Per- and Polyfluoroalkyl Substances: An Overview of the Health Effects on Human Body

1. Introduction

Pollution by organic substances is becoming a global problem not only for the environment but also for human health. Such substances include, for example, pharmaceuticals, pesticides, poly- and perfluorinated substances (PFAS), etc. PFAS are anthropogenic microcontaminants that, due to their abundant occurrence, have an adverse impact not only on the ecosystem, but also on human health. The most studied substances are perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS) [1,2,3]. Due to its potential multi-organ toxicity and adverse effects on human health, PFOS was the first compound to be regulated by the European Commission in 2006 (EU, 2006) [4]. It was listed in the Stockholm Convention on Persistent Organic Pollutants (POPs) in 2009 [5]. PFOA was listed in 2019 [6] and perfluorohexanesulfonic acid (PFHxS) in 2022 [7].

The first classification of PFAS was presented by Buck et al., [8], who defined them as fluorinated aliphatic substances containing one or more carbon atoms, where the aliphatic hydrogens may be partially or fully replaced by fluorine atoms. In 2018, the Organisation for Economic Co-operation and Development (OECD) defined PFAS as substances that contain -CnF2n+1 or -CnF2n- (n ≥ 1). The existing classification system was further modified by Buck et al. [9] so that PFAS are divided into two groups: polymers and non-polymers. Non-polymers are divided into per- and polyfluorinated substances, with the perfluorinated ones having polar hydrophilic groups such as carboxylate (COO-), sulfonate (SO₃-) or phosphate (OPO₃-) mostly attached to a hydrophobic carbon chain. Polymers are divided into fluoropolymers, fluorinated side-chain polymers and perfluoropolyether [10]. They are not bioavailable and do not accumulate in the environment or biological systems. They gradually break down into PFAS. Often, long-chain PFAS are replaced by new short-chain alternatives, such as GenX [11]. Several alternatives have been proposed to meet the market requirements of PFOS and PFOA. For example, chlorinated polyfluoroethylene ether sulfonic acid (Cl-PFESA) with the trade name F–53 B has been proposed as a replacement for PFOS [12]; the ammonium salt of 2,3,3,3-tetrafluoro-2-(1,1,2,2,3,3,3-heptafluoropropoxy)-propanoic acid under the trade name GenX has been proposed as an alternative to PFOA [13]. Several studies have shown that alternatives to PFOA and PFOS are found in environment at comparable or higher concentrations than PFOA and PFOS [14,15]. For example, concentrations of GenX in surface water samples are three times higher than PFOA in the North Sea [16]. Concentrations of F–53 B in aquatic and soil environments in China are similar to those of PFOS [13,17].

The hydrophobic and oleophobic parts of PFAS compounds cause them to act as surfactants. They are widely used in various industries and consumer products, such as carpets, textile impregnations, fire-fighting foams, electroplating, adhesives, protective coatings, insecticides, household cleaners, cosmetics, electronics, explosives, food packaging, tea bags, and many others from the 1940s to the present. The United States Environmental Protection Agency (US EPA) has set maximum concentrations of PFAS in drinking water at 4 ng/L for PFOA and PFOS and 10 ng/L for PFHxS in 2024. PFAS are ubiquitous substances, found in various environmental compartments, such as air, surface water, drinking water, groundwater, sediment, soil, plants, food, and animals. Routes of exposure to PFAS by inhalation and dermal contact include dietary intake, indoor air, and drinking water [18,19]. As a result, PFAS are found in multiple human tissues, such as blood, urine, hair, nails, urine, placenta and breast milk.

2. Properties of PFAS

PFAS contain of a hydrophobic chain of varying lengths made up of carbon and hydrogen atoms and a hydrophilic functional group at the end of the chain. The hydrogens can be fully or partially replaced by fluorine atoms. Accordingly, we divide the compounds into perfluorinated, when all hydrogen atoms are replaced by fluorine atoms, and polyfluorinated, when some hydrogen atoms (attached to at least one carbon) are replaced by fluorine atoms. According to the chemical structure, we divide PFAS into a long-chain group (number of carbons 7-13) called legacy PFAS and a short-chain group (number of carbons 6 or fewer atoms) terms emerging or alternative compounds. Polyfluoroalkyl substances can be chemically modified into perfluoroalkyl substances [20].

The most studied are the perfluoroalkyl acids (PFAAs), which include long-chain compounds, such as 9-carbon perfluorononanoic acid (PFNA); 8-carbon compounds PFOA and PFOS; 7-carbon perfluoroheptanoic acid (PFHpA), and PFHxS. PFAAs also include short-chain 6-carbon compound perfluorohexanoaic acid (PFHxA); 5-carbon perfluoropentanoic acid (PFPeA); 4-carbon perfluorobutanoic acid (PFBA), and perfluorobutanesulfonic acid (PFBS). The classification of PFAS is presented on Figure 1.

The most commonly monitored PFAS are PFOS and PFOA, which are detectable in up to 90% of the population in the USA. Molecular structures of selected PFAS are shown on Figure 2.

PFAS consist of two parts, namely: an alkyl chain and a functional group, which can be a carboxylate or sulfonate. The alkyl chain, in which hydrogen atoms are replaced by fluorine, is responsible for the physical and chemical properties of PFAS. Fluorine has the high electronegativity and ionization potential and low polarization. It is the most electronegative element in the periodic table, therefore the bond between carbon and fluorine is among the strongest covalent bonds with a dissociation energy of up to 513.5 kJ.mol^-1. The strength of the bond increases with the number of F atoms bound to carbon atoms. The strong bond between C and F is responsible for the thermal stability of PFAS and their resistance to degradation, hydrolysis, photolysis, which is related to their long-term persistence in various components of the environment, but also in the food chain and in the human body [21,22]. They are further divided into polymers and non-polymers, with the polymeric ones degrading to non-polymers over time. Due to the low polarizability of fluorine, PFAS have weak intra- and intermolecular interactions and, as a result, higher volatility and lower boiling point. They have low surface tension and thus good surface wettability. The oleophobic character is a consequence of weak van der Waals forces. The combination of hydrophobic and hydrophilic properties allows PFAS to interact in the environment, they have the ability to combine into micelles, as a result of which they are capable of long-distance transport in air, water, and the atmosphere. Their high mobility provides the possibility of being transported over long distances, for example to remote areas such as the Arctic or Antarctica. PFAS are soluble in both non-polar and polar solvents, and their boiling and melting points increase with chain length. PFAS with higher vapor pressure exist in the gaseous state, with lower vapor pressure they remain solid or liquid [21,22].

The most commonly monitored physicochemical parameters include pH, temperature, turbidity, salinity, electrical conductivity (EC), dissolved oxygen (DO) and environmental factors (such as chain length, seasonal variations, organic content, solubility, partition coefficient, redox potential) that affect the fate, mobility and occurrence of PFAS in the aquatic ecosystem [21,22].

The study by Ohoro et al. found that at high pH, ​​turbidity and dissolved oxygen, PFOS concentrations are high. Conversely, high electrical conductivity, temperature and salinity cause a decrease in PFOS concentrations in water. The protection of aquatic animals with regard to environmental safety, as well as human safety, is necessary [23].

