An engineered PET depolymerase to break down and recycle plastic bottles.

Present estimates suggest that of the 359 million tons of plastics produced annually worldwide1, 150-200 million tons accumulate in landfill or in the natural environment2. Poly(ethylene terephthalate) (PET) is the most abundant polyester plastic, with almost 70 million tons manufactured annually worldwide for use in textiles and packaging3. The main recycling process for PET, via thermomechanical means, results in a loss of mechanical properties4. Consequently, de novo synthesis is preferred and PET waste continues to accumulate. With a high ratio of aromatic terephthalate units-which reduce chain mobility-PET is a polyester that is extremely difficult to hydrolyse5. Several PET hydrolase enzymes have been reported, but show limited productivity6,7. Here we describe an improved PET hydrolase that ultimately achieves, over 10 hours, a minimum of 90 per cent PET depolymerization into monomers, with a productivity of 16.7 grams of terephthalate per litre per hour (200 grams per kilogram of PET suspension, with an enzyme concentration of 3 milligrams per gram of PET). This highly efficient, optimized enzyme outperforms all PET hydrolases reported so far, including an enzyme8,9 from the bacterium Ideonella sakaiensis strain 201-F6 (even assisted by a secondary enzyme10) and related improved variants11-14 that have attracted recent interest. We also show that biologically recycled PET exhibiting the same properties as petrochemical PET can be produced from enzymatically depolymerized PET waste, before being processed into bottles, thereby contributing towards the concept of a circular PET economy.

Biochemical and structural characterization of an aromatic ring-hydroxylating dioxygenase for terephthalic acid catabolism.

SignificanceMore than 400 million tons of plastic waste is produced each year, the overwhelming majority of which ends up in landfills. Bioconversion strategies aimed at plastics have emerged as important components of enabling a circular economy for synthetic plastics, especially those that exhibit chemically similar linkages to those found in nature, such as polyesters. The enzyme system described in this work is essential for mineralization of the xenobiotic components of poly(ethylene terephthalate) (PET) in the biosphere. Our description of its structure and substrate preferences lays the groundwork for in vivo or ex vivo engineering of this system for PET upcycling.

Mono-(2-ethyl-5-hydroxyhexyl) phthalate promotes uterine leiomyoma cell survival through tryptophan-kynurenine-AHR pathway activation.

Uterine leiomyoma is the most common tumor in women and causes severe morbidity in 15 to 30% of reproductive-age women. Epidemiological studies consistently indicate a correlation between leiomyoma development and exposure to endocrine-disrupting chemical phthalates, especially di-(2-ethylhexyl) phthalate (DEHP); however, the underlying mechanisms are unknown. Here, among the most commonly encountered phthalate metabolites, we found the strongest association between the urine levels of mono(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), the principal DEHP metabolite, and the risk of uterine leiomyoma diagnosis (n = 712 patients). The treatment of primary leiomyoma and smooth muscle cells (n = 29) with various mixtures of phthalate metabolites, at concentrations equivalent to those detected in urine samples, significantly increased cell viability and decreased apoptosis. MEHHP had the strongest effects on both cell viability and apoptosis. MEHHP increased cellular tryptophan and kynurenine levels strikingly and induced the expression of the tryptophan transporters SLC7A5 and SLC7A8, as well as, tryptophan 2,3-dioxygenase (TDO2), the key enzyme catalyzing the conversion of tryptophan to kynurenine that is the endogenous ligand of aryl hydrocarbon receptor (AHR). MEHHP stimulated nuclear localization of AHR and up-regulated the expression of CYP1A1 and CYP1B1, two prototype targets of AHR. siRNA knockdown or pharmacological inhibition of SLC7A5/SLC7A8, TDO2, or AHR abolished MEHHP-mediated effects on leiomyoma cell survival. These findings indicate that MEHHP promotes leiomyoma cell survival by activating the tryptophan-kynurenine-AHR pathway. This study pinpoints MEHHP exposure as a high-risk factor for leiomyoma growth, uncovers a mechanism by which exposure to environmental phthalate impacts leiomyoma pathogenesis, and may lead to the development of novel druggable targets.

Increasing the Soluble Expression and Whole-Cell Activity of the Plastic-Degrading Enzyme MHETase through Consensus Design.

