Review

Poly- and Perfluoroalkyl Substance (PFAS) Analysis in

Environmental Matrices: An Overview of the Extraction and
Chromatographic Detection Methods
Alessia Iannone 1 , Fabiana Carriera 1 , Cristina Di Fiore 1 and Pasquale Avino 1,2, *

1 Department of Agriculture, Environmental and Food Sciences, University of Molise, Via De Sanctis,
86100 Campobasso, Italy; [email protected] (A.I.); [email protected] (F.C.);
[email protected] (C.D.F.)
2 Institute of Atmospheric Pollution Research, Division of Rome, c/o Ministry of Environment and Energy
Security, Via Cristoforo Colombo 44, 00147 Rome, Italy
* Correspondence: [email protected]

Abstract: Per- and polyfluoroalkyl substances (PFASs) are carbon–fluorine compounds with widespread
industrial and domestic use, posing potential toxicological risks to humans and ecosystems. Several
analytical methods have been developed to assess the occurrence of PFASs in the environment, but a
standardized method, applicable to all matrices, is still lacking. This paper reviews the extraction and
chromatographic detection methods for PFAS assessment in environmental samples, considering
parameters such as the LOD, LOQ, and recoveries. Solid phase extraction (SPE) is commonly
used, showing high recovery rates for water, soil, and sediment samples using HBL and WAX
polymeric sorbents (85–100% and 93–111.5%, respectively). LC-MS has demonstrated low LODs
and LOQs in seawater (0.01–0.08 ng L−1 ; 0.03–0.24 ng L−1 ), marine sediment (0.002–0.018 ng g−1 ;
0.004–0.054 ng g−1 ), and dust (0.08–0.68 pg g−1 ; 0.26–2.25 pg g−1 ), indicating its sensitivity when
detecting trace PFAS levels. Evaluating PFASs is crucial for the development of future removal
strategies and risk assessments. Potential solutions including the use of PFAS substitutes and
innovative adsorption techniques for their adsorption could present promise in reducing their
Citation: Iannone, A.; Carriera, F.; Di environmental presence.
Fiore, C.; Avino, P. Poly- and
Perfluoroalkyl Substance (PFAS) Keywords: PFAS; environment; exposure; toxicological hazards; extraction approaches; detection
Analysis in Environmental Matrices: methods
An Overview of the Extraction and
Chromatographic Detection Methods.
Analytica 2024, 5, 187–202. https://
doi.org/10.3390/analytica5020012 1. Introduction
Academic Editor: Victoria Per- and polyfluoroalkyl substances (PFASs) are emerging contaminants, largely
Samanidou present throughout the environment. Data suggested by the Organisation for Economic
Cooperation and Development (OECD, 2018) report 4700 PFASs being currently available
Received: 10 March 2024 on the market [1]. Their chemical inertness, thermal stability, amphiphilicity linked to their
Revised: 11 April 2024
highly polarity, and strong carbon–fluorine (C-F) bonds allow them to be used in manyfold
Accepted: 15 April 2024
industrial applications (households and industrial products, retardant flame, waterproof
Published: 19 April 2024
clothing, food packaging, aqueous film-forming foams (AFFF), functional fluids for ma-
chinery, and anti-corrosion agents) and their following release into the environment [2–5].
The emission of PFASs can occur through direct or indirect routes. Industrial dis-
Copyright: © 2024 by the authors.
charges, landfills, agricultural runoff, firefighting training sites, and wastewater treatment
Licensee MDPI, Basel, Switzerland. plants (WWTPs), as well as atmospheric deposition, are direct sources which promote the
This article is an open access article distribution of PFASs to remote areas [4,6]. On the other hand, they can be indirectly re-
distributed under the terms and leased through the degradation of neutral and volatile PFAS precursors [7]. The persistence
conditions of the Creative Commons and bioaccumulation of PFASs in the environment have become a serious concern for the
Attribution (CC BY) license (https:// potential toxic effects on human and wildlife [8].
creativecommons.org/licenses/by/ Studies focused on the toxicological profiles of perfluorooctanoic acid (PFOA), perflu-
4.0/). orooctanesulfonic acid (PFOS), and perfluorohexanesulfonic acid (PFHxS) from the Agency

Analytica 2024, 5, 187–202. https://doi.org/10.3390/analytica5020012 https://www.mdpi.com/journal/analytica