3. Sources of PFAS Exposure

There are multiple routes of human exposure to PFAS, with the main sources of exposure being drinking water, air, indoor dust, soil, food, food contact materials, kitchen utensils, mother-to-fetal transmission and breastfeeding to newborns However, it is expected that 98% of the US human population may have detectable levels of PFAS in their blood. PFAS are carcinogens that disrupt the endocrine system, genotoxicity, immunotoxicity, reproductive toxicity, and hepatic toxicity [24].

In the adult population, food and drinking water are the main sources of exposure, but this is influenced by overall lifestyle and the level of contamination of drinking water. Infants fed formula may be among the most exposed due to high water intake per body weight [24,25,26].

Due to their exceptional properties, such as stability, water resistance, etc., they are widely used in industry and commercial products. They can enter the environment through various routes: industry, emissions, product use, lividation, and the result is insufficiently effective removal of PFAS by wastewater treatment plants, resulting in soil and water contamination. Their degradation and removal from the environment depend on the properties of PFAS, especially the chain length, type of head group, degree of ionization and fluorination. Accumulation in humans occurs through dermal contact, inhalation or ingestion and can cause organ-specific toxicity, such as hepatotoxicity, neurotoxicity, immunotoxicity, etc. Long-chain PFAS accumulate in the body for a longer period of time and can result in health problems such as hormonal imbalance, decreased immune function, liver damage, various developmental and reproductive anomalies, and an increased risk of cancer. The toxicokinetic profile of PFAS in humans consists of absorption, distribution, metabolism, and excretion [27]. PFAS absorption can occur orally, inhaled, and dermally. The oral route predominates (>90%), mainly through contaminated food and drinking water [28]. Inhalation can occur when handling consumer products or in work environments where products containing PFAS are manufactured [29]. Dermal contact occurs when using personal care products or when coming into contact with surfaces contaminated with PFAS [30]. The rate and extent of absorption in the human body depends on the physicochemical properties of PFAS, namely: molecular weight, chain length, functional groups, hydrophobicity, lipophilicity, solubility, partition coefficient, volatility Short-chain PFAS, such as PFBS, PFHxA are more soluble in water than long-chain PFOS and PFOA, which bioaccumulate in the body over a longer period of time [31].

Investigating the distribution of PFAS is a critical point for PFAS are known to cross the placenta and PFAS have also been found in the fetus. There have been concerns about adverse effects on fetal development, particularly through disruption of thyroid hormones, which are essential for normal fetal neurobehavioral development [32].

In a study by Høyer et al. [33], the association between prenatal exposure to PFHxS, PFHpA, PFNA and PFDA, PFOA and PFOS and behavior and hyperactivity in 5-9 year old children from Greenland and Ukraine was investigated. A country-specific parental standardized questionnaire was used to assess behavior in 1023 children. Parents rated their children’s behavior over the past six months. PFOS exposure levels were higher in Greenland compared to Ukraine. Foods such as fish, seabirds and marine mammals, which have high levels of trace elements, are a major part of the diet. This fact explains the differences in PFAS exposure. The results suggest that some of the associations between PFOS exposure and hyperactivity in some children may be due to postnatal exposure through breastfeeding. In the case of learning disabilities in children, more detailed investigation will be needed as levels of PFHxS and PFNA increase with decreasing PFOS levels [33].

Bharal 2024 et al. reviewed the persistence of PFAS, exposure routes, and toxicological profiles. They provided a comprehensive review of neurotoxicity studies and their mechanisms induced by PFAS exposure [34].

4. Analytical Techniques

The chemical structure of PFAS allows them to interact with the environment, which can result in toxic effects on humans and the environment, and therefore monitoring the impact of PFAS on human health is essential. Due to the relatively low concentration of PFAS in biological matrices and the complexity of the sample, continuous improvement of analytical methods is necessary, especially increasing the sensitivity and robustness of the method.

A review article by Kee et al. [35] describes human sample collection, sample preparation techniques, and instrumental techniques for sample analysis. When collecting, storing, and analyzing samples, the use of sampling material (tubes, sample containers, sampling tools) made of Teflon® (polytetrafluoroethylene, PTFE) should be avoided. Polypropylene (PP) and polyethylene (PE) materials are recommended. Since PFAS have the ability to adsorb to glass surfaces in aqueous environments, the use of glass devices should be avoided [36].

The most commonly used method for the determination of concentration of PFAS in various human samples (blood, plasma, serum, urine, hair, nails) is the combination of liquid chromatography (mainly UHPLC) with tandem mass spectrometry (MS/MS) using negative electrospray ionization (ESI). In addition, high-resolution mass spectrometry (HRMS) is used for non-targeted analysis for identifying new PFAS [37,38,39]

Less frequently, gas chromatography (GC) combine with a flame ionization detector (FID), an electron capture detector (ECD) or a mass spectrometer (MS) has been used as well [40]. GC-MS is used, especially for the analysis of volatile PFAS [41,42,43].

Eliminating matrix interferences and simultaneously increasing the detection limit of PFAS in various complex matrices are essential steps in the analysis of PFAS. Matrix effects can cause false signal increases or decreases. Various extraction techniques are used for sample preparation. Review Dhiman et al. deals with different extraction techniques, analysis and detection of PFAS in different matrices [44].

For accurate assessment of PFAS exposure, proper matrix selection is necessary. Plasma and serum are most commonly used for practical reasons. Non-invasive matrices such as urine, hair, and breast milk are advantageous in terms of ease of collection and efficiency, especially for infants and children. Urine is suitable for monitoring short-chain PFAS. Highly volatile PFAS do not accumulate in hair, and PFAS levels in breast milk may not accurately reflect maternal exposure because breastfeeding may affect the rate of PFAS excretion [45].

The key task is the pretreatment of the biological matrix. The most commonly pretreatment techniques are solid-phase extraction (SPE) (using mainly hydrophilic-lipophilic balanced (HLB) sorbents; weak anion exchanger (WAX); polymeric cartridges; polyacrylonitrile and divinylbenzene/carboxen/polydimethylsiloxane fibres) [46]; liquid-liquid extraction (LLE) [47,48]; solid-liquid extraction (SLE) [49]; ultrasound-assisted extraction [50,51]; dispersive solid-phase microextraction [52]; dispersive liquid-liquid microextraction [53] or accelerated solvent extraction (ASE) [54]. A review article by Comito et al. [55] provides an overview of selected analytical methods for the determination of PFAS in various biological matrices as well as sample pretreatment in the period 2003-2022.

In this review, Table 1 presents selected examples of PFAS determination in conventional biological matrices after 2022.

Nuclear magnetic resonance (NMR) is not sensitive to matrix components and offers clean spectra. It allows PFAS to be quantified without requiring high reproducibility and cost-effectiveness [76,77]. Advanced and novel analytical methods include fluorimetric assays, which measure the fluorescence signal of common fluorophores in solution. An interesting technique is molecular imprinting, which combines fluorescent dyes with molecularly imprinted polymers (MIPs). Electrochemical techniques are also used to detect PFAS, which help increase the sensitivity of the method [78].

5. Human Matrices

Measuring chemicals or their metabolites in different matrices is important for assessing human exposure and associated risks. About 77% of all published articles concerned blood, plasma and serum, as they best reflect internal exposure to PFAS. The disadvantage is the invasiveness of the collection, for example in newborns and children, due to ethical and humanitarian reasons, which limits the number of available samples. Therefore, non-invasive matrices such as hair and nails have several advantages including simple and easy sampling, transport, stable storage, and ethical acceptance. Moreover, hairs contain keratin, which is suitable for monitoring PFAS exposure. Another non-invasive sample is urine. Each matrix has its advantages and disadvantages. Hair and nails require simple collection, but due to external transfer they may contain various impurities that need to be removed [79].