The mono(2-hydroxyethyl) terephthalate hydrolase (MHETase) from Ideonella sakaiensis carries out the second step in the enzymatic depolymerization of poly(ethylene terephthalate) (PET) plastic into the monomers terephthalic acid (TPA) and ethylene glycol (EG). Despite its potential industrial and environmental applications, poor recombinant expression of MHETase has been an obstacle to its industrial application. To overcome this barrier, we developed an assay allowing for the medium-throughput quantification of MHETase activity in cell lysates and whole-cell suspensions, which allowed us to screen a library of engineered variants. Using consensus design, we generated several improved variants that exhibit over 10-fold greater whole-cell activity than wild-type (WT) MHETase. This is revealed to be largely due to increased soluble expression, which biochemical and structural analysis indicates is due to improved protein folding.

Biomimetic Mineralization of Large Enzymes Utilizing a Stable Zirconium-Based Metal-Organic Frameworks.

Enzymes are natural catalysts for a wide range of metabolic chemical transformations, including selective hydrolysis, oxidation, and phosphorylation. Herein, we demonstrate a strategy for the encapsulation of enzymes within a highly stable zirconium-based metal-organic framework. UiO-66-F4 was synthesized under mild conditions using an enzyme-compatible amino acid modulator, serine, at a modest temperature in an aqueous solution. Enzyme@UiO-66-F4 biocomposites were then formed by an in situ encapsulation route in which UiO-66-F4 grows around the enzymes and, consequently, provides protection for the enzymes. A range of enzymes, namely, lysozyme, horseradish peroxidase, and amano lipase, were successfully encapsulated within UiO-66-F4. We further demonstrate that the resulting biocomposites are stable under conditions that could denature many enzymes. Horseradish peroxidase encapsulated within UiO-66-F4 maintained its biological activity even after being treated with the proteolytic enzyme pepsin and heated at 60 °C. This strategy expands the toolbox of potential metal-organic frameworks with different topologies or functionalities that can be used as enzyme encapsulation hosts. We also demonstrate that this versatile process of in situ encapsulation of enzymes under mild conditions (i.e., submerged in water and at a modest temperature) can be generalized to encapsulate enzymes of various sizes within UiO-66-F4 while protecting them from harsh conditions (i.e., high temperatures, contact with denaturants or organic solvents).

Evaluating Mixtures of Urinary Phthalate Metabolites and Serum Per-/Polyfluoroalkyl Substances in Relation to Adolescent Hair Cortisol: The HOME Study.

Results of toxicological studies indicate that phthalates and per-/polyfluoroalkyl substances (PFAS), 2 classes of endocrine-disrupting chemicals, may alter the functioning of the hypothalamic-pituitary-adrenocortical (HPA) axis. We evaluated the associations of urinary phthalate metabolites and serum PFAS during gestation and childhood with adolescent hair cortisol concentrations (pg/mg hair) at age 12 years, an integrative marker of HPA axis activity (n = 205 mother-child pairs; Cincinnati, Ohio; enrolled 2003-2006). We used quantile-based g-computation to estimate associations between mixtures of urinary phthalate metabolites or serum PFAS and hair cortisol. We also examined whether associations of individual phthalate metabolites or PFAS with cortisol varied by the timing of exposure. We found that a 1-quartile increase in all childhood phthalate metabolites was associated with 35% higher adolescent hair cortisol (phthalate mixture ψ = 0.13; 95% confidence interval: 0.03, 0.22); these associations were driven by monoethyl phthalate, monoisobutyl phthalate, and monobenzyl phthalate. We did not find evidence that phthalate metabolites during gestation or serum PFAS mixtures were related to adolescent hair cortisol concentrations. We found suggestive evidence that higher childhood concentrations of individual PFAS were related to higher and lower adolescent hair cortisol concentrations. Our results suggest that phthalate exposure during childhood may contribute to higher levels of chronic HPA axis activity.

Accelerating the Phosphatase-like Activity of Uio-66-NH2 by Catalytically Inactive Metal Ions and Its Application for Improved Fluorescence Detection of Cardiac Troponin I.

