Nabumetone and flufenamic acid pose a serious risk to aquatic plants: A study with Chlamydomonas reinhardtii as a model organism.

The aquatic environment is constantly under threat due to the release of numerous pollutants. Among them, pharmaceuticals constitute a huge and diverse group. Non-steroidal anti-inflammatory drugs (NSAIDs) are increasingly found in water bodies, but knowledge about their potential toxicity is still low. In particular, there is a lack of information about their influences on aquatic plants and algae. We estimated the susceptibility of the microalgae Chlamydomonas reinhardtii to nabumetone (NBT) and flufenamic acid (FFA), focusing on photosynthesis. Due to the differences in the structures of these compounds, it was assumed that these drugs would have different toxicities to the tested green algae. The hypothesis was confirmed by determining the effective concentration values, the intensity of photosynthesis, the intensity of dark respiration, the contents of photosynthetic pigments, the fluorescence of chlorophyll a in vivo (OJIP test), and cell ultrastructure analysis. Assessment of the toxicity of the NSAIDs was extended by the calculation of an integrated biomarker response index (IBR), which is a valuable tool in ecotoxicological studies. The obtained results indicate an over six times higher toxicity of NBT compared to FFA. After analysis of the chlorophyll a fluorescence in vivo, it was found that NBT inhibited electron transport beyond the PS II. FFA, unlike NBT, lowered the intensity of photosynthesis, probably transforming some reaction centers into "silent centers", which dissipate energy as heat. The IBR estimated based on photosynthetic parameters suggests that the toxic effect of FFA results mainly from photosynthesis disruption, whereas NBT significantly affects other cellular processes. No significant alteration in the ultrastructure of treated cells could be seen, except for changes in starch grain number and autophagic vacuoles that appeared in FFA-treated cells. To the best of our knowledge, this is the first work reporting the toxic effects of NBT and FFA on unicellular green algae.

Evaluation of greenness and analytical performances of separative methods for chiral separation of novel lactam-based P2RX7-antagonists.

In this work, a preparative supercritical fluid chromatography (SFC) method was first developed to separate a series of chiral compounds evaluated as lactam-based P2RX7 antagonists. Subsequently, high-performance liquid chromatography, SFC, and capillary electrophoresis (CE) were comparatively investigated as QC tools to determine the enantiomeric purity of the separated isomers, including analytical performance and greenness. The screening of the best conditions was carried out in liquid and SFC on the nine derivatives and the amylose tris(3,5-dimethylphenylcarbamate)-based chiral stationary phase was found to be highly efficient. The same screening was carried out in CE and very different conditions, either in acidic or basic background electrolyte and different cyclodextrins used as chiral selectors, allowed the separation of six of the nine derivatives. 1-((3,4-Dichlorophenyl)carbamoyl)-5-oxopyrrolidine-2-carboxylic acid (compound 1) was chosen as a probe, and its semi-preparative separation by SFC and enantiomeric verification using the three techniques are presented. Its limit of detection and limit of quantification are calculated for each method. Finally, the greenness of each quality control method was evaluated.

Photoprotection and Antiaging Activity of Extracts from Honeybush (Cyclopia sp.)-In Vitro Wound Healing and Inhibition of the Skin Extracellular Matrix Enzymes: Tyrosinase, Collagenase, Elastase and Hyaluronidase.