5.1. Hair

Hair as a non-invasive matrix and contains 15-35% water, 63-95% protein, and 1-9% lipid, making it a suitable reservoir for hydrophobic compounds and their hydrophilic metabolites [79,80,81]. The review article Zhang et al. describes standardized procedures for measuring microorganic contaminants (MOC) in hair, sample collection and preparation, methods for determining chlorinated persistent organic pollutants, brominated flame retardants, non-persistent pesticides, per- and polyfluoroalkyl substances, phthalate esters, bisphenols and polycyclic aromatic hydrocarbons in hair, as well as conclusions from cohort and epidemiological studies. There are several factors that reduce the reliability of hair analysis. Detection of MOC is affected by age, gender, hair treatment and sampling method. Furthermore, it is necessary to distinguish between endogenous and exogenous contamination in hair, either by incorporation via blood or from the environment. Correlations between MOC concentrations in hair and in blood/urine are limited. Since hair has been less used in the past to assess the health risk of contamination with microorganic contaminants compared to blood/urine, there is a need to discuss techniques, experiments, and propose standardized procedures for their measurement [82].

In a study by Wang et al., concentrations of PFOS, PFOA, PFHxA, PFNA, PFDA, PFUnDA, PFDoA and PFHxS were determined in 39 human serum, urine, hair and nail samples by HPLC-MS/MS. PFHxA was detected only in nails and PFDoA was found in urine, hair and nails at very low concentrations. The concentrations of PFOS were 9.24 ng.mL^-1 in serum, 13.96 ng.L^-1 in urine, 0.58 ng.g^-1 in hair and 0.63 ng.g^-1 in nails and were higher in all matrices compared to other PFAS. A statistically significant difference in serum concentrations of PFOA and PFHxS was found between men and women. However, no statistically significant difference in PFAS concentrations was found between older and younger individuals in all matrices. The results of the study showed that the most suitable matrix for monitoring PFASs, especially PFOS, were nails [83,84].

PFAS can enter the hair in three ways: internal passive diffusion through the bloodstream; from the external environment and external transfer into hair shaft from sweat and sebum [85]. The memory effect of hair due to the accumulation of chemicals allows for retrospective analysis. In the case of hair, it is possible to increase the frequency of sample collection, as well as its amount, which is especially advantageous for toddlers. In an Indian study [86], e concentrations of 25 PFAS (6 PFSA, 13 PFCA, 6 PFAA precursors) were determined in 39 human hair samples. The effect of gender was also monitored. PFOA, PFOS and PFHxS were the most abundant in the samples. A highly significant correlation was found between PFOS and PFHxS. Women showed a higher incidence of PFAS than men. The article summarizes the results of PFAS analyses in hair obtained around the world. Compared to global results, PFAS values ​​in the Indian study reached lower concentrations [86].

Review article by Robin et al. presented the development and optimization of the method, including sample collection and preparation, extraction procedures, and instrumental techniques. Regarding hair sampling, the preferred site is the back of the head (posterior vertex, occipital region, or neck) or as close to the scalp as possible. Aluminum, envelopes, polypropylene tubes, or glass vials are used for hair storage [87].

Samples were mainly stored at room temperature, but storage in a refrigerator or freezer has also been reported. Combinations of aqueous and organic solvents; aqueous solvents; organic solvent and shampoo were used for hair decontamination. Hair samples were dried at room temperature or lyophilized. After external decontamination of the sample, hair was cut into small pieces or ground into powder. The samples were denatured with an organic solvent, most often methanol or by acid hydrolysis (HCl, acetic, formic, nitric acid) and denaturation followed by purification [88,89]. In general, LC-MS/MS or GC-MS/MS are applied for determination of polar or non-polar compounds, respectively. Prior to instrumental analysis, matrix interferents have to be removed usually by extraction methods (LLE, SPE or SPME) [90,91,92].

A Spanish study was aimed to determine PFBuA, PFPA, PFHxA, PFHpA, PFOA and PFOS in hair samples from children, women and men, assessing the potential relationship between PFAS concentration and age, gender, smoking and hair dyeing or coloring. The samples came from 42 volunteers, of whom 10 were children, 16 were women and 16 were men. The concentrations of PFAS ranged from 0.6-15.5 ng.g^-1, with PFHpA and PFOS being present in at least 86% and 76% of the analyzed samples, respectively. Longer-chain PFAS, such as PFOA and PFOS, are more commonly detected in hair samples, while shorter-chain PFAS (PFBuA) are more commonly detected in urine [93].

5.2. Placenta

The placenta is an important organ for the transfer of oxygen and nutrients between the mother and fetus. Various studies have focused on the capacity of the placenta to transfer PFAS and their alternatives. In a Chinese study, Pan et al. included 100 paired samples of human maternal and umbilical cord serum and found that chlorinated polyfluorinated ether sulfonates (6:2 and 8:2 Cl-PFESA) were detected in more than 99% of both matrices. Higher efficiency of placental transfer was associated with older maternal age, higher education, and lower glomerular filtration rate [94].

Gao et al. presented analysis of 132 paired samples of maternal and umbilical cord serum. The study showed that the extent of placental transfer of short-chain PFAS was in the range of 97-146% [95].

A review article by Blake et al. [96] focused on the placenta as a target organ and adverse pregnancy outcomes due to PFAS exposure. Specifically, these include prolonged gestation, pregnancy-induced hypertension, preeclampsia, gestational diabetes, and low birth weight. Studies by Chen [97] and Wang et al. [98] in human and animal models have shown that PFAS readily pass from maternal serum through the embryo to the placenta, which in the early stages functions as the liver, lungs, and kidneys for the embryo. Wang analyzed PFHpA, PFOA, PFNA, PFDA, PFUA, PFDoA, PFBS, PFHxS, PFOS and PFOSA in 369 pairs of maternal and cord serum in China. Almost all maternal and cord serum samples were found to contain all PFAS analyzed. All ten PFAS were found in both mothers and cord serum in almost all samples. Maternal and cord levels were closely correlated (r= 0.485-0.908) for all PFAS, except PFBS. The efficiency of transplacental transfer (TTE) was influenced by carbon chain length as well as functional group. Perfluoroalkylsulfonates had a lower maternal-fetal transfer ratio compared to perfluoroalkylcarboxylates [98].

In a review article, Liu et al. summarized the findings from previously published data on PFAS monitoring in maternal blood, cord blood, breast milk, placenta, amniotic fluid, fetal organs, dried blood spots of newborns, and infant serum; and on PFAS exposure during pregnancy and breastfeeding. PFAS concentrations determined in blood are relatively high, while in breast milk they are relatively low. They emphasized the importance of future research on PFAS isomers and enantiomers [99].