Compared with natural enzymes, nanozymes usually exhibit much lower catalytic activities, which limit the sensitivities of nanozyme-based immunoassays. Herein, several metal ions without enzyme-like activities were engineered onto Uio-66-NH2 nanozyme through postsynthetic modification. The obtained Mn+@Uio-66-NH2 (Mn+ = Zn2+, Cd2+, Co2+, Ca2+and Ni2+) exhibited improved phosphatase-like catalytic activities. In particular, a 12-fold increase in the catalytic efficiency (kcat/Km) of Uio-66-NH2 was observed after the modification with Zn2+. Mechanism investigations indicate that both the amino groups and oxygen-containing functional groups in Uio-66-NH2 are the binding sites of Zn2+, and the modified Zn2+ ions on Uio-66-NH2 serve as the additional catalytic sites for improving the catalytic performance. Furthermore, the highly active Zn2+@Uio-66-NH2 was used as a nanozyme label to develop a fluorescence immunoassay method for the detection of cardiac troponin I (cTnI). Compared with pristine Uio-66-NH2, Zn2+@Uio-66-NH2 can widen the linear range by 1 order of magnitude (from 10 pg/mL-1 µg/mL to 1 pg/mL-1 µg/mL) and also lower the detection limit by 5 times (from 4.7 pg/mL to 0.9 pg/mL).

Programmable Lanthanide Metal-Organic Framework for Ultra-Efficient Nucleic Acids Extraction and Interaction Analysis.

Efficient, mild, and reversible adsorption of nucleic acids onto nanomaterials represents a promising analytical approach for medical diagnosis. However, there is a scarcity of efficient and reversible nucleic acid adsorption nanomaterials. Additionally, the lack of comprehension of the molecular mechanisms governing their interactions poses significant challenges. These issues hinder the rational design and analytical applications of the nanomaterials. Herein, we propose an ultra-efficient nucleic acid affinity nanomaterial based on programmable lanthanide metal-organic frameworks (Ln-MOFs). Through experiments and density functional theory calculations, a rational design guideline for nucleic acid affinity of Ln-MOF was proposed, and a modular and flexible preparation scheme was provided. Then, Er-TPA (terephthalic acid) MOF emerged as the optimal candidate due to its pore size-independent adsorption and desorption capabilities for nucleic acids, enabling ultra-efficient adsorption (about 150% mass ratio) within 1 min. Furthermore, we elucidate the molecular-level mechanisms underlying the Ln-MOF adsorption of single- and double-stranded DNA and G4 structures. The affinity nanomaterial based on Ln-MOF exhibits robust nucleic acid extraction capability (4-fold higher than commercial reagent kits) and enables mild and reversible CRISPR/Cas9 functional regulation. This method holds significant promise for broad application in DNA/RNA liquid biopsy and gene editing, facilitating breakthroughs in analytical chemistry, pharmacy, and medical research.

Biomimetic delivery of emodin via macrophage membrane-coated UiO-66-NH2 nanoparticles for acute pancreatitis treatment.

Acute pancreatitis (AP) is a severe inflammatory condition with a rising incidence and high mortality rates, especially in severe cases. Emodin (ED), known for its potent anti-inflammatory properties, holds promise in addressing AP. However, its clinical application is hindered by limitations such as low bioavailability and insufficient target specificity. Herein, we developed a novel drug delivery system using macrophage membrane-coated UiO-66-NH2 nanoparticles loaded with ED (MVs-UiO-ED). UiO-66-NH2 was successfully synthesized and characterized, revealing an octahedral structure with a suitable size distribution. The successful loading of ED onto UiO-66-NH2 was confirmed by ultraviolet and infrared spectroscopy. Subsequently, MVs-UiO-ED was prepared by coating macrophage membrane-derived vesicles onto UiO-ED, resulting in a biomimetic delivery system. In vitro release studies demonstrated that MVs-UiO-ED exhibited a sustained-release profile, indicating its potential for prolonged drug circulation. An AP mouse model was established to evaluate the therapeutic efficacy of MVs-UiO-ED. Compared with the model group, MVs-UiO-ED significantly reduced serum levels of α-amylase and lipase, two indicators of pancreatitis severity. Furthermore, histopathological examinations revealed that MVs-UiO-ED ameliorated pancreatic tissue damage. This study underscores the potential of MVs-UiO-ED as an effective therapeutic approach for AP.

