belonging
point from baseline Bi. On the other hand, the intensity difference
from two neighboured baseline points DBi,i-1 lead to the following
lead to the following Eq. (3).
Equation 3. Asymmetric Least Squares Minimise the competing
sums:
X X 2
z> pi ðI i  Bi Þ2 þ l DBi;i1
where z is a pre-defined termination criterion and p is a weighting
vector that penalises negative differences of the Ii  Bi term, which
explains the asymmetric character of this concept. The weighting
parameter l specifies the baseline itself and has to be defined
manually by the operator. A modification of this approach is
adaptive iteratively reweighted penalized least squares (airPLS) by
Zhang et al. [50]. They focussed on re-weighting p within an
automatic loop process, which makes this baseline correction more
practicable.
RCF as well as ALS and airPLS are suitable but less often used,
because they are often not implemented in the spectrometer
software.
Next to signal disturbances caused by noneconstant baselines,
unwanted residues of atmospheric signals can appear in the
measured and background corrected spectra, which also in-
fluences the microplastics identification process and decreases
comparability. For compensation, automatic atmosphere sup-
pression is implemented in most spectrometer software applica-
tions. Commonly, these correction approaches use atmospheric
signals, e.g., carbon dioxide and water vapour from the measured
background or theoretical computed spectra, respectively, for an
empirical weighted subtraction from the measured sample spec-
trum. Thereby, discrete wavenumbers are monitored and
weighting factors are set automatically to minimise signal residues
[51,52].
3.3. Spectra normalisation
In vibrational spectroscopy of microplastics, signal intensity
levels vary a lot, as the cross section between light and sample
depends on geometrical properties [20,53]. Hence, an intensity
normalisation before analysing similarity of sample and reference
seems to be plausible, and therefore, this is also a well established
step before spectra library searching. Typically, the operator is not
involved into this data transformation, if microplastics identifi-
cation is performed by a spectrometer software. However, for any
manual identifications with external software, it could be useful
to know, how the normalisation algorithm works to avoid iden-
tification errors. The commonly used normalisation technique
changes the recorded intensity data (I) from a   ∞; þ∞½ to a
½0; 1 range. This means the smallest intensity becomes zero,
while the largest intensity becomes one, which can be achieved
by Eq. (4).
Equation 4. Data Normalisation
I  min I
I normalised ¼ (4)
max I  min I
However, this normalisation algorithm is only suitable, if the
smallest and largest intensities can be detected properly and are
characteristic data points for the examined microplastics. Over-
saturated signals, e.g. analysing frequencies that will not pass
spectrometer optics or completely be absorbed by the sample can
disturb data normalisation. In some cases, it can be suitable to cut
these frequencies from the data set.
Alternatively, instead using minimum and maximum of the
absorbance vector, the maximum intensity of a defined vibrational
band and the mean intensity of the baseline signal can be used for
normalisation [30]. However, this method needs an almost indi-
vidual treatment, which limits its application in high sample
throughput tasks.
A more objective and automatable approach is standard normal
(3)
variate (SNV) transformation [54]. This technique uses mean (I) and
standard deviation (sI ) of a vibrational spectrum for normalisation,
which can be seen in Eq. (5).
Equation 5. Standard Normal Variate Transformation
II
I normalised ¼ (5)
sI
In contrast to the minemax normalisation (Eq. (4)) SNV uses not
two reference points from the measured spectrum (minimum in-
tensity and maximum intensity) but SNV uses all data points to
calculate the reference values (mean and standard deviation sI ). On
the one hand, scattering variances are suppressed much better by
SNV, but on the other hand, the presence of matrix signals can
 G. Renner et al. / Trends in Analytical Chemistry 111 (2019) 229e238
influence mean (I) as well as standard deviation (sI ). Therefore,
SNV could lead to low HQIs.
3.4. Spectral range
Within a vibrational spectrum of microplastics, not every
spectral region contains relevant information. Therefore, it can be
helpful to reduce the recorded data to a more compact but highly
characteristic sub-spectrum. As it is shown in Fig. 4, this can in-
crease the selectivity of library searching [11,13,41,55]. Especially
for multivariate classification, selection of the considered spectral
range is an important factor [56].