Cyclopia sp. (honeybush) is an African shrub known as a rich source of polyphenols. The biological effects of fermented honeybush extracts were investigated. The influence of honeybush extracts on extracellular matrix (ECM) enzymes responsible for the skin malfunction and aging process-collagenase, elastase, tyrosinase and hyaluronidase-was analysed. The research also included assessment of the in vitro photoprotection efficiency of honeybush extracts and their contribution to the wound healing process. Antioxidant properties of the prepared extracts were evaluated, and quantification of the main compounds in the extracts was achieved. The research showed that the analysed extracts had a significant ability to inhibit collagenase, tyrosinase and hyaluronidase and a weak influence on elastase activity. Tyrosinase was inhibited effectively by honeybush acetone (IC50 26.18 ± 1.45 µg/mL), ethanol (IC50 45.99 ± 0.76 µg/mL) and water (IC50 67.42 ± 1.75 µg/mL) extracts. Significant hyaluronidase inhibition was observed for ethanol, acetone and water extracts (IC50 were 10.99 ± 1.56, 13.21 ± 0.39 and 14.62 ± 0.21µg/mL, respectively). Collagenase activity was inhibited effectively by honeybush acetone extract (IC50 42.5 ± 1.05 µg/mL). The wound healing properties of the honeybush extracts, estimated in vitro in human keratinocytes (HaCaTs), were indicated for water and ethanol extracts. In vitro sun protection factor (SPF in vitro) showed medium photoprotection potential for all the honeybush extracts. The quantity of polyphenolic compounds was estimated with the use of high-performance liquid chromatography equipped with diode-array detection (HPLC-DAD), indicating the highest mangiferin contents in ethanol, acetone and n-butanol extracts, while in the water extract hesperidin was the dominant compound. The antioxidant properties of the honeybush extracts were estimated with FRAP (2,4,6-Tris(2-pyridyl)-s-triazine) and DPPH (2,2-diphenyl-1-picrylhydrazyl) tests, indicating their strong antioxidant activity, similar to ascorbic acid for the acetone extract in both tests. The wound healing abilities, estimation of SPF in vitro and the direct influence on selected enzymes (elastase, tyrosinase, collagenase and hyaluronidase) of the tested honeybush extracts were analysed for the first time, indicating a high potential of these well-known herbal tea for antiaging, anti-inflammation, regeneration and protection of the skin.

Isolation of Citrus lemon extracellular vesicles: Development and process control using capillary electrophoresis.

A new and scalable method for the isolation of extracellular vesicles (EV) from Citrus lemon juice samples was developed. The methodology included preliminary preconcentration of the sample using ultrafiltration (UF) followed by size-exclusion chromatography (SEC) purification and final preconcentration of the eluates. Transmission electron microscopy and proteomic analysis showed that isolates contained exosome-like vesicles, exocyst-positive organelle (EXPO), and microvesicles. The efficiency of certain isolation steps was evaluated with total protein content assay (bicinchoninic acid assay, BCA), nanoparticles tracking analysis (NTA), and capillary electrophoresis (CE). A good correlation between CE, BCA, and NTA results was shown. The application of CE enabled the detection of soluble contaminants, macromolecular aggregates, and vesicles' heterogeneity. The fluorescent staining of encapsulated nucleic acids was proposed for the identity confirmation of EV detected in CE. The study demonstrates the CE as a comprehensive tool for monitoring of the EV isolation process.

Protein overproduction alters exosome secretion in Chinese hamster ovary cells.

Despite the abundance of available cell lines, nearly 70% of all recombinant therapeutic proteins today are produced in Chinese hamster ovary (CHO) cells. The impact of protein overproduction on the secretion of exosomes by CHO cells has been investigated here. Increased secretion of extracellular vesicles (EVs) by protein overexpressing CHO cells was demonstrated with protein content assay, nanoparticle tracking analysis, and capillary electrophoresis. Our results revealed that a protein overproduction might induce EVs secretion, which might be accompanied by the sequestration and loading of overexpressed proteins into the exosomes. These findings are of vital importance for the manufacturing of therapeutics in CHO expression systems due to the risk of product loss during downstream processing of culture medium as well as the application of exosomes as nanocarriers of therapeutic proteins. The study indicates also the importance of culturing process control.

Green alga Chlamydomonas reinhardtii can effectively remove diclofenac from the water environment - A new perspective on biotransformation.