Arbuscular mycorrhizal fungi by inducing watermelon roots secretion phthalates, altering soil enzyme activity and bacterial community composition to alleviate the watermelon wilt.

BACKGROUND: Long-term continuous cropping has resulted in the frequent occurrence of fusarium wilt of watermelon (Citrullus lanatus). AMF inoculation can alleviate the continuous cropping barrier and reduce the incidence of fusarium wilt of watermelon. Our previous study found that the root exudates of mycorrhizal watermelon can enhance watermelon resistance to this disorder. It is necessary to further isolate and identify the specific compounds in root exudates of mycorrhizal watermelon and explore their control effects on fusarium wilt of continuous cropping watermelon. RESULT: The results of this study showed that the root system of watermelon seedlings inoculated with AMF (Funneliformis mosseae or Glomus versiforme) secreted diisooctyl phthalate (A) and dibutyl phthalate (B). Compared with water treatment, treatment with 0.1 ml/L (A1, B1), 0.5 ml/L (A2, B2) and 1 ml/L (A3, B3) of A or B significantly increased soil enzyme activities, the numbers of bacteria and actinomycetes, and the bacteria/fungi ratio in the rhizosphere. Furthermore, the Disease indexes (DI) of A1 and B3 were 25% and 20%, respectively, while the prevention and control effects (PCE) were 68.8% and 75%, respectively. In addition, diisooctyl phthalate or dibutyl phthalate increased the proportions of Gemmatimonadetes, Chloroflexi, and Acidobacteria in the rhizosphere of continuous cropping watermelon, and decreased the proportions of Proteobacteria and Firmicutes, with Novosphingobium, Kaistobacter, Bacillus, and Acinetobacter as the predominant bacteria. Compared with the water treatment, the abundance of Neosphingosaceae, Kateybacterium and Bacillus in the A1 group was increased by 7.33, 2.14 and 2.18 times, respectively, while that in the B2 group was increased by 60.05%, 80.24% and 1 time, respectively. In addition, exogenous diisooctyl phthalate and dibutyl phthalate were shown to promote growth parameters (vine length, stem diameter, fresh weight and dry weight) and antioxidant enzyme system activities (SOD, POD and CAT) of continuous cropping watermelon. CONCLUSION: Lower watermelon fusarium wilt incidence in mycorrhizal watermelons was associated with phthalate secretion in watermelons after AMF inoculation. Exogenous diisooctyl phthalate and dibutyl phthalate could alleviate the continuous cropping disorder of watermelon, reduce the incidence of fusarium wilt, and promote the growth of watermelon by increasing the enzyme activities and the proportion of beneficial bacteria in rhizosphere soil. In addition, the low concentration of phthalate diisooctyl and high concentration of phthalic acid dibutyl works best. Therefore, a certain concentration of phthalates in the soil can help alleviate continuous cropping obstacles.

The effects of short-term and long-term phthalate exposures on ovarian follicle growth dynamics and hormone levels in female mice†.

Di(2-ethylhexyl) phthalate and diisononyl phthalate are widely used as plasticizers in polyvinyl chloride products. Short-term exposures to phthalates affect hormone levels, ovarian follicle populations, and ovarian gene expression. However, limited data exist regarding the effects of long-term exposure to phthalates on reproductive functions. Thus, this study tested the hypothesis that short-term and long-term exposure to di(2-ethylhexyl) phthalate or diisononyl phthalate disrupts follicle dynamics, ovarian and pituitary gene expression, and hormone levels in female mice. Adult CD-1 female mice were exposed to vehicle, di(2-ethylhexyl) phthalate, or diisononyl phthalate (0.15 ppm, 1.5 ppm, or 1500 ppm) via the chow for 1 or 6 months. Short-term exposure to di(2-ethylhexyl) phthalate (0.15 ppm) and diisononyl phthalate (1.5 ppm) decreased serum follicle-stimulating hormone levels compared to control. Long-term exposure to di(2-ethylhexyl) phthalate and diisononyl phthalate (1500 ppm) increased the percentage of primordial follicles and decreased the percentages of preantral and antral follicles compared to control. Both phthalates increased follicle-stimulating hormone levels (di(2-ethylhexyl) phthalate at 1500 ppm; diisononyl phthalate at 1.5 ppm) and decreased luteinizing hormone levels (di(2-ethylhexyl) phthalate at 0.15 and 1.5 ppm; diisononyl phthalate at 1.5 ppm and 1500 ppm) compared to control. Furthermore, both phthalates altered the expression of pituitary gonadotropin subunit genes (Cga, Fshb, and Lhb) and a transcription factor (Nr5a1) that regulates gonadotropin synthesis. These data indicate that long-term exposure to di(2-ethylhexyl) phthalate and diisononyl phthalate alters follicle growth dynamics in the ovary and the expression of gonadotropin subunit genes in the pituitary and consequently luteinizing hormone and follicle-stimulating hormone synthesis.