In Chinese study, there were tested 50 pairs of maternal and umbilical cord serum samples as well as placenta obtained from pregnant women living around the fluorochemical industrial park in Fuxin. 49 target PFAS in 11 classes were identified in human samples by HPLC-MS/MS [100]. PFBS, PFBA, and PFOA were the main contaminants from 21 target analytes of legacy PFAS in maternal and cord serum samples as well as placentas analyzed by HPLC-MS/MS. PFBS concentrations in maternal serum ranged from 3.6 to 140 ng.mL^-1; PFOA from 0.08 to 123 ng.mL^-1, and PFBA from 0.08 to 87 ng.mL^-1. PFBS, PFOA, and PFBA concentrations in newborns decreased by almost half compared to maternal serum. Similar concentrations were also measured in placenta samples. Using high-resolution MS (HRMS), 49 novel PFAS classified into 11 classes were identified in the primary screening, of which 20 novel congeners in 4 classes were discovered in human blood and placentas for the first time. The concentrations of most novel PFASs were higher in placentas and cord serum compared to maternal serum. In addition, the presence of novel PFAS in cord serum may have affected neonatal birth outcomes and serum thyroid hormone, sex hormone, and glucocorticoid levels [100].

5.3. Breast Milk

According to the World Health Organization, infants should be exclusively breastfed for the first 6 months [101], Mothers who cannot or do not want to breastfeed replace milk with formula, either in liquid form or diluted with drinking water. Infant formula manufacturers try to minimize the differences between infant formula and breast milk, and it is important to know that liquid sources of nutrition are safe for the baby. PFAS can be transmitted from mother to child through breastfeeding. Infants can also be exposed to PFAS through infant formula. A review article by LaKind et al. [102] compared concentrations of PFOA, PFOS, PFNA and PFHxS in breast milk and infant formula with screening values ​​for the listed PFAS in drinking water in children. Results from an earlier study confirmed by more recent data confirm that PFAS concentrations in breast milk are higher than screening levels for drinking water in children [103]. This is a global problem and it is essential that pregnant women and breastfeeding mothers have the necessary data available to inform their future decisions about breastfeeding.

Lamichhane et al. [104] studied the effect of maternal PFAS exposure on breast milk lipid composition, as well as the combined effect of exposed breast milk and breast milk lipid composition on infant growth. PFAS and lipid concentrations in maternal serum were measured in 44 mother-infant pairs using UPLC-Q/TOF MS. Lipidomic analysis of breast milk collected 2-4 days after delivery and in 3-month-old infants was also performed. Fecal biomarkers calprotein and beta defensin 2 were measured in the stool of infants aged 3, 6, 9, and 12 months to assess intestinal immunomodulatory function in the infant intestine. High PFAS exposure caused an increase in the ratio of acylated saturated and polyunsaturated fatty acids in triacylglycerols. Altered phospholipid composition due to high PFAS exposure has been linked to slower growth in infants. Maternal exposure to PFAS affects the quality of breast milk, which may affect the health and growth of children [104].

A Canadian study observed that the use of personal care products (e.g., hairsprays and gels, nailcare products, fragrances and perfumes, makeup, hair dye) may be associated with higher plasma concentrations of PFOA, PFOS, and PFHxS. Similar results were observed in breast milk. The study used prenatal plasma from 1.940 women at 6-13 weeks of gestation and human milk from 664 women 2-10 weeks postpartum. The frequency of use of personal care products was monitored during the 1st and 3rd trimesters, 1 to 2 days postpartum, and 2 to 10 weeks postpartum. Increased use of nail care products, fragrances, makeup, hair dye, hair sprays, and hair gels was observed in the 1st trimester, which caused increased plasma concentrations of PFOA and PFOS. Similar results were obtained in the 3rd trimester in plasma and in milk 2-10 weeks postpartum with the use of personal care products. In addition, the use of colored permanent hair dye 1-2 days postpartum may have caused higher concentrations of PFOA, PFOS, and PFNA in postpartum milk [105].

6. Health effects

PFAS exposure is associated with adverse health risks such as cancer, steroid hormone disruption, infertility, lipid and insulin dysregulation, higher cholesterol levels, liver and kidney disease, altered immunological and thyroid function, and cardiovascular effects [28,106,107]. In infants and children, PFAS exposure can cause adverse effects on infants and premature babies and can lead to reduced growth parameters, lower visual motor skills and attention deficit/hyperactivity disorder (ADHD) in childhood, lower levels of antibody concentrations against mumps and rubella, reduced lung and respiratory function, along with increased levels of glucocorticoids, progestogens, and uric acid [108,109,110,111].

The various sources, routes of exposure to PFAS and their adverse health effects on human health are illustrated in Figure 2.

PFOS and PFOA are representatives of PFAS, which are toxic to humans and animals. Due to their multiple toxicity, they have been banned in many countries and replaced by substitutes, which, however, have shown even higher toxicity. The most pronounced toxicity has been reproductive toxicity. Epidemiological studies have shown that PFOS and PFOA can cause a decrease in testosterone levels in humans, abnormal levels of sex hormones, an increase in the risk of infertility. PFAS can be absorbed through the intestinal or respiratory tract and subsequently cause reproductive disorders such as testicular cancer, testicular dysplasia syndrome and menstrual disorders [112].

Wee et al. provide a detailed review of the sources, transport routes, and potential toxicological impacts of PFAS. In addition, the article provides an overview of the adverse effects of PFAS on human health [113].

The Center for Disease Control (Centers for Disease Control (CDC, 2024) and Agency for Toxic Substances and Disease Registry (ATSDR,2021) reported that less consistent data are associated with effects on the immune system, disruption of thyroid hormones, increased risk of cancer [114,115].

Lipid Metabolism

The Agency for Toxic Substances and Disease Registry [115] stated that the most consistent health effect of PFAS is increased cholesterol in the adult population. The relationship between lipid levels and PFAS exposure was also presented in the study by Liu et al. [66]. In the study, 575 human serum samples were collected. 11 of 18 PFASs were detected in more than 80% of the samples. The highest serum concentrations were measured for PFOA (11.34 ng.mL^-1) and PFOS (7.64 ng.mL^-1). With increased exposure to PFASs, total TC and LDL concentrations also increased, while HDL and TG concentrations were not affected. PFUnDA and PFTrDA had a greater effect on blood lipid concentrations than other PFASs [66].

In a review article, Ho et al. [116] comprehensively assessed the effects of PFA exposure on LDL, HDL, total cholesterol TC, and triglyceride TG concentrations in human plasma, serum, and whole blood. The study found a positive relationship between PFOA-LDL, PFOA-TC, PFOS-TC, and PFNA-LDL. The relationships between PFAS, especially perfluoroundecanoic acid (PFUnDA), and triglycerides tended to be negative [116].

Gardener et al. [117] investigated the relationship between serum concentrations of PFDA, PFNA, PFOS, PFOA PFHxS and fasting serum concentrations of total cholesterol, triglycerides and insulin. The study included 433 pregnant women whose serum was collected in the third trimester. PFAS were examined in quartiles in relation to serum biomarkers, gestational age at birth and birth weight standardized for gestational age. Results showed that total cholesterol was positively associated with PFDA, PFNA, PFOS and triglycerides, but PFAS were not associated with fasting insulin. PFNA was associated with an increased likelihood of preterm birth (<37 weeks of gestation) [117].

A large Japanese study by Hasegawa et al. [65] investigated the association between 28 PFAS and lipid concentrations in both maternal and cord blood. The analysis included 20.960 pregnant women. Plasma samples were collected before 22 weeks of gestation to determine PFAS concentrations. Serum samples collected before, during, after 22 weeks of gestation, at birth, and from cord blood were used to determine total cholesterol and triglycerides. Using linear regression models, 7 PFAS were quantified in more than 80% of the women out of the 28 PFAS analyzed, namely PFOA, PFNA, PFDA, PFOS, PFUnA, PFHxS, PFTrDA. Of these, 6 had positive associations with maternal total cholesterol before 22 weeks of gestation, but no association with cord blood TC. In the case of triglycerides, a negative relationship was shown between 3 PFAS and maternal blood and a positive relationship between 4 PFAS and TG in cord blood [65].