The use of unicellular algae to remove xenobiotics (including drugs) from wastewaters is one of the rapidly developing areas of environmental protection. Numerous data indicate that for efficient phycoremediation three processes are important, i.e. biosorption, bioaccumulation, and biotransformation. Although biosorption and bioaccumulation do not raise any serious doubts, biotransformation is more problematic since its products can be potentially more toxic than the parent compounds posing a threat to organisms living in a given environment, including organisms that made this transformation. Thus, two questions need to be answered before the proper algae strain is chosen for phycoremediation, namely what metabolites are produced during biotransformation, and how resistant is the analyzed strain to a mixture of parent compound and metabolites that appear over the course of culture? In this work, we evaluated the remediation potential of the model green alga Chlamydomonas reinhardtii in relation to non-steroidal anti-inflammatory drugs (NSAIDs), as exemplified by diclofenac. To achieve this, we analysed the susceptibility of C. reinhardtii to diclofenac as well as its capability to biosorption, bioaccumulation, and biotransformation of the drug. We have found that even at a relatively high concentration of diclofenac the algae maintained their vitality and were able to remove (37.7%) DCF from the environment. A wide range of phase I and II metabolites of diclofenac (38 transformation products) was discovered, with many of them characteristic rather for animal and bacterial biochemical pathways than for plant metabolism. Due to such a large number of detected products, 18 of which were not previously reported, the proposed scheme of diclofenac transformation by C. reinhardtii not only significantly contributes to broadening the knowledge in this field, but also allows to suggest possible pathways of degradation of xenobiotics with a similar structure. It is worth pointing out that a decrease in the level of diclofenac in the media observed in this study cannot be fully explained by biotransformation (8.4%). The mass balance analysis indicates that other processes (total 22%), such as biosorption, a non-extractable residue formation, or complete decomposition in metabolic cycles can be involved in the diclofenac disappearance, and those findings open the prospects of further research.

Quantification and stability assessment of urinary phenolic and acidic biomarkers of non-persistent chemicals using the SPE-GC/MS/MS method.

Nowadays, people are exposed to numerous man-made chemicals, many of which are ubiquitously present in our daily lives, and some of which can be hazardous to human health. Human biomonitoring plays an important role in exposure assessment, but complex exposure evaluation requires suitable tools. Therefore, routine analytical methods are needed to determine several biomarkers simultaneously. The aim of this study was to develop an analytical method for quantification and stability testing of 26 phenolic and acidic biomarkers of selected environmental pollutants (e.g., bisphenols, parabens, pesticide metabolites) in human urine. For this purpose, a solid-phase extraction coupled with gas chromatography and tandem mass spectrometry (SPE-GC/MS/MS) method was developed and validated. After enzymatic hydrolysis, urine samples were extracted using Bond Elut Plexa sorbent, and prior to GC, the analytes were derivatized with N-trimethylsilyl-N-methyl trifluoroacetamide (MSTFA). Matrix-matched calibration curves were linear in the range of 0.1-1000 ng mL-1 with R > 0.985. Satisfactory accuracy (78-118%), precision (< 17%), and limits of quantification (0.1-0.5 ng mL-1) were obtained for 22 biomarkers. The stability of the biomarkers in urine was assayed under different temperature and time conditions that included freezing and thawing cycles. All tested biomarkers were stable at room temperature for 24 h, at 4 °C for 7 days, and at -20 °C for 18 months. The total concentration of 1-naphthol decreased by 25% after the first freeze-thaw cycle. The method was successfully used for the quantification of target biomarkers in 38 urine samples.

Development of the thin film solid phase microextraction (TF-SPME) method for metabolomics profiling of steroidal hormones from urine samples using LC-QTOF/MS.

In the present study, the development and optimization of a thin film solid phase microextraction method (TF-SPME) was conducted for metabolomics profiling of eight steroid compounds (androsterone, dihydrotestosterone, dihydroepiandrosterone, estradiol, hydroxyprogesterone, pregnenolone, progesterone and testosterone) from urine samples. For optimization of extraction method, two extraction sorbents (PAN-C18 and PS-DVB) were used as they are known to be effective for isolation of low-polarity analytes. The stages of sample extraction and analyte desorption were considered as the most crucial steps in the process. Regarding the selection of the most suitable desorption solution, six different mixtures were analyzed. As a result, the mixture of ACN: MeOH (1:1, v/v) was chosen in terms of the highest analytes' abundances that were achieved using the chosen solvent. Besides other factors were examined such as the volume of desorption solvent and the time of both extraction and desorption processes. The analytical determination was carried out using the ultra-high performance liquid chromatography coupled with high resolution tandem mass spectrometry detection in electrospray ionization and positive polarity in a scan mode (UHPLC-ESI-QTOF/MS). The developed and optimized TF-SPME method was validated in terms of such parameters as extraction efficiency, recovery as well as matrix effect. As a result, the extraction efficiency and recovery were in a range from 79.3% to 99.2% and from 88.8% to 111.8%, respectively. Matrix effect, calculated as coefficient of variation was less than 15% and was in a range from 1.4% to 11.1%. The values of both validation parameters (recovery and matrix effect) were acceptable in terms of EMA criteria. The proposed TF-SPME method was used successfully for isolation of steroids hormones from pooled urine samples before and after enzymatic hydrolysis of analytes.