The role of the aryl hydrocarbon receptor in mediating the effects of mono(2-ethylhexyl) phthalate in mouse ovarian antral follicles†.

Di(2-ethylhexyl) phthalate (DEHP) is a pervasive environmental toxicant used in the manufacturing of numerous consumer products, medical supplies, and building materials. DEHP is metabolized to mono(2-ethylhexyl) phthalate (MEHP). MEHP is an endocrine disruptor that adversely affects folliculogenesis and steroidogenesis in the ovary, but its mechanism of action is not fully understood. Thus, we tested the hypothesis that the aryl hydrocarbon receptor (AHR) plays a functional role in MEHP-mediated disruption of folliculogenesis and steroidogenesis. CD-1 mouse antral follicles were isolated and cultured with MEHP (0-400 µM) in the presence or absence of the AHR antagonist CH223191 (1 µM). MEHP treatment reduced follicle growth over a 96-h period, and this effect was partially rescued by co-culture with CH223191. MEHP exposure alone increased expression of known AHR targets, cytochrome P450 (CYP) enzymes Cyp1a1 and Cyp1b1, and this induction was blocked by CH223191. MEHP reduced media concentrations of estrone and estradiol compared to control. This effect was mitigated by co-culture with CH223191. Moreover, MEHP reduced the expression of the estrogen-sensitive genes progesterone receptor (Pgr) and luteinizing hormone/choriogonadotropin receptor (Lhcgr) and co-treatment with CH223191 blocked this effect. Collectively, these data indicate that MEHP activates the AHR to impair follicle growth and reduce estrogen production and signaling in ovarian antral follicles.

Phthalate and phthalate replacement concentrations in relationship to adiposity in a multi-racial cohort of children.

BACKGROUND/OBJECTIVE: Phthalates and phthalate replacements are used in multiple everyday products, making many of them bioavailable to children. Experimental studies suggest that phthalates and their replacements may be obesogenic, however, epidemiologic studies remain inconsistent. Therefore, our objective was to examine the association between phthalates, phthalate replacements and childhood adiposity/obesity markers in children. SUBJECTS/METHODS: A cross-sectional study was conducted in 630 racial/ethnically diverse children ages 4-8 years. Urinary oxidative metabolites of DINCH and DEHTP, three low molecular weight (LMW) phthalates, and eleven high molecular weight (HMW) phthalates were measured. Weight, height, waist circumference and % body fat were measured. Composite molar sum groups (nmol/ml) were natural log-transformed. Linear regression models adjusted for urine specific gravity, sex, age, race-ethnicity, birthweight, breastfeeding, reported activity level, mother's education and pre-pregnancy BMI. RESULTS: All children had LMW and HMW phthalate metabolites and 88% had DINCH levels above the limit of detection. One unit higher in the log of DINCH was associated with 0.106 units lower BMI z-score [ß = -0.106 (95% CI: -0.181, -0.031)], 0.119 units lower waist circumference z-score [ß = -0.119 (95% CI: -0.189, -0.050)], and 0.012 units lower percent body fat [ß = -0.012 (95% CI: -0.019, -0.005)]. LMW and HMW group values were not associated with adiposity/obesity. CONCLUSIONS: We report an inverse association between child urinary DINCH levels, a non-phthalate plasticizer that has replaced DEHP in several applications, and BMI z-score, waist circumference z-score and % body fat in children. Few prior studies of phthalates and their replacements in children have been conducted in diverse populations. Moreover, DINCH has not received a great deal of attention or regulation, but it is a common exposure. In summary, understanding the ubiquitous nature of these chemical exposures and ultimately their sources will contribute to our understanding of their relationship with obesity.