3.5. Spectrometer software versus manual evaluation
Up to this point it could already be figured out that microplastics
identification is not just a ‘one-click’ operation, but means a com-
plex process that has to be performed carefully. In this context, the
spectrometer software provides more and more rather useful tools
to walk through this procedure, and the major part of the micro-
plastics research community seems to accept these with thanks.
This can be assumed, since there are hardly reports identifying
microplastics based on non-implemented algorithms [11]. How-
ever, it is worth to look beyond the spectrometer software, as
microplastics analysis is not routine but characterised by a high
variability, which often means that each measured particle has to
be treated individually. A manual evaluation concept has the
advantage of a much higher flexibility as the number of possible
algorithms is almost unlimited.
Handling a huge number of spectra while performing accurate
individual data processing is challenging and research studies that
deal with microplastics imaging have already realised that spec-
trometer software is limited for this problem [17,28,57,58]. How-
ever, by creating own evaluation concepts, they succeeded very
well in identifying microplastics. In this context, Primpke et al.
developed a super fast and accurate identification method to
Fig. 4. Absorbance variance spectrum of a spectra library for microplastics identifi-
cation (containing 22 common polymers and matrix components). The Spectrum can
be divided into three regions: 4000e2750, 2750e1850 and 1850e700 cm1. It can be
observed that the variance (intensity) and specificity (number of individual signals) in
the fingerprint region (1850e700 cm1) are very high. Therefore, this region seems to
be suitable for microplastics identification, whereas the middle region
(2750e1850 cm1) cannot be recommended due to its low variance and specificity.
233
analyse microplastics that are enriched on alumina filters [17]. The
main difficulty was the huge amount of spectra ( > 1 mio). In the
end, they came up with a multi step identification process that
combines different HQI algorithms to reduce identification errors
below a 3% level.
Furthermore, Renner et al. developed a microplastics identifi-
cation method that focussed on maximising robustness to identify
even heavily weathered microplastics in an accurate way [11]. The
main idea was to use the strengths of peak search instead of full
spectrum search, and they succeeded in creating a fully automated
and highly robust vibrational band detection algorithm for FTIR
spectra. Using this algorithm, the identification accuracy could be
improved from 70% to 96% for diversely weathered microplastics.
4. From spectrum to hit quality index: how does library
searching work?
The most commonly used technique to evaluate vibrational
spectra of microplastics is library searching [14]. This bases on the
idea that each substance is connected to a highly characteristic
vibrational spectrum, which is called “finger print” of a substance.
To that end, a spectrum of an unknown sample is compared to a
reference spectra library, and if the similarity reaches a certain
level, the unknown sample can be assigned [9,13,59e61]. However,
even if this sounds very familiar to most researchers that use
vibrational spectroscopy to identify microplastics, there are many
parameters like hit quality index threshold, type of algorithm or
baseline correction that influence this procedure dramatically, and
there still exists no standardised workflow for spectra evaluation
[14,16].
4.1. Reference spectra library
Identification results of microplastics analysis are strongly
connected to the reference spectra library which was used. Most
commercially available databases (from spectrometer manufac-
turers or third parties) are divided into several categories including
polymer references. Next to polymers, it is highly recommended to
consider typical matrix components like wood, wool, cellulose,
proteins, seaweeds, fish parts, human hair or skin particles and add
their spectra to the search field in order to avoid misidentifications
[7,9,17,62]. As the number/diversity of relevant microplastics that
are found in environmental samples only consists of a handful
references, many research groups create their own laboratory in-
ternal database to ensure that measurement parameters of sample
and references are equivalently defined [11,13,17]. This approach is
highly recommended, due to the fact that measurement parame-
ters can influence library searching, which will be discussed later. In
the following, typical considered references of the main groups of
polymers for microplastics identification are listed.
Furthermore, most commercially available spectra libraries are
based on non-degraded pure and often synthetic reference mate-
rials, while most samples are degraded and may contain residues of
diverse contaminations, e.g., sand, biofilms etc. Additionally, addi-
tives or fillers can lead to additional signals, and therefore, in-
fluences the measured spectra significantly, which is confusing if
the corresponding database documentation does not contain any
information about them.