PFAS exposure may affect lipid metabolism in pregnant women. Elevated plasma triglyceride levels may cause preeclampsia or hypertension. Yang et al. measured 11 PFAS in serum of 436 pregnant women and investigated the association with lipid parameters, namely TC, TG, HDL, LDL, PFOS, PFOA, PFHxS, PFUdA, PFNA, PFDA and PFHpS were detected in serum with a detection rate of more than 70%. The following associations were found positive association of PFHxS with TC, HDL and LDL; negative association of PFUdA with HDL PFDA with LDL; PFOA, PFNA, PFDA and PFUdA with LDL/HDL. The results indicate the possibility of an impact of PFAS exposure on lipid metabolism in pregnant women [118].

In Italian study, there was conducted a cross-sectional analysis of 319 pregnant women aged 14-48 years, who came from areas with high exposure to PFAS through drinking water. Concentrations of PFOA, PFOS and PFHxS, total cholesterol (TC), HDL-C and LDL-C were determined. Elevated TC, LDL-C and HDL-C values ​​increased gradually and may have adverse effects on both the mother and the fetus. Associations between PFAS and lipid markers varied throughout all trimesters of pregnancy. In the first trimester, the relationship between PFOS and TC and between PFHxS and HDL-C was positive and similar to that in non-pregnant women, with a reversal during the third trimester. There was an inverse relationship between PFOA and PFHxS and TC and LDL-C. The results highlight the importance of the correct timing of PFAS measurements during pregnancy [119].

Diabetes Mellitus

Exposure to persistent organic pollutants in the environment may be one of the risk factors for the development of diabetes mellitus, which has a growing prevalence worldwide.

In the study [120], there was investigated the relationship between PFAS exposure and the risk of developing type 2 diabetes mellitus (T2DM). 252 T2DM cases and 252 controls were examined and the results showed that serum PFHxS and PFHpA were significantly positively associated with the risk of developing T2DM at the lower levels [120].

Xu et al. [121] tested the hypothesis that PFAS can cause gestational diabetes mellitus (GDM) by modulating glucose metabolism. The study included 171 women with GDM and 169 controls and determined 15 PFASs including homologues of PFOA and PFOS; precursors of PFSA and PFCA and alternatives of PFOS and PFOA, which were detected in more than 70% of maternal serum samples. The highest concentrations in maternal serum were measured for PFOA 7.43 ng/ml, PFOS 4.23 ng/ml and chlorinated polyfluorinated ether sulfonate in the ratio 6:2 (6:2 Cl-PFESA). Exposure to PFOA, PFNA, PFDA, PFUnDA, PFDoA, PFHxS and PFOS was significantly higher in women with GDM compared to controls. Concentrations of PFOA, PFOS, PFUnDA, PFDoA and 6:2Cl-PFESA were associated with impaired glucose homeostasis in pregnancy and an increased risk of GDM, while an inverse association with GDM was found for PFHxS, 4:2FTS and 6:2FTS [121].

A Korean study [122] presented the relationship between serum PFAS concentrations and the prevalence of prediabetes and prediagnostic diabetes. This is the first study to describe a strong positive association between serum PFAS concentrations and prediabetes and the prediagnostic stage of diabetes. The study included 2709 participants aged ≥ 19 years. Prediagnostic diabetes and prediabetes were determined based on glycated hemoglobin HbA1c values. Serum PFOA, PFNA, PFDeA, PFHxS and PFOS were determined by HPLC combined with tandem mass spectrometry. Concentrations of individual PFAS compounds as well as mixtures were associated with a higher risk of prediabetes. Significant positive associations were found between serum PFOS and PFHxS and increased HbA1c values ​​and a higher risk of prediagnostic diabetes. The results suggest that PFAS exposure may contribute to impaired glucose homeostasis in the prediabetic stage and thus to the development of diabetes [122].

A Chinese study [123] included 495 women, 165 with GDM and 330 controls. GDM was measured between 24-28 weeks of gestation by glucose test. PFOA, PFOS, PFBS, PFDoA, PFUA, PFDA, PFHpS, PFNA,PFHxS, PFDS, PFHpA, PFOSA in serum were determined by UPLC-Q/TOF MS. PFHpA, PFDS and PFOSA were all present in ≤ 80% of samples and were therefore not considered further. PFOS, PFOA, PFBS, PFDoA were measured at significantly higher concentrations in maternal serum in early pregnancy, which may be associated with a higher risk of GDM [123].

Disorders in Women

There are many studies on PFAS exposure and male fertility, and most of them show that PFAS exposure leads to impaired male fertility [124]. The available results obtained from studies focusing on women have been quite inconsistent. Therefore, more systematic reviews and meta-analyses have been conducted to examine the evidence on the effect of PFAS exposure during pregnancy on maternal fertility, in addition to quantitative assessment of maternal PFAS concentrations during pregnancy and the risk of fertility and infertility.

Therefore, Wang et al. [125] conducted a meta-analysis in a review article. A total of 5468 records from 4 databases were searched, and after gradual elimination, 13 articles that fully met the inclusion criteria were left and were used for the meta-analysis. The results showed that exposure to (PFOA) was negatively associated with female fertility and positively associated with the odds ratio for infertility. Exposure to (PFOS) was negatively associated with the odds ratio for fertilization. The obtained effect values ​​for exposure to PFNA), (PFDA) and perfluorohexanesulfonate (PFHxS) did not provide sufficient evidence of an association with female fertility [125].

Preeclampsia is a condition of hypertensive disorder during pregnancy that can be a cause of maternal mortality or can be associated with low birth weight, prenatal death and preterm birth. Tian et al. [126] studied the possible association between preeclampsia and serum PFAS concentrations in 82 women with preeclampsia and 169 healthy women. 15 PFAS were analyzed in maternal serum before delivery. Concentrations of PFOA and 6:2Cl-PFESA were associated with an increased risk of developing preeclampsia, especially in primiparous women expecting a girl. Increased PFOA concentrations were significantly associated with increased blood pressure. In relation to neonatal development, a negative relationship was found between exposure to PFOS, PFNA, PFUnDA and 6:2Cl-PFESA and birth weight. Regression models indicated that significantly higher risks of low birth weight occurred in women with preeclampsia compared to normal pregnancies. This observational study considered not only legacy PFAS but also new alternatives and included a low birth weight parameter to better elucidate the possible effects of PFAS exposure on adverse birth outcomes [126].

Preeclampsia was also addressed in a study by Thompsom et al. [127] which examined the association between hypertensive disorders of pregnancy (preeclampsia and gestational diabetes) and exposure to 4 PFAS: PFOA, PFOS, PFHxS, and PFNA in a sample of 513 African-American mothers. Serum samples were collected from mothers between 8 and 14 weeks of pregnancy: Regression models were used to assess associations. Mean PFOS and PFHxS levels were lower in patients with preeclampsia compared to patients without hypertensive disorders. PFAS concentrations were not associated with gestational hypertension or preeclampsia. In this study, no association was found between serum PFAS concentrations measured in early pregnancy and hypertensive disorders of pregnancy [127].