Analytica 2024, 5 188

for Toxic Substances and Disease Registry (ATSDR) have suggested ranges of human
half-lives of 2.1–10.1 years, 3.1–27 years, and 4.7–35 years, respectively [9]. The same com-
pounds have also been observed to have half-lives in ranging from 1.2 to 3.3 days for the
northern leopard frogs (Rana pipiens), a bioindicator frequently used for ecosystem health
monitoring [10]. Generally, PFASs are present in varying forms of anions, zwitterions,
cations, or neutral compounds within the environment [11].
The transport of PFASs from the environment to the human body occurs though
several environmental media, such as water, sediments, air, and dust [12]. Therefore, they
can enter the human body through digestive, respiratory, and cutaneous pathways [13].
Liver and blood are the main accumulative pathways of PFASs [14–16]. Levels of PFHxS,
PFOA, and perfluorononanoate (PFNA) have been observed in liver and blood tissues in the
mean concentrations of 0.153, 0.301, and 0.244 ng g−1 , with the highest mean concentration
of PFOS being in liver (6.277 ng g−1 ) [17,18]. Moreover, a higher concentration of PFASs in
tumor liver tissue (mean 64.3, range 5.70–303 ng g−1 ) has been found when compared to
non-tumor liver tissues, confirming the major exposure of patients with cancer to oxidative
stresses [19]. Nonetheless, the investigations about the combined effects and toxicological
interactions have also confirmed the increasing cytotoxicity of PFASs to the human cell
liver line (HepG2) with increasing carbon chain lengths [20,21]. The fate of PFASs can reach
long distant organs, such as lungs and their bronchial epithelial cells [22]. The pulmonary
epithelium and the layer of lung surfactants (LS) covering the cells are the primary portal
of inhaled pollutants, creating a physical barrier of entry between the outside environment
and blood circulation [23]. In the study of Pye et al. [24], it was shown that environmental
protein contained in dust particles inhaled can act as a transport carrier of PFASs into the
respiratory system, modifying the lung surfactant function and inducing pro-inflammatory
and allergic responses [25–27].
Considering the fate of PFASs in the environment and their potential negative effects,
it is indispensable to develop an accurate and sensitive analytical method with which to
control them. Among the extraction procedure of PFASs, solid phase extraction (SPE) is
the main protocol used for the extraction, purification, and pre-concentration of PFASs
in environmental media [7]. Moreover, it is crucial to understand the behavior of PFAS–
sorbent interactions at different conditions in order to optimize the extraction procedure [28].
For example, the hydrophilic–lipophilic balanced (HBL) sorbent exhibits a low affinity
for short-chain PFASs (perfluorosulfonic acid (PFSA)) and perfluoroalkyl carboxylic acid
(PFCA)), and a greater affinity for the zwitterionic PFASs. Otherwise, the weak anion
exchanger (WAX) polymeric cartridges (e.g., Oasis WAX and Strata-X) exhibits better
recovery rates for short chain homologues; the presence of tertiary amine functional groups
enhances the retention of anionic PFASs via electrostatic interactions [29,30]. For PFAS
monitoring, liquid chromatography–mass spectrometry (LC-MS/MS) is the main analytical
procedure applied. Specifically, high-performance liquid chromatography (HPLC) and
ultra-high pressure liquid chromatography (UHPLC), coupled with triple quadrupole mass
spectrometers (QqQ) or triple quadrupole tandem mass spectrometry (MS/MS), show
a higher sensitivity for PFAS quantification [3,31]. However, gas chromatography (GC)
coupled with a flame ionization detector (GC-FID), an electron capture detector (GC-ECD),
and a mass spectrometer (GC-MS) has been employed as well [32]. The present review
aims to summarize the current extraction and analytical approaches for the determination
of PFASs in environmental matrices and details like water, soil, air, and dust in order to
underline the absence of a standardized methodology for such analytes. Furthermore, a
critical analysis of the current knowledge about both analytical methods, as well as the
toxicological aspects, has been provided. This review also considered the regulation of
force thus far in order to analyze its effectiveness on the protection of both human and
wildlife health statuses.
Analytica 2024, 5 189

2. Literature Research
The literature research was conducted, searching the most recent studies (from 2015
to 2023) on Scopus and Google Scholar databases. To this purpose, the keywords selected
were “per- and polyfluoroalkyl substances”, “environment”, “toxicology”, “extraction
techniques”, and “analytical methods of detection”. A total of 1188 studies emerged. The
studies were first analyzed by reading their titles and abstracts, and 556 studies were
excluded. Afterwards, only studies which met the aim of the present review (no = 632)
were screened by reading the full text. Hence, 80 out of the 632 were selected for the review.

3. Toxicology and Risk Assessment of PFAS

Toxicity studies of PFASs have shown the detrimental effects on humans and on the
environment [8,33,34]. Due to the amphiphilic properties, these compounds can attach
to proteins and be transported in the blood, kidney, and liver, causing multiple adverse
effects, such as perturbations in thyroid, liver, and kidney disease, reproductive toxicity,
high cholesterol, and immunosuppression [35,36]. A work by Poothong et al. (2020) [37]
showed an average concentration of 0.80 ng mL−1 and 0.90 ng mL−1 of PFASs in the blood
serum of people <41 and >41 years old, respectively. Furthermore, a positive correlation
was demonstrated between the PFAS concentration in blood serum and those found in
indoor dust. The highest levels of PFASs were detected in the human whole blood, serum,
and plasma of samples collected from people aged between 20 and 66 years old. The
levels were higher in both the serum and plasma (10 ng mL−1 ) as compared to the whole
blood (6.3 ng mL−1 ) [38]. The entering of PFASs into the blood circulation could led
to the transport of PFASs into other organs and their consequent accumulation within
them. A concentration ranging between 0.04 and 0.19 ng g−1 was found in human kidney
tissue, likely due to environmental exposure [39]. The potential toxicity of PFASs (PFOA
and PFOS) is likely correlated to the extended half-life that they could have due to the
re-adsorption in the renal tubule; specifically, in the proximal convoluted tubule and the
distal convoluted tubule [40]. The distribution and bioaccumulation of PFASs in different
tissues can be affected by their physicochemical properties. Experiments conducted on rat
models conducted by Cao et al. (2022) [41] suggested the key role of binding proteins, like
ASBT and NTCP, involved in the enterohepatic circulation in the reuptake of PFASs from
the intestinal lumen and the accumulation in blood and the liver. Concentrations of PFASs
in the liver are rarely reported. A work by Baumert et al. (2023) [42] showed for the first
time the concentration of PFASs (PFOA and PFOS) measured simultaneously in the plasma
and livers of 64 individual Teen-LABS participants. The highest concentration of PFOA
and PFOS were detected in plasma (1–10 ng mL−1 ). Likewise, a high PFOS concentration
(1–10 ng g−1 ) was observed in the liver. The accumulation of PFOS and PFOA in the liver
and plasma tissues, as studied using the intratissue Mass Balance Model (MBM), was
explained by the role of membrane lipids as carriers of hydrophobic, long-chain PFASs
in the liver and by the albumin-binding hydrophobic PFASs, which are strongly present
in plasma. Moreover, the strong correlation between plasma and liver concentrations
(r2 = 0.991), found by means of the multilinear regression (MLR) model, suggested the
ability to predict the concentration of PFASs in liver from those found in plasma. Basing on
the epidemiological evidence of PFAS toxicity, deleterious impacts on liver metabolome,
specifically bile acid metabolism, have been shown following environmental exposure
to PFASs [43]. It was observed that the increase in circulating cholesterol levels, as well
as the alteration of bile acid and fatty acid metabolites, could affect the enterohepatic
circulation and energy metabolism [44], and consequently dysregulate the lipid metabolism
and sterol homeostasis, leading to the development of non-alcoholic fatty liver disease
(NAFLD) [45] and its more severe form, non-alcoholic steatohepatitis (NASH). The first
potential exposure to PFASs begins in utero, specifically for people who transfer the fetus
through the placental passage [46,47]. Trophoblast cells form the outermost layer of the
blood—-placental barrier, which vehicles the PFAS transfer. In particular, the cell line JEG-3
Analytica 2024, 5 190