A Mixture of Urinary Phthalate Metabolite Concentrations During Pregnancy and Offspring Social Responsiveness Scale Scores.

BACKGROUND: Phthalates are a group of chemicals with ubiquitous exposure worldwide. Exposures to phthalates during pregnancy may play a role in autism spectrum disorder (ASD) etiology by disrupting hormone levels or directly impacting fetal neurodevelopment. However, there is little research quantifying the aggregate effect of phthalates on child ASD-related behaviors. METHODS: We used data from two prospective pregnancy and birth cohorts-the Health Outcomes and Measures of the Environment (HOME) and the Early Autism Risk Longitudinal Investigation (EARLI). HOME is a general population cohort while participants in EARLI were at higher familial risk for ASD. Using quantile g-computation and linear regression models, we assessed the joint and individual associations of a mixture of six phthalate metabolites during pregnancy with child ASD-related traits measured by Social Responsiveness Scale (SRS) scores at ages 3-8 years. RESULTS: Our analyses included 271 participants from HOME and 166 participants from EARLI. There were imprecise associations between the phthalate mixture and SRS total raw scores in HOME (difference in SRS scores per decile increase in every phthalate = 1.3; 95% confidence interval [CI] = -0.2, 2.8) and EARLI (difference in SRS scores per decile increase in every phthalate = -0.9; 95% CI = -3.5, 1.7). CONCLUSIONS: The cohort-specific effect sizes of the pthalates-SRS associations were small and CIs were imprecise. These results suggest that if there are associations between phthalate metabolites during pregnancy and child SRS scores, they may differ across populations with different familial liabilities. Further studies with larger sample sizes are warranted.

Endocrine disruption and male reproductive disorders: unanswered questions.

Maternal exposure to endocrine-disrupting chemicals (EDCs) in human pregnancy is widely considered as an important cause of adverse changes in male reproductive health due to impaired foetal androgen production/action. However, the epidemiological evidence supporting this view is equivocal, except for certain phthalates, notably diethyl hexyl phthalate (DEHP). Maternal phthalate exposure levels associated with adverse reproductive changes in epidemiological studies are several thousand-fold lower than those needed to suppress foetal androgen production in rats, and direct studies using human foetal testis tissue show no effect of high phthalate exposure on androgen production. This conundrum is unexplained and raises fundamental questions. Human DEHP exposure is predominantly via food with highest exposure associated with consumption of a Western style (unhealthy) diet. This diet is also associated with increased exposure to the most common EDCs, whether persistent (chlorinated or fluorinated chemicals) or non-persistent (phthalates, bisphenols) compounds, which are found at highest levels in fatty and processed foods. Consequently, epidemiological studies associating EDC exposure and male reproductive health disorders are confounded by potential dietary effects, and vice versa. A Western diet/lifestyle in young adulthood is also associated with low sperm counts. Disentangling EDC and dietary effects in epidemiological studies is challenging. In pregnancy, a Western diet, EDC exposure, and maternal living in proximity to industrial sites are all associated with impaired foetal growth/development due to placental dysfunction, which predisposes to congenital male reproductive disorders (cryptorchidism, hypospadias). While the latter are considered to reflect impaired foetal androgen production, effects resulting from foetal growth impairment (FGI) are likely indirect. As FGI has numerous life-long health consequences, and is affected by maternal lifestyle, research into the origins of male reproductive disorders should take more account of this. Additionally, potential effects on foetal growth/foetal testis from the increasing use of medications in pregnancy deserves more research attention.

Longitudinal associations between urinary biomarkers of phthalates and replacements with novel in vivo measures of placental health.