There is an association between PFAS and reduced birth weight, but it may be complicated by glucose status due to the effects of PFAS on fetal growth and placental transport. Wang 2023 analyzed 1405 mother-child pairs and investigated whether maternal glucose levels were associated with prenatal PFAS exposure and birth weight. Plasma concentrations of PFOA, PFOS, PFNA, PFDA, PFUA, and PFHxS were determined in the first trimester. Plasma glucose was measured at 24-28 weeks of gestation. The results suggest that infants born to mothers with hyperglycemia may be sensitive to PFAS exposure [125].

Zhang et al. investigated whether PFAS exposure is associated with Polycystic ovary syndrome (PCOS). A total of 502 women undergoing reproductive treatment were included in the study and 9 PFAS were measured in serum samples. Higher serum concentrations of PFOS and PFHxS were associated with a higher risk of PCOS. No associations were found for PFOA [128].

The results of this study were consistent with the results of a case-control study by Zhan et al., which included 366 women with PCOS and 577 controls. Interestingly, the concentrations of PFOA were several times higher than in the discussed article (7.2 versus 1.7 ng.mL^-1), while the concentrations of PFOS (3.9 and 3.5 ng.mL^-1) and PFHxS (0.22 and 0.8 ng.mL^-1) were found. The results support the possible adverse effects of PFAS on female reproductive function, but further studies are needed to confirm the findings [129].

The review article by Yi et al. summarizes the impact of PFAS exposure on the development of four reproductive health outcomes in women studied to date, namely polycystic ovary syndrome, endometriosis, primary ovarian insufficiency, and diminished ovarian reserve [130].

Thyroid Hormones

Thyroid hormones play an important role in regulating health, in brain development, depression, and obesity. Thyroid hormone deficiency during pregnancy can cause reduced IQ or neurological damage in children. In addition, it can cause intellectual disability or growth retardation in adolescents. PFAS are potential thyroid disruptor [131,132].

He et al. [133] investigated the relationship between exposure to 30 PFAS and thyroid function in 194 Chinese children aged 3-17 years using multiple statistical models. Free triiodothyronine, free thyroxine (FT4), and thyrotropin were tested as indicators of thyroid function, and subclinical hypothyroidism was diagnosed. PFAS and their alternatives were associated with changes in thyroid hormone levels and subclinical hypothyroidism. Higher concentrations of perfluorohexanoic acid (PFHpA) caused a decrease in FT4 levels and an increased likelihood of subclinical hypothyroidism. The measured PFOA concentration (median) of 23.22 μg.L^-1 was comparable to some previous studies. For example, in America, a concentration of 29.3 μg.L^-1 was found in children aged 1-17 years [134] in adults 23.1 μg.L^-1 [135], in pregnant women 23.22 μg.L^-1) [136]. The PFOS concentrations of 4.31 μg.L^-1 and PFNA 1.79 μg.L^-1 (median) in this study in children were similar to the PFOS and PFNA concentrations in adults (5.17 μg.L^-1 and 1.00 μg.L^-1) [137] and adolescents (PFOS 7.78 μg.L^-1and PFNA 1.01 μg.L^-1) [138]. Higher serum PFOA concentrations cause decreased FT4 levels and increased TSH levels. A cohort study in Spain obtained opposite results in children aged 15–17 years [139].

In a US study [140] higher serum PFOA levels were associated with lower TSH levels in adolescent females, which are different results from the discussed study in children. These different results may be related to variations in the regions and populations of each study.

In the study by Lebeaux et al. the authors investigated the association between maternal serum concentrations of PFAS (PFOA, PFOS, PFNA, and PFHxS) during pregnancy with levels of thyroid-stimulating hormone (TSH), total thyroxine (TT4), total triiodothyronine (TT3), free thyroxine (FT4), and free triiodothyronine (FT3) measured in maternal and cord serum. The study involved 468 pregnant women and their children. The results show that there is no strong association between maternal serum PFAS concentrations measured in the second trimester and maternal and cord serum thyroid hormones [132]. A cross-sectional study by Xie et al. examined thyroid function parameters and serum PFAS concentrations in adolescents. Concentrations of 18 PFAS were measured, including 11 PFCAs, 4 PFSAs, and 3 novel PFASs (4:2, 6:2, and 8:2Cl-PFESA). The study included 836 adolescents aged 11-15 years who came from a high-PFCA-exposed area near a fluorochemical industrial plant. Elevated FT3 and decreased FT4 levels were found. Using logistic and linear regression, a significant negative correlation was found between PFOA and FT4 and a positive correlation between PFHxS and FT3, with both PFOA and PFHxS being assessed as risk factors [141].

Wang et al. conducted a study to investigate the effect of PFAS exposure on serum metabolome and its association with thyroid cancer. The study included 746 thyroid cancer cases and 746 controls. LC-MS/MS was used to determine the concentrations of PFDoA, PFUnDA, PFDA, PFNA, PFOA, PFOS, PFHxS, PFHxA, PFHpA, PFBS, PFPeA. Exposure to PFHxA and PFDoA was associated with an increased risk of thyroid cancer, while PFOA and PFHxS were associated with a decreased risk [142].

Liver Disorders

General Liver Toxicity

PFAS are known to accumulate in various organs. Many studies have shown an association between PFAS exposure and liver toxicity, as well as the impact of PFAS on liver homeostasis, lipid and bile acid metabolism, and hepatocarcinogenesis. In a review article, Maerten et al discuss the role of PFAS in liver toxicity, the mechanisms underlying the hepatotoxic effects of PFAS, and fill the knowledge gap for a better assessment of PFAS risks [143].

In humans, the highest concentrations of PFAS were measured in the liver, kidneys, and lungs. PFOSD predominates in the liver, while PFBA predominates in the lungs and kidneys. Many studies [144,145,146,147] reported that exposure to PFAS (PFOS, PFOA, PFNA, PFHxS, fluorohexanoic acid, perfluoro-3,5,7,9,11-pentaoxadocanoic acid and 6:2 chlorinated polyfluorinated ether sulfonate (6:2 Cl-PFESA) can lead to liver damage in humans. Markers of liver damage include alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyltransferase (GGT), alkaline phosphatase and serum bilirubin, the increased values ​​of which cause an increase in inflammatory markers. PFOS, PFOA, PFBA increase ALT activity [148].

Dyslipidemia, Steatosis and Steatohepatitis

This article summarizes studies that have evaluated the relationship between serum PFAS and steatosis and steatohepatitis as a consequence of dyslipidemia. PFNA correlates with LDL, PFOS with total cholesterol, and PFOA with both LDL and TC [149,150,151,152].

PFOS, PFOA, and PFHxS have been associated with an increased risk of developing steatohepatitis [153,154]. PFAS exposure can lead to hepatic steatosis, which is manifested by increased inflammatory markers and biomarkers of liver injury.

Hepatocarcinogenesis

The International Agency for Research on Cancer considers both PFOS and PFOA to be probable human carcinogens in humans at high exposure and in humans with testicular and kidney tumors. The association of PFOS and PFOA with hepatocarcinogenesis has been demonstrated in laboratory animals, but not in humans. In conclusion, it can be stated that exposure to PFAS can cause: an increase in serum markers of liver damage; an increase in cholesterol and hepatic lipid accumulation in humans; interference with bile acid metabolism by affecting hepatic transporters [115].