is relevant for the presence of multinucleated trophoblasts, which simplify the exchange of
nutrients, waste, and gases between maternal and fetus compartments in the placenta [48].
However, the concentration and ability of PFASs to cross the placental barrier depends
both on passive and active diffusion and on their binding affinity to proteins in human
serum. A work conducted by Lu et al. (2021) [49] reported the highest level of PFOSs and
PFOAs (mean: 0.457 and 0.242 ng g−1 ww, respectively) in placental human samples. They
observed that the binding affinities of PFASs to human serum albumin (HAS) increased
with the chain length. However, due to the difficult transfer of PFAS–protein complexes
through the placental barrier, the concentration of free PFASs in the blood is an important
factor for the placental passive diffusion [50]. As for the active transport process, PFASs act
as a vehicle for the organic anion transporters (OAT) placed on the placenta membrane.
Although the binding behavior of PFASs as placental transporters is still unknown, an
increase in binding affinities to OAT4 was observed with the chain length of PFASs [49].
Prenatal exposure to PFASs may be harmful for the child’s health. Studies have suggested
negative outcomes regarding cognitive and neurobehavioral development [51,52], the risk
of asthma [53], allergies, and infectious diseases [54]. However, although the toxicity
of PFASs at high doses of exposure (acute toxicity) is well documented, there are few
studies focusing on chronic effects on health. A work from Kristensen et al. (2013) [55]
investigated the possible long-term effects of in utero exposure to PFOAs and PFOSs
on female reproductive functions. A significant association between prenatal exposure to
PFOAs and menarches occurring at a later age was observed, with a difference of 5.3 months
between the lowest and highest tertiles of the PFOA concentration. This delay could have
psychological consequences on human health, but no significant association between PFAS
exposure and the menstrual cycle length, as well as the reproductive hormone levels or
the number of antral follicles in young adulthood, was observed. Moreover, the only
PFOA association with the age at the time of the menarche occurring could be explained
via the easily transfer of PFOAs through the placenta barrier. Epidemiological studies
on mice exposed to PFASs for 12 weeks showed elevated serum cholesterol levels, sterol
metabolites, and bile acids, with some sexual dimorphic effects [44]. Moreover, a recent
work by Maxwell et al. (2024) [55] on male mice exposed to PFASs for 18 weeks showed
the influence on sperm methylation, which interfered with the epigenetic regulation of the
gene expression of offspring liver and fat. This result could be significant in predicting the
long-term offspring gene expression and the link between environmental PFAS exposure
and the health of future generations. Dust particles inhaled, as well as dermal contact
with household cleaning products have also been indicated as a potential source of PFAS
exposure [56].
Emerging evidence has suggested associations between air pollutants, like PFASs, and
lung damage [23,57,58]. It was observed that fine particulate matter with an aerodynamic
diameter of less than 2.5 µm (PM 2.5) can readily enter the lower airways, causing respira-
tory and cardiovascular diseases, including asthma, chronic obstructive pulmonary disease
(COPD), pulmonary fibrosis, cancer, type-2 diabetes, neurodegenerative diseases, and even
obesity [59]. However, these pollutants are also spread in indoor environments (home and
public spaces) and inhaled via air or dust. Animal studies conducted by Gustafsson et al.
(2022) [60] revealed that the concentration of PFOAs on lung tissue was almost 2.5 times
higher when compared to the liver and kidney following the inhalation of and oral exposure
to PFOA-coated house dust. In addition, data obtained via US EPA 2017 suggested dust
ingestion rates of 50–100 mg day−1 (toddlers) and 20–60 mg day−1 (adult), with daily PFOA-
equivalent intakes exceeding the daily threshold fixed by the European Food Safety Authority
(EFSA) (1.3–1.5 ng kg−1 body weight (bw) d vs. 0.63 ng kg−1 bw d−1 ) [61].

4. Regulation
Ecological and potential human health effects of PFASs have caused regulations and
restrictions regarding their use to be implemented. In 2018, the “Global PFC Group”,
led by the Organisation for Economic Co-Operation and Development (OECD) and the
Analytica 2024, 5 191

United Nations Environment Programme (UNEP), published a list of over 4700 PFASs
that contain–Cn F2n – (n ≥ 3) or Cn F2n OCm F2m – (n and m ≥ 1) moieties [1]. Among these,
PFOSs and PFOAs are the most known and frequently used PFAS, listed as persistent
organic pollutants (POPs) under the Stockholm Convention, together with their related
salts [62]. In January 2023, the National Authorities of Germany, Denmark, the Netherlands,
Norway, and Sweden submitted a proposal of the reduced application of PFASs to the
European Chemicals Agency (ECHA), with the aim to reduce PFAS emissions into the
environment, and to make products and processes safer for people (ECHA/NR/23/01) [63].
This objective is in line with the ambitions of the EU Chemicals Strategy and the Zero
Pollution Action Plan, which assumes the reduction levels of pollutants in air, water, and
soil, and creates a toxic-free environment (European Commission, 2021) [64]. Despite the
emergency of PFASs being a common focus of investigation for scientific and political
communities, their toxicity, their potential risks on health, as well the exposure levels of
humans are still uncertain. The lack of evidence between public health adverse impacts and
their behavior has led governments to set different guidelines for some PFASs. Moreover,
the lack of standardized analytical methods and stable isotope-labeled internal standards,
the interdependence of programs, legislative support, and funding represent important
challenges, which challenge setting uniform PFAS values among different countries. There-
fore, with the aim of enforcing the effective measures of the protection of both health
and the environment, it could be necessary to adopt integrated approaches, including
the prevention, monitoring, and control of PFASs, the use of alternative chemicals, which
should be less toxic and should not persist in the environment, and a strong cooperation
among governments.