STUDY QUESTION: What is the longitudinal association between gestational phthalate exposure and in vivo placental outcomes? SUMMARY ANSWER: Phthalates were adversely associated with placental microvasculature, stiffness, and presence of calcification, with different metabolites associated with different outcomes. WHAT IS KNOWN ALREADY: Phthalate exposure is ubiquitous and implicated as a contributor to adverse pregnancy outcomes, possibly through impacts on the placenta. STUDY DESIGN, SIZE, DURATION: A total of 303 women were recruited in early pregnancy and prospectively followed for up to eight visits across gestation in the Human Placenta and Phthalates study. PARTICIPANTS/MATERIALS, SETTING, METHODS: At each visit, women provided urine samples and underwent placental ultrasounds. Urine was analyzed for 18 metabolites of phthalates and replacements. We took the geometric mean of repeated measurements to reflect pregnancy-averaged phthalate or replacement exposure for each participant (n = 303). Placental microvasculature, stiffness, and microcalcification presence were quantified from ultrasounds at each visit. Higher scores reflected worse placental function for all measures. Generalized linear mixed models were created to estimate the association between pregnancy-averaged exposure biomarker concentrations and repeated outcome measurements for microvasculature and stiffness. Gestational age at the time of calcification detection was modeled using Cox proportional hazards models. MAIN RESULTS AND THE ROLE OF CHANCE: Monocarboxyisononyl phthalate and summed di(2-ethylhexyl) phthalate metabolites were associated with impaired microvasculature development, such that an interquartile range increase in concentration was associated with 0.11 standard deviation increase in the microvasculature ratio, indicating poorer vascularization (95% CI: 0.00, 0.22); 0.11 [95% CI: -0.01, 0.22], respectively. Monoethyl phthalate was associated with increased placental stiffness (0.09 [95% CI: -0.01, 0.19]) while summed di-iso-butyl phthalate metabolites and monobenzyl phthalate were associated with increased hazard of calcification detection (hazard ratios: 1.18 [95% CI: 0.98, 1.42]; 1.13 [95% CI: 0.96, 1.34]). LIMITATIONS, REASONS FOR CAUTION: Outcomes used in this study are novel and further investigation is needed to provide clinical context and relevance. WIDER IMPLICATIONS OF THE FINDINGS: We found evidence of associations between select phthalate biomarkers and various aspects of in vivo placental health, although we did not observe consistency across placental outcomes. These findings could illustrate heterogeneous effects of phthalate exposure on placental function. STUDY FUNDING/COMPETING INTEREST(S): This research was supported in part by the Intramural Research Program of the NIH, National Institute of Environmental Health Sciences (ZIA ES103344), and NIEHS T32ES007018. The authors declare that they have no competing interests to disclose. The findings and conclusions in this report are those of the authors and do not necessarily represent the official position of the Centers for Disease Control and Prevention. Use of trade names is for identification only and does not imply endorsement by the CDC, the Public Health Service, or the US Department of Health and Human Services. TRIAL REGISTRATION NUMBER: N/A.

Ligand binding properties of three odorant-binding proteins in striped flea beetle Phyllotreta striolata towards two phthalate esters.

Odorant-binding proteins (OBPs) initiate insect olfactory perception and mediate specific binding and selection of odorants via uncertain binding mechanisms. We characterized the binding characteristics of four OBPs from the striped flea beetle Phyllotreta striolata (SFB), a major cruciferous crop pest. Tissue expression analysis revealed that the two ABPII OBPs (PstrOBP12 and PstrOBP19) were highly expressed mainly in the antenna, whereas the two minus-C OBPs (PstrOBP13 and PstrOBP16) showed a broad expression pattern. Competitive binding assays of cruciferous plant volatiles showed that PstrOBP12, PstrOBP16 and PstrOBP19 had very strong binding capacities for only two phthalate esters (Ki < 20 µM), and PstrOBP13 specifically bound to four aromatic volatiles (Ki < 11 µM). Fluorescence quenching assays displayed that two phthalate esters bound to three PstrOBPs via different quenching mechanisms. PstrOBP12/PstrOBP16-diisobutyl phthalate and PstrOBP19-bis(6-methylheptyl) phthalate followed static quenching, while PstrOBP12/PstrOBP16-bis(6-methylheptyl) phthalate and PstrOBP19-diisobutyl phthalate followed dynamic quenching. Homology modelling and molecular docking displayed that PstrOBP12-diisobutyl phthalate was driven by H-bonding and van der Waals interactions, while PstrOBP16-diisobutyl phthalate and PstrOBP19-bis(6-methylheptyl) phthalate followed hydrophobic interactions. Finally, behavioural activity analysis demonstrated that phthalate esters exhibited different behavioural activities of SFB at different doses, with low doses attracting and high doses repelling. Overall, we thus revealed the different binding properties of the three PstrOBPs to two phthalate esters, which was beneficial in shedding light on the ligand-binding mechanisms of OBPs.