The study by Dai et al. investigated the correlations between PFAS concentrations and biomarkers of liver function. The study included 227 patients, of whom 197 had hepatocellular carcinoma and 30 suffered from severe liver disorders such as cirrhosis and hepatitis. 21 PFAS were determined, with the highest concentrations in serum, then in blood and urine. The concentrations were higher in the group of patients with HCC. Higher concentrations of PFAS are achieved in men compared to women. The highest concentrations in blood and serum were PFOA, PFOS, PFBS and PFHxS. The highest concentration was determined in urine for PFBA and PFPeA. The concentrations of PFBS, PFHpS and PFHxPA correlated with increased values ​​of alkaline phosphatase (ALP), aspartate aminotransferase (AST) and alpha-fetoprotein (AFP) (p < 0.05). The results provide a good basis for further study on the possible hepatotoxicity of PFAS [155].

Hypertension

PFAS exposure is associated with adverse health effects, such as dyslipidemia, hypertension, and hypertensive disorders of pregnancy (HDP), specifically preeclampsia and gestational hypertension. The link between PFAS and HDP is discussed in a review article by Erinc et al. [156].

Gestational hypertension (GH) is new-onset hypertension occuring after 20 weeks of pregnancy. A Chinese study evaluated the relationship between 13 PFAS and gestational hypertension and blood pressure (BP) during pregnancy. 826 pregnant women were included, with blood samples collected at 16 weeks of gestation. The highest concentrations of 11.99, 8.81 and 5.43 ng.mL^-1 were measured for PFOA, PFOS and PFHxS respectively. 5.57% of women developed GH. A lower probability of GH was demonstrated for PFOS, PFDA, PFUdA and PFDoA. Associations were found between PFAS and lower systolic and diastolic blood pressure values ​​in the third trimester. PFDA and PFUdA affected systolic blood pressure only in pregnant women whose fetus was female [157].

The study [158] investigated the relationship between PFAS exposure and hypertensive disorders of pregnancy (i.e. preeclampsia, gestational hypertension), which can cause maternal and neonatal morbidity and mortality. The study included 1558 pregnant women and measured PFHxS, PFOS, PFOA, PFNA, PFDA, EtFOSAA, MeFOSAA, PFOSA in plasma samples. The parameters examined were preeclampsia, gestational hypertension, and trimester-dependent diastolic and systolic blood pressure. An association was found between PFOS, PFOA, and PFHxS concentrations and gestational hypertension, but not with preeclampsia. Analysis of the PFAS mixture indicated a positive association with gestational hypertension, while the associations for PFOA and PFHxS remained. Furthermore, a positive association was observed between high concentrations of PFOS and PFOA and diastolic blood pressure in the 2nd and 3rd trimesters, as well as for the PFAS mixture in both trimesters. For systolic blood pressure, an association was demonstrated only for PFOA in the second and third trimesters. The results indicate a possible impact of PFAS exposure on blood pressure regulation during pregnancy [158].

Cancer

The International Agency for Research on Cancer classified PFOA as a Group 1 carcinogen and PFOS as a Group 2B carcinogen in 2023 [159]. The United States Environmental Protection Agency (EPA) has set a recommended health level of 70 ng.L^-1 for lifetime exposure to PFOS and PFOA [160]. Given the increasing number of epidemiological studies on PFAS and their adverse effects on humans, it has become imperative to address the carcinogenic mechanisms. The aim of the review by Zheng et al. was to update the evidence on the association between PFAS exposure and cancer. It presents a review of the literature published between 2019-2024 on the potential relationship between PFAS (PFOS, PFOA, PFHxS, PFHxA, PFNA) and various types of cancer, such as breast, testicular, prostate, colorectal, kidney, liver, lung, thyroid, leukemia, melanoma. In addition, the article also addresses potential carcinogenic mechanisms such as endocrine disruption, lipid metabolism, epigenetic alteration, oxidative stress, immunosuppression, and chronic inflammation [142,155,161,162,163,164,165,166,167,168].

Obesity

Chen et al. 2024 investigated the effects of PFAS exposure (PFOA, PFOS, PFUdA, PFNA, PFHxS, PFDeA, and 2- (N-methyl-perfluorooctane sulfonamide) acetic acid) on obesity. 11,090 individuals were analyzed and linear and logistic regression models were used to evaluate the effects. The role of inflammatory markers (neutrophils, lymphocytes, and alkaline phosphatase) and oxidative stress markers (gamma-glutamyltransferase, total bilirubin, and uric acid) was also examined. Lymphocytes, alkaline phosphatase, and total bilirubin were significantly associated with both obesity and type 2 diabetes mellitus. PFAS exposure was positively associated with the development of obesity and T2DM and mediated by inflammation and oxidative stress. PFNA was positively associated with obesity and T2DM, while PFOA was negatively associated with T2DM. PFDeA was positively associated with hypertension but negatively with obesity and T2DM [169].

Averina et al. investigated associations between PFAS and dyslipidemia, hypertension and obesity in adolescents. A cross-sectional study was conducted on 940 adolescents with a mean age of 16.4 years who had either hypertension, obesity or dyslipidemia. Serum concentrations of PFHxS, PFOA, PFOS, PFNA, PFDA, PFUnDA were measured by UHPLC-MS/MS. The results showed a positive association of PFOS, PFNA, PFDA and PFUnDA with apolipoprotein B, LDL and total cholesterol. Concentrations of PFOS, PFOA were positively associated with the risk of hypertension, and PFHxS and PFHpS with obesity [170].

In a study by Geiger et al., the association between PFOA and PFOS and BMI and waist circumference was investigated in a representative sample of 2473 American children. Elevated concentrations of PFOA may indicate an association with overweight and obesity in children. This topic requires further research, which will include, for example, abdominal adiposity, fat distribution, etc. [171].

Autism

Autism spectrum disorder (ASD) is a complex of neurodevelopmental disorders characterized by deficits in social interaction and communication, as well as the presence of restricted, stereotyped interests and repetitive behaviors [172]. Although there is a possibility of a potential association between PFAS and the development of ASD, large studies are needed to confirm this. Six case-control studies and one cohort study have evaluated the possible association between PFAS and the development of ASD. The results have been highly inconsistent. The studies by Liew et al. (220 with ASD and 550 controls) and Lyall et al. examined prenatal exposure to PFAS and the association with ASD (533 children with ASD and 433 controls) and found no consistent evidence that maternal plasma concentrations of PFAS were associated with an increased risk of ASD in a Danish population [173,174]. Long et al. measured PFAS levels in amniotic fluid of 75 children with ASD and 135 controls and found an inverse association between PFAS and risk of ASD, which may be related to the weak estrogenic activities of PFAS [175]. Skogheim et al. studied 400 children with ASD and 980 controls and confirmed an association between PFOA and ASD in boys [176].

Oh et al. investigated the association between prenatal exposure to PFAS and an increased risk of autism spectrum disorders. The study included 173 mother-child pairs, with children aged 3 years confirmed to have autism spectrum disorders. Nine PFAS were measured in maternal serum, namely PFOA, PFOS, PFHxS, PFNA, PFDA, PFUnDA, PFDoDA, MeFOSAA, EtFOSAA. Positive associations were found for PFOA and PFNA and an increased risk of autism spectrum disorders, while PFHxS showed negative associations. Given that exposure to individual and combined PFASs had different effects on the risk of ASD depending on the advanced age of the mother at delivery, further study with a larger sample size is needed [177].