5. PFAS Analysis on Environmental Matrices

5.1. Surface Water, Underground Water, Wastewater
Consistent levels of PFASs have been identified in several environmental matrices. Pre-
dominantly, the aquatic environment is the most polluted [65]. Water is an essential element
for the entire terrestrial ecosystem, and its pollution is a threat for all living creatures [66].
For this reason, an accurate evaluation of the concentrations of perfluorinated compounds
in various aqueous matrices is necessary. The procedure of PFAS analysis for water samples
usually consists of SPE, followed by LC-MS [67–72]. Table 1 summarizes the analytical
procedures of samples from some recent studies on the analysis of PFASs in surface water,
wastewater, and groundwater, and its recoveries, LOD, and LOQ. The comparison shows
differences in the types of cartridges, extraction solvents, and analytical parameters used
to determine different recovery, LOD, and LOQ rates. Aqueous samples usually contain
a low level of PFASs (ng L−1 or pg L−1 ). Therefore, greater attention must be paid to the
sample preparation, focusing on the clean-up of the sample matrix background and the
pre-concentration of PFASs [65]. Brumovsky et al. (2018) [28] assessed the influence of the
type of sorbent, the matrix pH, the salinity, and the eluent on PFAS recovery. Firstly, they
used three different SPE sorbents for the study: StrataTM -X, Oasis® hydrophilic–lipophilic
balance (HLB), and Oasis® weak anion exchange (WAX). Thereafter, they prepared three
matrices with different salinities: tap water, artificial seawater, and a 1:1 mixture of the two.
They obtained the highest recovery of the studied PFASs using the Oasis® HLB sorbent,
with methanol (MeOH) as the eluent, at a pH = 8, and with 50% (brackish water) or 100%
(marine water) seawater content (82.8% and 78.3%, respectively). They found slightly lower
values using the StrataTM -X sorbent at the same pH and salinity values (75.6% and 71.3%,
respectively). On the other hand, the Oasis® WAX sorbent provided lower recoveries. This
sorbent allows for the washing of the adsorbed analytes using MeOH, as PFCAs and PFSAs
are reluctant to be eluted under these conditions, representing an advantage when the
removal of interfering organics is required. Similarly, Janda et al. (2019) [73] evaluated
two sorbents with weak anion exchange properties (Oasis WAX and Strata X-AW) for the
optimization of the SPE recoveries at different pHs, and two stationary phases for LC
separation for the determination of PFCAs with chains from C2 to C8 , as discussed later.
Analytica 2024, 5 192

The obtained PFCA recoveries from C4 to C8 were between 76 and 133% for both SPE
materials, regardless of the sample pH, whereas the extraction efficiencies of trifluoroac-
etate (TFA) and perfluoropropanoate (PFPrA) depended on the sample pH. They obtained
PFPrA recoveries between 93 and 103% for all pH values using the Oasis WAX sorbent,
while the recoveries with Strata X-AW were acceptable in the pH range from 3.0 to 5.0
(from 94 to 114%), and then decreased at a pH of 6.0 (36%). They achieved the quantitative
extraction of TFA between pH 3.0 and 5.0 (95 to 103%) using the Oasis WAX sorbent, while
the Strata X-AW sorbent recovered TFA at 99% (pH 3.0) with reduced extraction recoveries
at higher sample pH levels. They selected the Oasis WAX sorbent for the SPE of surface,
groundwater, and drinking water samples, and adjusted the sample pH to 3.9 ± 0.1 before
extraction. They performed the analysis using a liquid chromatography–tandem mass spec-
trometry (LC-MS/MS) system with a core–shell C18 column and an acidified eluent. On
the other hand, Lockwood et al. (2019) [68] demonstrated the use of µSPE as an alternative
to conventional SPE for PFAS extraction. The authors compared the efficiency of four
sorbents for the extraction of 13 PFASs from ultrapure water. The sorbents studied were
C18 , aminopropyl silica (APS) phase, pentafluorophenyl (PFP), and diol sorbent. Among
them, C18 provided good retention for most compounds, while APS was the only phase
which was capable of retaining the compound in the chain short perfluorobutanoic (PFBA).
For these reasons, the combination of the reversed-phase C18 sorbent and APS (as a 50:50
mixture) was chosen to pack the µSPE cartridge. This method reduced the extraction times
to 5 min compared to conventional SPE methods, and achieved similar results while using a
smaller sample volume. Additionally, the automation of the extraction method was demon-
strated using an eXact3 digital syringe and an ePrep® sample preparation workstation.
Regarding the separation and detection of PFASs in water samples, LC-MS is the most used
technique [65,67–74]. In the study reported above, Janda et al. (2019) [73] also studied two sta-
tionary phases for LC. The authors evaluated a mixed-mode phase, offering both ion exchange
and HILIC (Obelisc N) and a core–shell RP column (Kinetex C18 ) for chromatographic sepa-
ration. Using the Obelisc N column, it was possible to obtain a separation of the studied
PFCAs in ultrapure water; however, in the environmental samples, the separation was
disturbed by matrix effects. For this reason, the study continued using the Kinetex C18
column. In the final method, a binary gradient was applied (eluent A: 2 mM ammonium
formate (AF) and 0.2% formic acid (FA) in H2 OmQ /MeOH (4:1, v/v); eluent B: 2 mM AF in
MeOH). FA was added to eluent A, as it was observed to promote the retention of TFA and
PFPrA. Mulabagal et al. (2018) [69] investigated the presence of PFASs in estuarine water
using ultra-high performance liquid chromatography–triple quadrupole mass spectrometry
(UHPLC-QqQ-MS). Several parameters were examined to optimize the analysis as follows:
column type, solvents in the mobile phase, flow rate, and column temperature. Best results
were obtained with an Agilent ZORBAX RRHD Eclipse Plus C18 column, thermostated at
a T of 40 ◦ C, and a binary solvent gradient consisting of 5 mM AF in water and MeOH.
In addition, a guard column was used to achieve reproducible retention times. Following
method optimization, recovery experiments were performed via adding 23 PFAS analytes
and the internal standard (ISTD) to ambient water samples. Zhu et al. (2020) [75] investi-
gated a simple method of the derivatization of PFOAs, which achieves the enabling of the
separation of its isomers/enantiomers via the use of gas chromatography–electron capture
negative ionization mass spectrometry (GC-ECNI-MS) detection. (S)-1-phenethyl chloride
(PhEtCl) was used as a derivatizing agent; this reacts with PFOA enantiomers to form
PFOA (R)-1-phenylethyl esters. Derivatization was performed via adding 100 mL of 5%
PhEtCl in acetone (ACE) to a standard extract. The GC analysis was carried out by means
of a non-chiral GC column (HP-5MS column). The method was subsequently applied to
river water samples for measuring PFOA isomer/enantiomer concentrations. Samples
were preliminary treated with an Oasis WAX plus SPE cartridge, and then were subjected
to a derivatization step. Various PFOA isomers/enantiomers, mainly 3m-PFOA, 4m-PFOA,
6m-PFOA, and L-PFOA, were detected in the river water sample, with concentrations of
5.2, 3.8, 41, and 3400 ng L−1 , respectively.
Analytica 2024, 5 193