 Polymers:Polyethylene, Polypropylene, Polystyrene, Polyamide,
Polyvinyl chloride, Polylactide, Polyethylene terephthalate,
Polycarbonate, Polyurethane, Poly(methyl methacrylate), Poly-
butadiene, Polyoxymethylene
 Matrices:Cellulose, Chitin, Wood, Wool, Seaweeds, Human hair,
Human skin particles, Sand, Carbon black
234 G. Renner et al. / Trends in Analytical Chemistry 111 (2019) 229e238
Of course, these lists are not exhaustive and may need to be
extended.
4.2. Hit quality index
Identifying a substance with library searching is based on sim-
ilarity analysis, and at the end of discrete calculations, which will be
discussed later, every compared entry of the reference spectra li-
brary is characterised by an individual numeric value called “hit
quality index” (HQI). This parameter describes similarity of two
spectra (reference and sample) in a normalised range between 0 ‰
and 1000‰ (0e1 or 0 %e100%, respectively). In this context, there
are several research groups and guidelines that recommend at least
an HQI of 700‰ for microplastics identification [9,63e66]. Never-
theless, the HQI difference (DHQI) to other possible, also high rated
candidates is a second very important value that is often overseen
and which describes the selectivity of the assignment. In this
context, there exist two suitable approaches to define a threshold
for DHQI e constant value of DHQI  100‰ [20] or a relative value
of DHQI  HQI  0.1 [20]. Unfortunately, both concepts will prohibit
to distinguish between similar polymers or sub classes of polymers
like low and high density polyethylene (LDPE & HDPE) or poly-
amide 6 and polyamide 6.6 (PA 6 & PA 6.6). Even a significant dif-
ference between some polyamides and naturally occurring chitin or
human skin particles is often not observable [67]. However, it is
important to underline, that the identification is only assisted by
HQI [20] and if at least one criterion (HQI or DHQI) is not fulfilled,
the identification should be re-evaluated by manual inspection
[13].
The HQI itself is heavily effected by many parameters like signal-
to-noise ratio or the presence of baselines that are significantly
different from zero, which can be observed in Fig. 5. Therefore, it is
highly recommended to monitor spectra quality and improve
Fig. 5. Suitable spectra preprocessing workflow for microplastics identification based
exemplarily on a polypropylene microplastic particle. Denoising was performed by
SavitzkyeGolay smoothing with a span ¼ 11 cm1 . Baseline effects were compensated
by airPLS with l ¼ 6  109 and p ¼ 0:5. The fingerprint region was defined between
700 and 1850 cm1. The Hit quality index (HQI) was calculated by Pearson's r2.
measurements physically in the first place and numerically by data
preprocessing in the second place. For the latter, useful tools and
techniques were presented above in Section 3.
4.3. Common HQI algorithms
The HQI describes similarity of a measured sample to a reference
spectrum, and therefore, it is the main decision criterion for
microplastics identification. However, there are many different
concepts about describing data similarity, and considering this, it is
not surprising that there are several algorithms that can calculate
the HQI and none of them leads to identical results [11,17,68e70].
Additionally, every algorithm has its pros and cons, which means
that there will be no gold standard approach that can be used for
any kind of data. In the following, the strength and weaknesses of
the common algorithms will be discussed.
4.3.1. Euclidean Distance
In essence, Euclidean distance calculates the squared absorbance
differences for every data point from normalised sample (s) and
reference (r) spectrum, which can be obtained from Eq. (6).
Equation 6. Euclidean Distance
" #1
jjs  rjj2
HQI ¼ 1 þ (6)
jjrjj2
At this point, it has to be considered that there is no general
equation, which is used for calculation. Instead, many spectrometer
manufacturers use individual variants of the shown Eq. (6), but
fortunately, they are mostly documented quite well within the user
manuals [51]. Although Euclidean distance is a standard library
searching algorithm, it cannot be recommended for microplastics
identification because it is very susceptible for baseline effects and
matrix interferences like the presence of biofilms on the sample
surface [11,51].
4.3.2. Pearson's r2
One of the most used algorithms to calculate HQIs for spectra
library searching is Pearson's r2. In contrast to Minkowski distance
approaches, the correlation of two datasets is a relative method (Eq.
(7)).