Respiratory Diseases

The term lung function includes respiratory, metabolic, defense and other functions of the lungs. Changes in lung function indicate the risk and severity of clinical symptoms and their impact on quality of life. Children, as a risk population, are more likely to be exposed to PFAS due to higher respiratory rates, interactions with contaminated surfaces (e.g. crawling on the floor), etc. As a result of environmental exposure to PFAS in children, respiratory burden arises, which raises concerns about the rapid development of pulmonary systems [178].

It is very important to determine the relationship between PFAS exposure and respiratory diseases, such as asthma. Meta-analysis processed bibliographic data from various studies that were dedicated to diagnosing asthma in children under 17 years of age, revealing a connection between PFOA exposure and a higher risk of developing asthma as well as between PFOS and lung function disorders [179].

Exposure to PFAS in pregnant women can cause respiratory problems in children after birth. Among the main factors that affect lung development in the prenatal period are, for example, premature birth, placental insufficiency, maternal tobacco smoking, etc.

Kung et al. [180] in their study investigated the relationship between intrauterine and postnatal PFAS exposure and lung function in children. The study used 165 umbilical cord blood plasma and serum samples from 8-year-old children. The average concentrations of PFOA, PFOS, PFNA and PFUA were determined by HPLC-MS/MS. The monitored lung functions were FEV1 (forced expiratory volume in one second), FVC (forced vital capacity), PEF (peak expiratory flow) and FEV1/FVC. The average concentrations of selected analytes were higher in umbilical cord blood than in serum. PFAS were not shown to significantly affect lung function in children. However, they found that PFOS concentrations were significantly inversely correlated with lung function in children with lower birth weight and suffering from allergic rhinitis [180].

Kidney Diseases

Chronic kidney disease has a high incidence and mortality rate, especially in patients with diabetes and hypertension. PFAS may be hazardous to kidney function. Liang 2023 et al. investigated the relationship between PFOAS, PFOS and Cl-PFESA, heavy metals (cadmium, lead, arsenic) to glomerular filtration rate and chronic kidney disease. PFOA and heavy metals were positively correlated with chronic kidney disease. PFOA, PFOS, CI-PFESA and arsenic were negatively associated with glomerular filtration rate. The study provided epidemiological evidence that Cl-PFESA alone and together with heavy metals may contribute to kidney damage [181].

A nationwide cross-sectional study was conducted in 13,979 adults in the US to examine the association of serum PFAS with the risk of uric acid and hyperuricemia. Even at low concentrations, the dominant PFOA may have caused an increase in uric acid and an increased risk of hyperuricemia and contributed to a decline in kidney function [182].

Cardiovascular Diseases

PFAS can play a role in the development of cardiovascular diseases (myocardial infarction, ischemic stroke, heart failure). A Swedish cohort study examined the association between PFAS exposure and cardiovascular disease in 2278 participants aged 45-75 years. PFOA, PFOS, and PFHxS were measured in plasma. A second independent study measured PFHxS, PFOA, linear isomer of PFOS, PFNA, PFDA, and PFUnDA in plasma in 1016 participants aged 70 years. Neither study found an increased risk of cardiovascular disease at slightly elevated levels of PFAS [183].

Potential associations between PFAS and values ​​of pre-selected proteomic biomarkers in plasma were observed in a Swedish study by Dunder et al. PFOA, PFOS and PFHxS were measured by untargeted metabolomics and 249 proteomic biomarkers in plasma in 2342 participants were measured by proximity extension assay. Epidermal growth factor receptor (EGFR) and paraoxonase type 3 (PON3) levels were positively associated with all three PFAS, while resistin (RETN) and urokinase surface plasminogen activator receptor (uPAR) showed inverse associations with all three PFAS [184].

Bone Metabolism

PFAS exposure is associated with a number of disorders and may also act as a toxic agent for the skeleton. PFAS interfere with many hormones that affect bone metabolism. An association has been found between PFAS exposure and lower bone mineral content and bone mineral density in children and adolescents [185]. A study in Ohio included 197 adolescents aged 12 years, from whom serum was collected. Concentrations of PFOA, PFOS and PFNA were determined using the SPE-LC-MS/MS method. Higher concentrations of PFAS caused a decrease in the BMC (bone mineral content) z-score. The adverse effects of PFAS can be mitigated by higher calcium intake, increased physical activity and a better quality diet [185].

Neurodevelopment in Child

PFAS can cross the placenta, and prenatal exposure to PFAS has been associated with neurodevelopmental disorders in children, increased risk of neuropsychological problems, and abnormal behavior in adulthood. The effects of PFAS on infant neurodevelopment were investigated in a study by Zhou et al., which included 1285 mother-infant pairs. PFOS, PFOA, PFHxS, and 62Cl-PFESA were confirmed in more than 90% of samples. Each increase in PFAS concentration resulted in poorer communication domain scores. The decrease in communication domain scores in 6-month-old infants was mainly attributed to the increase in PFOS concentration. The findings indicated that PFAS may adversely affect neurodevelopment in children [186].

An association between prenatal exposure to legacy PFAS and children’s intelligence and executive functioning was observed in a Canadian cohort study of mothers and infants. An association with child gender was also examined. Plasma concentrations of PFOA, PFOS and PFHxS were measured in the first trimester, and 522 children were assessed for full-scale, performance and verbal IQ using the Wechsler Preschool and Primary Intelligence Scale. Results indicated that each two-fold increase in prenatal exposure to PFOA, PFOS and PFHxS was inversely associated with performance IQ, but only in males. No significant association was found in females [187].

Cerebral Palsy

It is known that PFAS can be transported to the fetus across the placenta. Thyroid hormone levels may be disrupted during pregnancy. Insufficient hormone levels during a critical stage of brain development can lead to neurodevelopmental disorders such as cerebral palsy (CP), which is one of the most common physical and motor disabilities in childhood. The author Vilhelmsson et al. [188] investigated the association between prenatal PFAS exposure and the risk of CP. The study included 322 CP cases, 258 premature infants, and 343 controls. Serum samples collected at 10-14 weeks of gestation were obtained from a biobank, and concentrations of PFOA, PFOS, PFNA, and PFHxS were determined. No associations were observed between prenatal exposure to PFAS and the risk of CP, except in premature infants, but the results were not entirely consistent and therefore require further study.

The Swedish study discussed was preceded by only one study that investigated whether prenatal exposure to PFAS increases the risk of congenital cerebral palsy. This Danish study followed 83,389 newborns and mothers, identified 156 CP cases and randomly selected 550 controls. 6 PFAS were identified in more than 90% of samples and 16 PFAS in maternal plasma collected in early and mid-pregnancy. Higher concentrations of mainly PFOS and PFOA in maternal plasma increased the risk of CP in boys. In girls, no association was found between PFAS and CP [189].

7. Conclusions

PFAS are characterized by high toxicity, stability and slow degradation, which causes long persistence in various components of the environment, from where they accumulate in the human body through plants and animals and cause adverse health effects. The toxicological properties of PFAS in humans are not sufficiently studied due to the large number of compounds. Our review is based on scientific studies to clarify the presence of per- and polyfluoroalkyl substances in various, conventional and unconventional human matrices, such as hair, nails, urine, placenta and breast milk, but especially to health problems caused by PFAS exposure. Exposure to these toxicants has caused a variety of adverse health consequences, therefore, various biomonitoring approaches in humans are necessary. Exposure data, together with modifiable environmental risk factors, are crucial for risk assessment and facilitate the development of new strategies to prevent and mitigate the adverse effects of PFAS.