Table 1. Summary of the analytical procedure for the analysis of PFASs in surface water, wastewater,
and groundwater and their recoveries (%), LOD (ng L−1 ), and LOQ (ng L−1 ). N/A = not available,
a = pg/injection.

Extraction and
Matrix Analysis Recoveries LOD LOQ Ref.
Pretreatment
High pressure liquid
Surface and
Oasis WAX cartridge chromatography–mass
underground 80–120 N/A 0.2–5 [65]
SPE online spectrometry
water
(HPLC-MS)
Ultra-high performance Above 50
Oasis WAX cartridges
Seawater liquid chromatography except PFBA in 0.001–0.331 N/A [67]
SPE
(UHPLC-MS) seawater (<10)
Surface water µSPE cartridge UHPLC-MS 86–111 0.29–6.6 N/A [68]
Ultra-high performance
liquid chromatography
Waters Oasis PRiME
Estuarine water tandem mass 78.5–112.6 0.48–1.68 a 1.7–5.4 a [69]
HLB cartridges SPE
spectrometry
(UHPLC-MS/MS)
Ultra-performance liquid
chromatography–
Surface water Waters Oasis HLB
tandem mass 72–98 0.02–0.03 0.04–0.06 [70]
and rainwater cartridges SPE
spectrometry
(UPLC-MS/MS)
Surface water WAX SPE cartridges LC-MS/MS 68.5–118 0.5–10 2–20 [71]
Oasis WAX cartridges
Groundwater LC-MS/MS 73–137 0.03–0.06 0.03–1.27 [72]
SPE
Surface water
Oasis WAX cartridges
and LC-MS/MS 83–107 0.1–5.5 0.6–26 [73]
SPE
groundwater
Oasis® cartridge SPE
Seawater and Envi-CarbTM LC-MS/MS 63.2–112.9 0.01–0.08 0.03–0.24 [74]
cartridge
Derivatization using
Surface water GC-MS 92 0.050–0.15 N/A [75]
(S)-1-PhEtCl
Liquid chromatography
triple quadrupole mass
Surface water Strata-X cartridge SPE 44–100 N/A 0.01–2.00 [76]
spectrometer
(LC-QqQ-MS)
Water Oasis HLB Plus
Groundwater UPLC-MS/MS 70–130 0.02–0.03 0.04–0.06 [77]
Short cartridges SPE

5.2. Soil and Sediments

Soil contains over 40 classes of PFASs, usually originating from human activities, such
as industrial operations and waste disposal [78]. Table 2 summarizes the extraction, pre-
concentration, and analysis techniques of some recent studies, along with their respective
recoveries, LOD, and LOQ. The main differences among the studies are in the types of
cartridges and analysis techniques used. Sammut et al. (2019) [77] conducted the extraction
procedure using MeOH as the solvent and a short Waters Oasis HLB Plus cartridge for
the SPE. They used UPLC-MS/MS with an ACQUITY UPLC BEH C18 column for the
separation. Zhong et al. (2021) [79] used MeOH as the extraction solvent, and performed a
clean-up using the SPE with Supelco® ENVI-Carb cartridges. They performed the analysis
and quantification via a LC-QqQ-MS system, using an ACQUITY UPLC BEH C18 column
for the separation. They obtained recoveries ranging from 96 to 127.1%, while the LOD and
LOQ were 0.002–0.018 ng g−1 dry weight (dw) and 0.004–0.054 ng g−1 dw, respectively.
Simon et al. (2022) [69] optimized a fast and simple SPE-free extraction method, based on
liquid–solid extraction (LSE) using acidified MeOH. They compared the method with an
additional SPE purification step, and showed that the clean-up step was unnecessary, as it
Analytica 2024, 5 194

resulted in an underestimation of the organically bound extractable fluorine (EOF) concen-

trations. They carried out the analysis via by high resolution-continuum source-graphite
furnace molecular absorption spectrometry (HR-CS-GFMAS). Peng et al. (2013) [80] de-
veloped a different technique to detect fluorotelomer alcohols (FTOH), using a derivatiza-
tion reaction and liquid chromatography–electrospray mass spectrometry (LC-ESI-MS) in
combination with low-energy collision-induced dissociation tandem mass spectrometry
(CID-MS/MS). They used acetonitrile (ACN) as the solvent, and a WAX cartridge for
the SPE for the extraction. They performed the derivatization by adding dansyl chloride
(DNS) in acetonitrile and 4-(dimethylamino)-pyridine (DMAP) in dichloromethane (DCM)
as the catalyst. They used ultra-performance liquid chromatography collision-induced
dissociation tandem mass spectrometry (UPLC-CID-MS/MS) with a Waters ACQUITY
UPLC BEH phenyl column for the analysis. Overall, the method recoveries ranged from
67 to 83%. The LOQ for all FTOHs were 0.017–0.060 ng g−1 . They applied the method to
analyze six marine sediment samples. All FTOHs, except for 10:1 FTOH, were detected,
and the total concentrations of FTOHs were 0.19–0.52 ng g−1 dw. The developed method
provides a new method to sensitively determine FTOHs in environmental matrices.