Inhibitory effects of phthalate esters (PAEs) and phthalate monoesters towards human carboxylesterases (CESs).

Phthalate esters (PAEs), accompanied by phthalate monoesters as hydrolysis metabolites in humans, have been widely used as plasticizers and exhibited disruptive effects on the endocrine and metabolic systems. The present study aims to investigate the inhibition behavior of PAEs and phthalate monoesters on the activity of the important hydrolytic enzymes, carboxylesterases (CESs), to elucidate the toxicity mechanism from a new perspective. The results showed significant inhibition on CES1 and CES2 by most PAEs, but not by phthalate monoesters, above which the activity of CES1 was strongly inhibited by DCHP, DEHP, DiOP, DiPP, DNP, DPP and BBZP, with inhibition ratios exceeding 80%. Kinetic analyses and in vitro-in vivo extrapolation were conducted, revealing that PAEs have the potential to disrupt the metabolism of endogenous substances catalyzed by CES1 in vivo. Molecular docking results revealed that hydrogen bonds and hydrophobic contacts formed by ester bonds contributed to the interaction of PAEs towards CES1. These findings will be beneficial for understanding the adverse effect of PAEs and phthalate monoesters.

Long-term exposure to the mixture of phthalates induced male reproductive toxicity in rats and the alleviative effects of quercetin.

Phthalates (PEs), such as di(2-ethylhexyl) phthalate (DEHP), dibutyl phthalate (DBP) and butyl benzyl phthalate (BBP) could cause reproductive and developmental toxicities, while human beings are increasingly exposed to them at low-doses. Phytochemical quercetin (Que) is a flavonoid that has estrogenic effect, anti-inflammatory and anti-oxidant effects. This study was conducted to assess the alleviative effect of Que. on male reproductive toxicity induced by the mixture of three commonly used PEs (MPEs) at low-dose in rats, and explore the underlying mechanism. Male rats were treated with MPEs (16 mg/kg/day) and/or Que. (50 mg/kg/d) for 91 days. The results showed that MPEs exposure caused male reproductive injuries, such as decreased serum sex hormones levels, abnormal testicular pathological structure, increased abnormal sperm rate and changed expressions of PIWIL1 and PIWIL2. Furthermore, MPEs also changed the expression of steroidogenic proteins in steroid hormone metabolism, including StAR, CYP11A1, CYP17A1, 17ß-HSD, CYP19A1. However, the alterations of these parameters were reversed by Que. MPEs caused male reproductive injuries in rats; Que. inhibited MPEs' male reproductive toxicity, which might relate to the improvement of testosterone biosynthesis.

Effects of 1, 2-bis (2,4, 6-tribromophenoxy) ethane and bis (2-ethylhexyl) tetrabromophthalate on serum metabolic and lipid profiles in male rats.

This study explored the effects of 1, 2-bis (2,4, 6-tribromophenoxy) ethane (BTBPE) and bis (2-ethylhexyl) tetrabromophthalate (TBPH) on serum metabolites and lipids in male Sprague-Dawley (SD) rats. Rats were orally gavaged 250 mg/kg bw of BTBPE and 500 mg/kg bw of TBPH for 28 consecutive days. Serum samples were collected for metabolomics and lipidomics analysis. Orthogonal partial least squares discriminant analysis (OPLS-DA) was used to explore changes in rat metabolic patterns. Least absolute shrinkage and selection operator (LASSO) regression models were established using serum levels of total thyroxine (TT4), free thyroxine (FT4), and rats' grouping information as variables to screen for robust differential substances. SuperPred was the database to obtain potential targets. The metabolomics and lipidomics results showed that BTBPE and TBPH had an impact on rat metabolic patterns, affecting pathways such as vitamin B6 synthesis. For BTBPE treatment, pyridoxal and ceramide (Cer) 24:0;4O were selected as differential substances related to thyroid hormones. For TBPH treatment, dehydroascorbic acid, acylcarnitine (CAR) 19:0, and diglyceride (DG) 38:4 were selected as differential substances related to thyroid hormones. Serotonin 2c receptor and cyclooxygenase-2 were chosen as potential targets of BTBPE and TBPH, respectively. In conclusion, this study found that BTBPE and TBPH impacted the metabolism of rats, and this effect may be related to changes in thyroid function.