Urinary concentration of selected nonpersistent endocrine disrupting chemicals-reproductive outcomes among women from a fertility clinic.

Parabens and benzophenones are compounds widely used in cosmetics and personal care products. Although human exposure is widespread there is a limited number of epidemiological studies assessing the relationship between exposure to these chemicals and female reproductive health. The aim of the study is to explore the relationship between paraben and benzophenone concentrations and reproductive outcomes among women attending a fertility center. This prospective cohort included 450 women undergoing in vitro treatment (IVF) at fertility clinic in Poland. The validated gas chromatography ion-tap mass spectrometry to assess concentrations of parabens in urine (methyl (MP), ethyl (EP), propyl (PP), butyl paraben (BP)) and benzophenone-3 (BP-3) was used. To explore the relationship between concentrations of examined chemicals and reproductive outcomes (methaphase II (MII) oocyte yield, total oocyte yield, implantation rate, fertilization rate, clinical pregnancy, live births), multivariable generalized linear mixed model was used for the analysis. Increased exposure to butyl paraben was associated with a significant decrease in MII oocyte count (p = 0.007) when exposure to BP was treated as the continuous variable. Additionally, the exposure to BP in the highest quartile of exposure also decreases MII oocyte count (p = 0.02) compared to the lowest quartile. Urinary concentrations of BP were not related to total oocyte count, fertilization and implantation rate, clinical pregnancy, and live birth when the exposure variable was continuous variable or in the quartiles of exposure. Exposure to MP, EP, PP, the sum of examined parabens, and benzophenone-3 were not related to any of the examined reproductive outcomes. Exposure to butyl paraben was associated with a decrease in MII oocyte count among women attending fertility clinic rinsing concerns that exposure may have a potential adverse impact on embryological outcomes. The results emphasize the importance to reduce chemicals in the environment in order to minimize exposure. As this is the first study showing such an association, further research is needed to confirm these novel results in other populations.

Reviewing the variability in urinary concentrations of non-persistent organic chemicals: evaluation across classes, sampling strategies and dilution corrections.

Various biomonitoring studies have been carried out to investigate the exposure of populations by measuring non-persistent organic chemicals in urine. To accurately assess the exposure, study designs should be carefully developed to maximise reproducibility and achieve good characterization of the temporal variability. To test these parameters, the intraclass correlation coefficients (ICCs) are calculated from repeated measurements and range from poor (<0.4) to excellent (≥0.75). Several studies have reported ICCs based on diverse study designs, but an overview, including recommendations for future studies, was lacking. Therefore, this review aimed to collect studies describing ICCs of non-persistent organic chemicals, discuss variations due to study design and formulate recommendations for future studies. More than 60 studies were selected, considering various chemical classes: bisphenols, pyrethroids, parabens, phthalates, alternative plasticizers and phosphate flame retardants. The variation in ICCs for an individual chemical was high (e.g. ICC of propyl paraben = 0.28-0.91), showing the large impact of the study design and of the specific exposure sources. The highest ICCs were reported for parabens (median = 0.52), while lowest ICCs were for 3-phenoxybenzoic acid (median = 0.08) and bisphenol A (median = 0.20). Overall, chemicals that had an exposure source with high variation, such as the diet, showed lower ICCs than those with more stable exposure sources, such as indoor materials. Urine correction by specific gravity had an overall positive effect on reducing the variability of ICCs. However, this effect was mostly seen in the adult population, while specific compounds showed less variation with creatinine correction. Single samples might not accurately capture the exposure to most non-persistent organic chemicals, especially when small populations are sampled. Future studies that examine compounds with low ICCs should take adequate measures to improve accuracy, such as correcting dilution with specific gravity or collecting multiple samples for one participant.

Comparative study of the percutaneous permeation and bioaccumulation of a cyclic siloxane using frozen-thawed and nonfrozen ex vivo human skin.