Equation 7. Pearson's r2
2 32
 2 P
covðs; rÞ 6 ðs  sÞðr  rÞ 7
HQI ¼ ¼ 4qffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi qffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi5 (7)
ss ,sr P 2 P 2
ðs  sÞ , ðr  rÞ
where covðs; rÞ is the covariance of sample and reference and de-
scribes those signals that both have in common. ss ,sr is the
product of sample and reference standard deviations that describe
signals from both spectra individually. In contrast to Euclidean
distance, Pearson's r2 is a weighted approach and does not treat
every data point in the same way, but prefer extrema (mostly
vibrational bands). As it can be obtained from Fig. 3, baseline effects
also decrease data similarity but less strong than Euclidean distance
and especially periodically non-linear baselines just disturb
slightly. The latter is an interesting phenomenon, as cosine
modulated baselines can appear in transmission measurements of
microplastics, and Pearson's r2 seems to be robust against this
disturbance factor. Nevertheless, accurate baseline correction is a
very important step before performing library searching, especially
in presence of linear baselines which can appear during ATR ex-
periments [20,53].
 G. Renner et al. / Trends in Analytical Chemistry 111 (2019) 229e238 235

4.3.3. Derivative approaches 4.3.4. Peak search

A suitable way to identify microplastics is to combine similarity In former days, computing power was strongly limited, and HQI
calculation and baseline correction to decrease sources of errors calculations based on point-by-point methods like Euclidean dis-
during library searching. With respect to a high sample tance or Pearson's r2 were not practical at all due to too long
throughput, automated concepts become more and more impor- computing times. To that end, the measured spectrum consisting of
tant, and one easy but effective approach is library searching based many data points was reduced to a much more compact list of
on derivative spectra. Concretely, the recorded spectra are differ- significant peak positions and their intensities. This minimises the
entiated numerically and are compared to differentiated reference calculation times, as this approach processes significantly fewer
spectra with already established comparison algorithms like data points.
Euclidean distance or Pearson's r2. This mathematical trans- In essence, peak tables of sample and reference are compared
formation has two interesting effects that improve similarity for HQI calculation as it follows in Eq. (8).
analysis. On the one hand, broad banded interferences are Equation 8. Peak Search
underestimated, which can be understood as some kind of base-
line correction [71,72]. A side-effect or consequence of this phe- No: of common Peaks in Sample & Reference
HQI ¼ (8)
nomenon is also a discrimination of broad vibrational bands, No: of Peaks in Reference
which can improve selectivity of library searching for micro-
Optional, peak intensities can be used for weighting.
plastics that mostly show sharp signals, whereas attached bio-
While in mass spectrometry, peak search is one of the most used
films or other bio-organic matter show many broad vibrational
evaluation concepts, this technique is seldom used except for
bands due to the great variety of the monomer sequences, which
chemical imaging, where often only a few but characteristic
can be observed in Fig. 3.
vibrational bands are used for identification [17,28]. However, it
On the other hand, the mean spectral absorbance, which is an
could be shown that microplastics identification based on peak lists
important parameter for Pearson's r2 calculation becomes more
P from vibrational spectra can improve result accuracy a lot [11]. The
similar to the baseline of the spectrum. This results in ðs  sÞðr 
P 2 P 2 success of this approach depends admittedly on a proper peak
rÞ, ðs  sÞ and ðr  rÞ becoming almost zero for baseline
characterisation, which regretfully increases the complexity of
data points, which means, vibrational bands are even more
peak searching approaches.
preferred for HQI calculation, if the data were differentiated before.
However, numerical differentiation is also connected to a
decreasing of signal-to-noise ratio, which disturbs similarity as
5. Identification concepts based on multivariate classification
shown in Fig. 3. To that end, it is highly recommended to perform
not simple differentiation but SavitzkyeGolay differentiation,
Multivariate classification models have become very popular in
which includes a smoothing to increase signal-to-noise ratio [40].
many research fields, e.g. authenticity testing or forensic analysis
Fortunately, this technique is implemented in many spectrometer
[73e76], and therefore, it is plausible to look if these concepts are
software applications.
also suitable for microplastics identification. In parallel to

Table 1
Strengths and weaknesses of common and suitable data preprocessing methods for microplastics identification based on vibrational spectroscopy.