Table 2. Summary of the analytical procedure for the analysis of PFASs in soil and sediment and their
recoveries (%), LOD (ng g−1 ), and LOQ (ng g−1 ). N/A = not available, a = ng mL−1 , b = method
quantification limit (MQL, ng g−1 ), c = method detection limit (MDL, ng g−1 ).

Extraction and
Matrix Analysis Recoveries LOD LOQ Ref.
Pretreatment
Sea and river
Oasis® cartridge SPE and 0.002–0.042
sediment and LC-MS/MS 78.9–120 c 0.006–0.1 b [74]
ENVI-CarbTM cartridge
soil
Extraction with acetic acid
(AcOH) and MeOH;
Sediment LC-QqQ-MS 44–100 N/A 0.04–8.00 [76]
cleaning up by Strata-X
cartridge SPE
Water Oasis HLB Plus
Soils UPLC-MS/MS 70–130 0.02–0.50 0.04–0.60 [77]
Short cartridges SPE
Oasis WAX cartridges,
Soils LC-MS 52–167 0.100–400 c 0.01–0.30 b [78]
MeOH as solvent
Marine ENVI-Carb cartridges SPE,
LC-MS 96–127.1 0.002–0.018 0.004–0.054 [79]
sediments MeOH as solvent
Derivatization dansyl
Liquid chromatography–
chloride (DNS) in ACN
electronspray
under catalysis of 4-
Marine ionization–mass
(dimethylamino)-pyridine 67–83 0.006–0.016 0.017–0.060 [80]
sediment spectrometry
(DMAP) and
(LC-ESI-MS) combined
WAX and silica cartridges
with CID-MS/MS
clean-up
Soils LLE using acidified MeOH HR-CS-GFMAS 79–117 3.43 10.30 [81]
High performance liquid
chromatography–
MeOH-based extraction, electrospray ionization 0.013–0.089
Sediment 86.7–113 a N/A [82]
Oasis WAX SPE cartridges tandem mass
spectrometry
(HPLC-ESI-MS/MS)
Pre-treatment with
Sediment Oasis-HLB SPE and MeOH HPLC-ESI(-)-MS/MS 72–101.9 N/A 0.01–0.08 b [83]
solvent
ACN and MeOH as the
Sediment UPLC-MS/MS 80–115 0.02–1.11 a N/A [84]
extraction solvent
Analytica 2024, 5 195

5.3. Air and Dust

As reported above, many studies have focused on the measurement of PFAS concen-
trations in surface water, groundwater, soil, and sediments, while much less is known about
atmospheric concentrations. The determination of volatile PFASs is crucial for studying
their sources and fate. Volatile PFASs can be present in air and can be transformed into
highly persistent PFCAs [85]. Dimzon et al. (2017) [86] investigated and validated a method
for determining volatile PFASs in the air and water of industrial and municipal wastewater
treatment plants (WWTP) in the Netherlands and Germany. During the study, a sampling
and enrichment method was developed for volatile PFASs in air samples using Oasis
HLBTM SPE cartridges. They used a low-volume air sampling device for air sampling. They
spiked the cartridge with 20 µL of an enrichment control standard solution, attached it to a
membrane pump, and eluted the analytes with 2 mL of n-pentane (C5 H12 ). Furthermore,
1 µL of the sample was analyzed using GC-MS. They performed the mass analysis in
a programmed selected ion monitoring (SIM) mode. They used different temperature
ramp rates in the oven program for the GC separation of the different PFASs. They also
chose a splitless injection to avoid system losses. They obtained an approximate MDL of
1 ng m−3 in the air, except for fluorotelomer acrylates (FTACs), which they could detect
at >40 ng m−3 . They obtained recoveries between 60 and 120% for all analytes using
the volatilization and direct addition methods. Humans might be exposed to PFASs via
indoor air and dust. Padilla-Sánchez et al. (2016) [87] developed and validated a fast and
sensitive analytical method for the analysis of a large number of PFASs in indoor dust.
They collected dust samples from vacuum cleaner bags, extracted them by adding MeOH,
and then filtered them with a syringe coupled to a 0.45 µm PP filter. They mixed the
extract with 0.25 mg of activated charcoal. They performed the analyses using ultra-high
performance liquid chromatography–time-of-flight mass spectrometry (UHPLC-TOF-MS),
equipped with an Oasis SPE column that allowed on-line SPE cleaning prior to analysis. In
this study, the detection performed using ESI-MS was used, operating in sensitivity mode
and negative ionization mode (NIM). They obtained MDLs and MQLs in ranges from 0.008
to 0.846 ng g−1 and 0.027 to 2.820 ng g−1 , respectively, and recoveries in ranging from 58 to
114%. Similarly, Ao et al. (2019) [88] collected dust from carpet and floor surfaces using a
domestic vacuum. They used accelerated solvent extraction (ASE) for the extraction and
SPE for the clean-up of the samples. They used an Oasis WAX cartridge for the SPE process.
The analysis was conducted using an UPLC-MS/MS. They obtained LODs and LOQs in
the range of 0.08–0.68 pg g−1 and 0.26–2.25 pg g−1 , respectively, for the target analytes.
They obtained recoveries from 80.2 to 95.2%. PFASs have also been detected in remote
regions, such as the Atlantic and Antarctic regions. Wang et al. (2015) [89] investigated the
PFAS presence in the atmosphere across these regions. They conducted the air sampling
onboard a research vessel. They collected a total of 33 air samples using a high-volume
air sampler. They used a cartridge containing polyurethane foam and polystyrene divinyl
benzene copolymer (PUF/XAD® -2) for sampling. They carried out the extraction in a
Soxhlet extractor using DCM for 16 h. The analysis was performed using GC-MS in SIM
mode with a positive chemical ionization (PCI) mode. They used a SUPELCO WAX 10
column for the separation, MeOH as the reagent gas, and helium as the carrier gas. They
obtained an MDL in the sampled air, ranging from 0.01 to 0.10 pg m−3 . They determined
the total gas phase concentration of 12 PFAS, which ranged from 2.8 to 68.8 pg m−3 , with
an average value of 23.5 pg m−3 . Camoiras González et al. (2021) [90] also used the PUF in
their study, where they monitored the air in developing countries with a passive sampler.
They exposed the PUFs for 3 months and then the extracted the analytes using a Soxhlet
with a 60:40 methyl tert-butyl ether (MTBE)/MeOH mixture for 24 h. They performed the
clean-up of the samples in the first period of the study using an ENVI-Carb SPE tube, and
collected them with MeOH. In the second period of the study, they stacked the ENVI-Carb
SPE tubes under a SPE-WAX cartridge. All the sample were analyzed with a LC-MS/MS,
with ESI operating in NIM. Recoveries were in the range of 30–150% for all target analytes,
and LOQs for PFSAs and PFAAs were between 11 pg/PUF and 14 pg/PUF. Wild birds have
Analytica 2024, 5 196