Literature shows contradictory information regarding the effect of freezing the excise skin ex vivo on the diffusion of substances into the skin. Few studies indicate that storing the human or animal skin in a frozen state decreases the barrier properties after thawing. Therefore, to understand the properties of frozen skin, we evaluated the effect of storage of ex vivo human skin (2 weeks at -20 °C) on the penetration of stratum corneum and permeation into deeper skin layers (epidermis, and dermis) as well as to the receptor fluid by octamethylcyclotetrasiloxane (D4) a representative test compound of cyclic siloxanes. The main research were preceded by checking the integrity of nonfrozen ex vivo human skin in comparison to the frozen-thawed one by using the Electrical Resistance technique (ER) and the fluorescence microscopy. Samples collected in the skin absorption experiment were analyzed by gas chromatography equipped with a flame ionization detector (GC-FID). The results of this study demonstrated that freezing of excised ex vivo human skin at -20 °C for up to 14 days does not alter the permeability of D4 in a statistically significant manner. Thus, our results confirmed the validity of using skin storage conditions for testing the penetration and permeation of xenobiotics recommended by the OECD, EMA, and WHO guidelines.

Synthetic Pyrethroids Exposure and Embryological Outcomes: A Cohort Study in Women from Fertility Clinic.

Pyrethroids exposure has been associated with adverse reproductive outcome. However, there is no study that explores the effect of environmental exposure and embryological outcomes. This question was addressed in a prospective cohort of couples undergoing fertility treatment. The study aims to assess the association between urinary metabolites of synthetic pyrethroids and embryological outcomes (MII oocyte count, top quality embryo, fertilization and implantation rate). We included 450 women aged 25−45 undergoing assisted reproductive technology (ART) cycle at Infertility Clinic in Poland. Urine samples were collected at the time of fertility procedure(s) to assess four urinary synthetic pyrethroids concentrations (3-phenoxybenzoic acid (3PBA), cis-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane carboxylic acid (cis-DCCA), trans-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane carboxylic acid (trans-DCCA), cis-2,2-dibromovinyl-2,2-dimethylocyclopropane-1-carboxylic acid (DBCA)) using validated gas chromatography ion-tap mass spectrometry and calculated for each cycle-specific metabolite. To evaluate the effect of environmental exposure to synthetic pyrethroids and embryological outcomes (methaphase II (MII) oocyte yield, top quality embryo, fertilization rate, implantation rate), multivariable generalized linear mixed analyses with random intercepts were prepared. Urinary 3-PBA concentrations decrease MII oocyte count (p = 0.007) in the fourth quartile (>75 percentile) compared to women in the first quartile (≤25 percentile). Additionally, when 3-PBA was treated as continuous variable, the negative association between exposure to pyrethroids and MII oocyte count was also observed (p = 0.012). Exposure to other pyrethroid metabolities (CDCCA, TDCCA, DBCA) was not related to any of the examined embryological outcomes. Exposure to synthetic pyrethroids may be associated with poorer embryological outcome among couples seeking fertility treatments. As this is the first study on this topic, the results need to be confirmed in further studies.

Quality Control of Bacterial Extracellular Vesicles with Total Protein Content Assay, Nanoparticles Tracking Analysis, and Capillary Electrophoresis.

Extracellular vesicles (EVs) were isolated from Pectobacterium zantedeschiae culturing media using direct ultracentrifugation (UC), iodixanol cushion ultracentrifugation (ICUC), and iodixanol density gradient ultracentrifugation (IDGUC) techniques. The isolates were characterized with total protein content assay (bicinchoninic acid assay, BCA), nanoparticles tracking analysis (NTA), and capillary electrophoresis (CE). A satisfactory correlation (R2 > 0.94) between quantitative results obtained with BCA, NTA and CE was achieved only for isolates obtained with the IDGUC. The correlation between protein content and CE was proved to be related to the isolates' purity. The NTA was found unable to provide reliable information on EVs quantity in samples isolated with UC and ICUC, due to the co-isolated particulate impurities. Moreover, the work reports polysaccharides, used as culturing media components, as a potential source of bias of quantitation with total protein content assay and NTA. The study demonstrates the advantageous selectivity of CE in quality control of EVs and its ability to differentiate subpopulations of EVs of Pectobacterium.

Silicone Wristbands in Exposure Assessment: Analytical Considerations and Comparison with Other Approaches.