Task Method Strengths Weaknesses Applied in

Denoising moving average fast & easy decreases sharp signals [77,78]
SavitzkyeGolay smoothing fast & easy constant smoothing [79,80]
wavelet filter conserve signal shapes complex
multivariate denoising conserve signal shapes complex
Baseline Correction SavitzkyeGolay differentiation automatable broad signals disappear [11,42,43,81]
rolling-circle filter conserve most signals hardly automatable
asymmetric least squares conserve most signals hardly automatable [80,82]
adaptive iteratively conserve most signals hardly automatable
reweighted penalized
least squares
Spectral Region full spectrum good to distinguish less good for microplastics [83e85]
(400e4000 cm1) between polyamide & classification
bio(organic) mass
fingerprint ( < 1850 cm1) highly characteristic data [11,43,86]
Normalisation minemax normalisation automatable error-prone [17,43]
with noisy data
reference peak good for quantifications less good for microplastics [82,84,87,88]
classification
standard normal good to normalise less good if [79,81]
variate transformation repeated measurements aging effects
vary highly
Library Search Euclidean distance baselines and matrix interferences [59,82]
falsify results
2
Pearson's r baselines and matrix interferences [17,80]
falsify results
Euclidean distance matrix interferences [42]
1st derivative falsify results
Pearson's r2 1st derivative suitable for most matrix interferences [17,42]
microplastics falsify results
Peak Search very robust hardly automatable [11,85]
multivariate classification automatable complex, “black box” [43,81,85,89]
236 G. Renner et al. / Trends in Analytical Chemistry 111 (2019) 229e238
conventional library searching, multivariate classifications charac-
terise similarities between data sets, or samples and references,
respectively. In general, those approaches can investigate a dataset
that contains diverse reference microplastics spectra and extract so
called ‘classifiers’. These describe the relevance of each data point
in a spectrum for a discrete classification. Not only vibrational
bands are taken into account but relevant signals, which can be the
same but in real systems this is less often the case. Therefore, once
again, data preprocessing also influence multivariate classification
models and has to be performed carefully including a detailed
parameter validation.
As one of the few, Paul et al. applied multivariate classification to
identify soil samples that are contaminated with microplastics [89].
They successfully pointed out how powerful these techniques can
be, as they detected 1 %wt. microplastics within an integral mea-
surement setup for near infrared spectroscopy.
Halstead et al. used principal component analysis (PCA) to
identify ingested microplastics and natural fibres in fish [56].
Concretely, they added vibrational spectra from samples and ref-
erences to a PCA and performed a cluster analysis for identification.
In this study SavitzkyeGolay second order derivative baseline
correction was applied, and the spectral range was reduced to the
finger print region.
In summary, Paul et al., as well as, Halstead et al. could suc-
cessfully prove that multivariate classifications of environmental
microplastics are fit for purpose.
6. Suitable strategies to identify microplastics
As already mentioned before, identification accuracy of micro-
plastics depends on data quality, and in the first place, optimising
measurement conditions should be the option of choice. Never-
theless, a properly performed spectra processing is important, as
not all disturbances can be avoided by adjusting measurement
parameters. To that end, Fig. 5 and Table 1 contain a practical
workflow and a method overview for microplastics identification.
Before finalising the microplastics identification results, it is
highly recommended to check for plausibility. Hereby, two aspects
play a significant role. On the one hand, the identification result
itself has to be looked at critically, meaning the specific polymer
that was found. Most of the identified microplastics from the
environment was characterised as fragments from packaging ma-
terials or synthetic textiles, e.g., polyethylene, polypropylene,
polystyrene, polyamide, polyvinyl chloride, polylactide, poly-
ethylene terephthalate. Findings beyond this list should in partic-
ular be reviewed carefully and compared to other microplastics
studies. On the other hand, sampling location has to be considered,
due to different mobilities of diverse microplastics in different
environmental compartments. Floating microplastics, e.g., poly-
ethylene or polypropylene are commonly found on water surfaces
or beaches, while polymers with higher densities, such as polyvinyl
chloride or polyamide are enriched in the sediments. Deviations
from this concept should be checked critically. However, it has to be
underlined that results that do not fit with these aspects are not
automatically wrong and results that do fit are not automatically
correct.
7. Conclusion
Identification of microplastics is a key step in many studies that
deal with microplastics in the environment. In contrast to other
methodological aspects, data preprocessing is described in much
less detail, which often impedes transparency of the results.
However, this review article could demonstrate that microplastics
identification depend strongly on data preprocessing. Fortunately,
there already exist suitable techniques in literature that ensure
highly accurate results. To that end, it is worth it to apply these
methods rationally and keep in mind their specific strengths and
weaknesses.
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