also proven useful in the biomonitoring of environmental contaminants. For these reasons,
in a recent study, Groffen et al. (2021) [91] developed an analytical method to determine
a broad range of PFASs, using bird feathers as indicators of PFAS contamination. The
feathers were collected from the ground, having fallen from free-range domestic chickens
near a fluorochemical plant in Belgium. Simultaneously, non-contaminated feathers were
collected at a site with low PFAS contamination. The samples were washed to remove soil
and dust, cut into small pieces, and, for extraction, 10 mL of 100% MeOH and 10 ng of bulk
labeled ISTD solution were added. Subsequently, the samples were vortexed for 1 min and
left in the dark for 24 h at room temperature. After centrifugation, the supernatant was
dried, and then reconstituted with 2 mL of a 2% ammonium hydroxide (AH) solution in
ACN, vortexed for at least 1 min, and filtered through a 0.2 µm Supor polyethersulfone
ion chromatography Acrodisc 13 mm syringe filter (PES). For the analysis, UPLC-MS/MS
with a negative electrospray ionization mode (ESI-) was employed. An ACQUITY BEH
C18 column was used for the separation. Different gradients of FA and water were used
during the analysis. Target analytes were quantified using the multiple reaction monitoring
(MRM) of two diagnostic transitions per analyte. The extraction recovery ranged from
67.3% to 112%, while the LOD and LOQ values for all the analytes were between 0.09 and
25.8 ng g−1 wet weight (ww) and 0.29 and 86.0 ng g−1 ww, respectively.

6. Discussion
Numerous studies focusing on the extraction and analysis of PFASs suggest SPE as the
best procedure for extracting PFASs from environmental samples. This technique shows
high versatility as it is used to extract different categories of contaminants (PAHs, phthalates,
PCB, pesticides), including PFASs from a wide variety of samples (biological, food, and
environmental). Moreover, it is useful as a preconcentration and clean-up approach for
the isolation and separation of the analyte, removing all the interference that could affect
the chromatographic analysis. The concentration of the analyte in a small volume solvent
enhances the accuracy and reproducibility of the analysis. Therefore, many parameters
such as temperature, matrix pH, salinity, and the choice of specific adsorbent need to be
optimized in order to achieve a better extraction efficiency. Despite the versatility of SPE
being seen as a benefit, it can also be a drawback due to the potential lack of selectivity.
A few sorbents like Oasis HLB and WAX Oasis are used for the extraction of numerous
target compounds, which are simultaneously retained [92].
Regarding the water samples, the best recoveries of PFASs were achieved using Oasis
WAX (73–137%) and Water Oasis HLB Plus Short SPE (70–130%) cartridges at pH values
between 6 and 8; on the other hand, slightly lower recoveries (44–100%) at pH values
between 3 and 5 are shown with Strata-X cartridge. However, the µSPE procedure, when
applied as an alternative to traditional SPE, bypassed the limitations of SPE via the use
of smaller adsorbent sizes (<5 µm), leading to an increase in the extraction efficiency
and lower solvent volumes (2 mL) and extraction times (5 min). Hence, the optimized
mixture of C18:APS adsorbents (50:50) has allowed us to obtain satisfying recoveries
for short- and medium-chain PFASs (C4-C10) and sulfonated fluorotelomers (96–105%)
with RDS < 14%, excepting for PFOSs (recovery: 129%; RSD < 29%). In addition, the
derivatization procedure based on the identification of PFAS isomers/enantiomers has
proven to be particularly useful for the evaluation of the potential toxicity of PFASs.
Specifically, isomers/enantiomers of PFOAs, such as 3m-PFOA, 4m-PFOA, 6m-PFOA,
and L-PFOA, are detected in river water samples at concentrations of 5.3, 3.8, 41, and
3400 ng L−1 , respectively. Derivatives of 5m-PFOA, 4.5 dm-PFOA, and 2m-PFOA are not
detected, whereas the recovery of 3C4-PFOA was 92%. Although the derivatization process
allows us to reduce the persistence of PFASs, attention should be paid to the production of
by-products (i.e., PFHpS, PFHxS, PFPeS, PFBS, PFOA, PFHxA, and PFBA), which could
have unknown effects on health and the environment. Furthermore, difficulties that could
be found are a loss of analytes in traces during the pretreatment and derivatization steps,
matrix effects with detection difficulties resulting in false positives, and the evaporation
Analytica 2024, 5 197