Humans are exposed to numerous potentially harmful chemicals throughout their lifetime. Although many studies have addressed this issue, the data on chronic exposure is still lacking. Hence, there is a growing interest in methods and tools allowing to longitudinally track personal exposure to multiple chemicals via different routes. Since the seminal work, silicone wristbands (WBs) have been increasingly used to facilitate human exposure assessment, as using WBs as a wearable sampler offers new insights into measuring chemical risks involved in many ambient and occupational scenarios. However, the literature lacks a detailed overview regarding methodologies being used; a comprehensive comparison with other approaches of personal exposure assessment is needed as well. Therefore, the aim of this review is fourfold. First, we summarize hitherto conducted research that employed silicone WBs as personal passive samplers. Second, all pre-analytical and analytical steps used to obtain exposure data are discussed. Third, we compare main characteristics of WBs with key features of selected matrices used in exposure assessment, namely urine, blood, hand wipes, active air sampling, and settled dust. Finally, we discuss future needs of research employing silicone WBs. Our work shows a variety of possibilities, advantages, and caveats associated with employment of silicone WBs as personal passive samplers. Although further research is necessary, silicone WBs have already been proven valuable as a tool for longitudinal assessment of personal exposure.

Urinary Bisphenol A Concentrations and Parameters of Ovarian Reserve among Women from a Fertility Clinic.

BACKGROUND: Human exposure to environmentally widespread endocrine disruptors, especially bisphenol A (BPA), has been suggested to affect reproductive health. Animal studies indicate that BPA may play a role in the process of reproduction and impact on maturing oocytes, meiotic cell division or fertilization rate. Nevertheless, data regarding the effects of exposure to BPA on women's ovarian function are still limited. Therefore, the aim of the current study is to assess the effects of environmental exposure to BPA on ovarian reserve. METHODS: The study participants consisted of 511 women in reproductive age (25-39 years) who attended an infertility clinic for diagnosis, due to the couples' infertility. BPA urinary concentrations were assessed by the validated gas chromatography ion-trap mass spectrometry method. The ovarian reserve was assessed using ovarian reserve parameters: Hormones concentrations: E2 (estradiol), FSH (follicle stimulating hormone), AMH (anti-Müllerian hormone), and AFC (antral follicle count). RESULTS: In the present study, the negative association between BPA urinary concentrations and AMH (p = 0.02) and AFC (p = 0.03) levels was found. Exposure to BPA was not related to other examined parameters of ovarian reserve (FSH, E2). CONCLUSIONS: Our results suggest that BPA exposure may affect women ovarian reserve parameters and reduce ovarian reserve. As this is one of the first studies of its kind, the findings need confirmation in a further investigation.

Investigation of selected parameters of capillary zone electrophoresis method for analysis of isolates of outer membrane vesicles.

The capillary zone electrophoresis (CZE) has recently been proposed by our group as a novel technique for outer membrane vesicles (OMVs) characterization (J. Chromatography 1621 (2020) 461047). In present work the impact of selected parameters of CZE method on OMVs isolates analysis was assessed. It was shown that the extension of sample injection plug length significantly improves the detectability of macromolecular aggregates in CZE. Moreover, a negligible adsorption of OMVs to both uncoated and polymer-modified (poly(DMA-GMA-MAPS)) capillary walls was proven. Finally, the relaxation effect as well as deformation/polarization of vesicles were demonstrated to affect OMVs electrophoretic mobility. The significance of these findings was discussed.

DeltaF508 CFTR Hetero- and Homozygous Paediatric Patients with Cystic Fibrosis Do Not Differ with Regard to Nutritional Status.

The purpose of this study was to compare the nutritional status between deltaF508 CFTR hetero- and homozygous paediatric patients with cystic fibrosis. We assessed the percentage profiles of fatty acids measured in erythrocyte membranes and the serum levels of vitamins A, D3, E and K1 in the studied groups. We also measured the weights and heights and calculated the body mass indexes (BMIs). The studied groups consisted of 34 heterozygous and 30 homozygous patients. No statistically significant differences were found in the serum vitamins or erythrocyte membrane fatty acid profiles between the hetero- and homozygous patient groups, except for heptadecanoic acid (p = 0.038). The mean percentiles of height, weight and BMI did not differ significantly between the two groups. The homozygous and heterozygous paediatric patients with cystic fibrosis were similar in terms of their nutritional statuses.

Development and validation of a gas chromatography method coupled with flame ionization detector for quantitative analysis of fragrance allergens in aromas for e-cigarettes.

