Detection of Endocrine Disruptors in Water around Landfills

[Objectives]This study was conducted to explore the occurrence levels of endocrine disruptors(EDCs)in rural areas around a county landfill in Tongren City.[Methods]The water around the landfill was sampled and analyzed.A solid-phase extraction and high performance liquid chromatography-tandem mass spectrometry(SPE-UPLC-MS/MS)method was established for the determination of 27 EDCs.After the HLB solid-phase extraction column was activated,a water sample,which was adjusted with phosphoric acid to a pH of 2(±0.5)and added with 500 mg of disodium EDTA,was loaded,and 5 ml of water and 20%methanol water was added for washing.Next,10 ml of elution solution was added for elution,and the collected eluate was evaporated under reduced pressure at 40℃to near dryness,and 1 ml of reconstitution solution was added to a constant volume.An ACQUITY UPLC BEH C18(100×2.1 mm,2.6μm)chromatographic column was adopted for LC separation by gradient elution with pure water solution-acetonitrile as the mobile phase.For MS detection,the MRM mode was adopted for collection,and the positive and negative ion modes were switched for simultaneous determination,and the internal standard method was used for quantification.[Results]The correlation coefficient R2 was greater than 0.99 in the linear range of each target substance.The limits of quantitation in the method were between 0.05 and 2.00 ng/L,and the recoveries ranged from 75.3%to 105.7%.[Conclusions]The method has high sensitivity,good accuracy and strong practical value.

Endocrine disruptors and estrogenic effects on male reproductive axis

Endocrine disruptors （e.g., polychlorinated biphenyls [PCBs], dichlorodiphenyl-trichloroethane [DDT], dioxin, and some pesticides） are estrogen-like and anti-androgenic chemicals in the environment. They mimic natural hormones, inhibit the action of hormones, or alter the normal regulatory function of the endocrine system and have potential hazardous effects on male reproductive axis causing infertility. Although testicular and prostate cancers, abnormal sexual development, undescended testis, chronic inflammation, Sertoli-cell-only pattern, hypospadias, altered pituitary and thyroid gland functions are also observed, the available data are insufficient to deduce worldwide conclusions. The development of intra-cytoplasmic sperm injection （ICSI） is beyond doubt the most important recent breakthrough in the treatment of male infertility, but it does not necessarily treat the cause and may inadvertently pass on adverse genetic consequences. Many well-controlled clinical studies and basic scientific discoveries in the physiology, biochemistry, and molecular and cellular biology of the male reproductive system have helped in the identification of greater numbers of men with male factor problems. Newer tools for the detection of Y-chromosome deletions have further strengthened the hypothesis that the decline in male reproductive health and fertility may be related to the presence of certain toxic chemicals in the environment. Thus the etiology, diagnosis, and treatment of male factor infertility remain a real challenge. Clinicians should always attempt to identify the etiology of a possible testicular toxicity, assess the degree of risk to the patient being evaluated for infertility, and initiate a plan to control and prevent exposure to others once an association between occupation/toxicant and infertility has been established.

Testing Potential Effect of Environmental Endocrine Disruptors in Cow Milk on Reproductive Index in Female Rats

To study the effect of endocrine disruptor chemicals in cow milk on female reproductive system. Methods A two-generation reproduction was conducted according to U. S. FDA standard. Milk was fed in special bottle to Wistar rats of both sexes through two successive generations （F0 and F1） in the milk group while artificial milk was fed to rats in the control group. Twenty-four rats of each sex were mated in each group. Measurements were made according to this guideline. Results Reproductive parameters in the milk group such as fertility index, gestation index, weights of uterus and ovary, days of vaginal opening, estrous cycles, histological morphological changes were comparable to those in the control group. However, the means of body weight had some differences. The body weight gains increased significantly in the milk-treated group in F1 and F2 generation compared with those in the control group. The concentration of insulin-like growth factor-1 （IGF-1） in blood in the milk group was comparable to that in the control group, but the standard deviation changed greatly in the milk-treated rats. Conclusion Endocrine disruptor chemicals in milk have no severe effects on the female reproductive system.