and loss of volatile compounds such as PFAS esters when transferred from the aqueous to
the organic phase. In addition, the analysis can be limited since only a particular class of
PFASs with the same fractional group can by analyzed via derivatization [93].
For soil and sediment samples, the derivatization of fluorotelomeric alcohols (FTOH)
with dansyl chloride (DNS) in acetonitrile and 4-(dimethylamino)-pyridine (DMAP) in
dichloromethane (DCM) has demonstrated satisfactory recoveries (67 ± 6.0–83 ± 9.4%)
and matrix effects < 15%. Methanol is the solvent mostly used for PFAS extraction from
soil and marine sediments, showing interesting recoveries in combination with Oasis HLB
70–130%), ENVICARB (96–127.1%) and Oasis WAX (52–167%) cartridges.
Chromatographic analysis conducted using UHPLC-MS and LC-QqQ-MS are proved
to be the most sensitive, achieving low LOD and LOQ values in water (0.001–0.331 ng L−1 ;
0.01–2.00 ng L−1 ) and sediment (0.002–0.18 ng g−1 ; 0.004–0.054 ng g−1 ). Combined
with the conventional SPE procedures, they have been shown to be reliable and repro-
ducible by identifying the most relevant PFASs, such as PFOS (<LOD-5.91 ng g−1 ), PFOA
(LOD-0.58 ng g−1 ), and PFDA (<LOD-1.05 ng g−1 ) in real sediment, soil, and water sam-
ples. Moreover, although these analytical techniques are widely developed, they are only
effective for the determination of a limited number of analytes. When compared to water
and soil matrices, the number of studies focused on determining PFASs in the atmosphere
are still limited. Analytical methodologies based on sampling with air samplers and en-
richment using HLB and Oasis WAX cartridges have reported good recoveries of PFASs in
air (60–120%) and dust (58–114%). Moreover, the UHPLC-TOF-MS analysis demonstrated
a good sensitivity (MDL = 0.008–0.846 ng g−1 ; MQL = 0.027–2.820 ng g−1 ), finding PFAS
concentrations in real indoor dust samples in the range from 0.02 (PFHxS, NIPH 1) to
132 ng g−1 (6:2-diPAP, NIPH 4).

7. Conclusions
The relevant use of PFASs in various industrial sectors has raised considerable con-
cerns about their harmful effects on the environment, humans, and the entire ecosystem.
The ingestion and inhalation of contaminated air particles form the pathways of environ-
mental PFAS exposure. Toxicological and detrimental hazards on respiratory, digestion,
cardiovascular, and neurological systems have been proven, thus highlighting the necessity
of developing a sensitive analytical method for their detection and removal. However, the
widespread occurrence at a low level concentration (pg L−1 to mg L−1 ) into the environ-
ment has enforced governments to set limits and restrictions regarding their applications. In
this review, several analytical approaches for the extraction and following chromatographic
separation of PFASs in environmental matrices have been discussed. SPE procedures
followed by LC-MS analysis are the main approaches applied. The average recoveries
using Oasis® s HLB (85–100%) and Oasis® WAX (93–111.5%) cartridges have been con-
firmed as providing the best performance for the purification and PFAS extraction in water,
sediment, and soils samples. In addition, air and dust samples have also showed recov-
eries of PFASs between 60 and 120% and 80.2 and 95.2% using HLBTM SPE and Oasis
WAX cartridges. The results obtained could be explained via the flexibility of the Oasis®
HLB (N-vinylpyrrolidone-divinylbenzene copolymer) sorbent for the extraction of a large
variety of compounds of different polarities, as well as by the similar structure to the
Oasis® HLB of the Oasis® WAX sorbent (styrene-divinylbenzene polymer modified by a
weak anion-exchange group piperazine), highly affine to anionic short-chain compounds
at high pH levels. Very low LODs and LOQs were observed in seawater (0.01–0.08 ng L−1 ;
0.03–0.24 ng L−1 ), marine sediment (0.002–0.018 ng g−1 ; 0.004–0.054 ng g−1 ), and dust
(0.08 to 0.68 pg g−1 ; and 0.26 to 2.25 pg g−1 ) through LC-MS analysis. There are many
advancements and innovative approaches being developed to improve the extraction,
analysis, monitoring, and remediation of PFASs in the environment. These include the
use of passive sampling techniques and alternative analytical techniques, which can help
in identifying and monitoring PFASs in the environment. For example, the use of the
NTA-HRMS (non-target analysis-high-resolution mass spectrometry) approach allows
Analytica 2024, 5 198

us to identify the structures of unknown PFASs, distinguishing ions which are slightly
different in their mass/charge ratio with high mass resolution, selectivity, and sensitivity.
Advanced water and wastewater treatment technologies are effective for PFAS removal, but
may require high operating costs and generate toxic waste. Therefore, PFAS removal and
degradation technologies (electrocoagulation, foam fractionation, bioremediation, chemical
oxidation/reduction, advanced oxidation/reduction processes), as well as PFAS adsorp-
tion on various materials (porous organic polymers (POPs) or low-cost carbon-derived
materials) are being explored for an efficient PFAS remediation from both water and soil. In
addition, the use of PFAS substitutes, such as carboxylic acid fluorotelomer 6:2 (6:2-FTCA),
perfluoroalkyl carboxylates (PFECAs), GenX, ADONA, PFPiA, perfluoroalkyl ether car-
boxylic acid (PFESA), perfluorobutanesulfonic acid (PFBS), and perfluorohexanoic acid
(PFHxA), which are less harmful than some specific PFASs (PFOA and PFOS), can help
reduce the toxicological risks associated with them.

Author Contributions: Conceptualization, P.A. and C.D.F.; methodology, F.C.; software, F.C.; vali-
dation, A.I., C.D.F. and F.C.; formal analysis, A.I.; investigation, F.C.; resources, A.I.; data curation,
F.C.; writing—original draft preparation, A.I.; writing—review and editing, F.C.; visualization, A.I.;
supervision, P.A.; project administration, P.A.; funding acquisition, C.D.F. All authors have read and
agreed to the published version of the manuscript.
Funding: This research received no external funding.
Data Availability Statement: Not applicable.
Conflicts of Interest: The authors declare no conflicts of interest.

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