Aromas can give smell and/or flavor to a variety of products in the cosmetic and food industry. They are also used as e-cigarette additives. Some of those substances are recognized as fragrance allergens and can cause allergic reactions so there is a need to control their use. Gas chromatography is the method of choice for analyzing fragrance allergens because of their low boiling points. This study aimed to develop and validate a robust and simple method for the analysis of fragrance allergens in aromas for e-cigarettes. A method using gas chromatography coupled with a flame ionization detector was developed for 25 fragrance allergens. Optimized parameters were sample diluent, internal standard, and carrier gas. The output method was the one previously developed and optimized. The linearity of the method was >0.994 over the range of 0.5-40 µg/mL. Accuracy and precision were within the acceptance range. Limits of detection and quantification were determined, and calibration curves were plotted. The method was applied to three real samples of aromas. Thirteen fragrance allergens were detected. Concentrations varied in the range of dozens to thousands of µg/mL showed that concentrations of fragrances in aromas for e-cigarettes can be high and varies among products.

Concentrations of urinary biomarkers and predictors of exposure to pyrethroid insecticides in young, Polish, urban-dwelling men.

Pyrethroid insecticides are a class of pesticides with multiple agricultural and residential applications. However, widespread use of these chemicals may pose a threat to human health. Biomarkers of pyrethroid exposure are frequently detected in populations around the world, but some groups may be underrepresented. Moreover, there is an ongoing debate on factors contributing to pyrethroid burden in humans. To address these problems, we measured urinary biomarkers of pyrethroid exposure in urine samples from 306 young men living in urban area of Lódz, Poland, and gathered questionnaire data to identify predictors of exposure. Limit of detection (LOD) of gas chromatography-mass spectrometry (GC-MS) method was 0.1 ng/mL for all quantified pyrethroid metabolites, namely cis-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane carboxylic acid (cis-DCCA), trans-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane carboxylic acid (trans-DCCA), cis-3-(2,2-dibromovinyl)-2,2-dimethylcyclopropane carboxylic acid (cis-DBCA), and 3-phenoxybenzoic acid (3-PBA). Detection rate ranged from 32% (cis-DBCA) to 76% (trans-DCCA). Concentrations of urinary biomarkers in studied sample were in lower range of these observed in similar studies, with unadjusted geometric means (GMs) of most prevalent biomarkers, trans-DCCA and 3-PBA, equal to 0.268 and 0.228 ng/mL, respectively. As for questionnaire data, the statistical analysis revealed that non-dietary factors, especially dog ownership and pesticide use on household pets, contribute significantly to urinary trans-DCCA and 3-PBA concentrations (p ≤ 0.009). Moreover, a few dietary sources of exposure were identified, such as seeds and nuts consumption for 3-PBA (p < 0.001) and vegetable juice intake for trans-DCCA (p = 0.015). Multivariate analyses further highlighted the importance of non-dietary factors in pyrethroid exposure. Compared to other works, our results confirm widespread exposure to pyrethroids observed in other studies and stress the role of residential pyrethroid use in pyrethroid burden in humans.

Triclosan exposure and in vitro fertilization treatment outcomes in women undergoing in vitro fertilization.

Triclosan (TCS) is a widespread environmental endocrine-disrupting chemical. Animal and in vitro studies suggested that triclosan may affect homesostasis of sex and thyroid hormones and impact on reproduction. Due to limited data derived from human epidemiological studies, this study was performed to examine the association between urinary concentration of triclosan and in vitro reproductive outcomes (methaphase II (MII) oocyte yield, top quality embryo, fertilization rate, implantation rate, and clinical pregnancy) among women from infertility clinic. The study participants were enrolled in an Infertility Center in Poland. A total of 450 women aged 25-45 (n = 674 IVF cycles) provided urine samples. The urinary concentrations of triclosan were evaluated using validated gas chromatography ion-tap mass spectrometry method. Clinical outcomes of IVF treatment were abstracted from patients electronic chart records. Triclosan was detected in urine of 82% of women with geometric mean 2.56 ± 6.13 ng/mL. Urinary concentrations of triclosan were associated with decrease implantation rate (p = 0.03). There were no association between other examined IVF outcomes: MII oocytes, embryo quality, fertilization rate, and exposure to triclosan. As this is one of the first study on this topic, studies among larger and more diverse population are needed to confirm the results.