Endocrine disruptor chemicals as obesogen and diabetogen: Clinical and mechanistic evidence

Obesity is becoming an inevitable pandemic all over the world.The World Obesity Federation predicts in the 2022 World Obesity Atlas that one billion people worldwide,including 1 in 5 women and 1 in 7 men,will be living with obesity by 2030.Moreover,the prevalence of diabetes is increasing worldwide,and diabetes is becoming more of a public health problem.Increased insulin resistance due to obesity and deficiency in insulin secretion are the two main causes of type 2 diabetes mellitus(T2DM).An exogenous chemical or mixture of chemicals that interferes with any aspect of hormone action was defined as endocrine-disrupting chemicals(EDCs).Bisphenol A(BPA),the first known EDC,was synthesized and was considered to be estrogenic.Global production of BPA has increased progressively from 5 to 8 million tons(MT)between 2010 and 2016.Furthermore,researchers estimated that the production should reach 10.2 MT by 2022.The human population is exposed to EDCs in daily life in such forms as pesticides/herbicides,industrial and household products,plastics,detergents,and personal care products.The term obesogen was used for chemicals that promote weight gain and obesity by increasing the number of adipocytes and fat storage in existing adipocytes,changing the energy balance,and finally regulating appetite and satiety.Besides the obesogenic effect,EDCs can cause T2DM through alteration inßcell function and morphology and insulin resistance.In this review,we provide clinical and mechanistic evidence regarding EDCs as obesogen and diabetogen.However,those studies are not enough methodologically to indicate causality.In this respect,randomized clinical trials are needed to investigate the association between obesogen,diabetogen and the related metabolic clinical picture.

Reduced Life Expectancy Model Analyses of Exposure Time Effects of Endocrine Disruptors to Teleost Fishes Based on Effect Concentration of Hepatic Biomarkers

In this current paper, the exposure time effects on four endocrine disruptors and teleost fishes were evaluated using the reduced life expectancy (RLE) model based on the effect concentration (EC50) of available literature published. The result on the regression analysis over different exposure times has demonstrated that the EC50 of hepatic biomarkers falls with increasing exposure times in a predictable manner. The slopes of the regression equations reflect the strength of the toxic effects on the various teleost fish. The EC50 reduction over time can be interpreted based on the bioconcentration process, which can be used to understand transfer routes of the compounds from water to fish body. RLE model also provides useful information in assessing the toxic effects on fish life expectancy as a result of the occurrence of compounds.

Human exposure to a mixture of endocrine disruptors and serum levels of thyroid hormones:A cross-sectional study

Exposure to endocrine disruptors(EDCs) could disrupt thyroid hormone homeostasis. However, human epidemiological studies reported inconsistent observations, and scarce information on the effect of a mixture of chemicals. The aim of the present study was to examine the associations of multiple chemicals with thyroid hormones among adults from China. We measured serum levels of thyroid hormones and urinary levels of 11 EDCs, including six phthalate metabolites, bisphenol A(BPA), bisphenol F(BPF), bisphenol S(BPS), perchlorate, and thiocyanate among 177 healthy adults without occupational exposure. Associations of multiple urinary analytes with serum thyroid hormones were examined by performing general linear regression analysis and bayesian kernal machine regression analysis. These EDCs were detected in almost all samples. After adjusting for various covariates, we observed only BPF significantly associated with total thyroxin(TT4)(β=-0.27, 95% confidence interval(CI) [-0.41,-0.14]), total triiodothyronine(TT3)(β=-0.02 95% CI [-0.03,-0.01]), free T4(fT4)(β=-0.02, 95% CI [-0.03,-0.01]), and free T3(fT3)(β=-0.04, 95% CI [-0.07,-0.01]), and mono-(2-ethyl-5-oxohexyl) phthalate(MEOHP) and monoethyl phthalate(MEP) positively associated with TT4(β=0.24, 95% CI [0.01, 0.48]) and fT4(β=0.02, 95% CI [0.01, 0.04]), respectively. Moreover, we observed significant dose-response relationships between TT4 and the mixture of11 EDCs, and BPF was the main contributor to the mixture effect, suggesting the priority of potential effect of BPF on disrupting thyroid function under a real scenario of human exposure to multiple EDCs. Our findings supported the hypothesis that human exposure to low levels of EDCs could alter thyroid hormones homeostasis among non-occupational healthy adults.

Degradation of endocrine disruptor bisphenol A in drinking water by ozone oxidation

The ozone oxidation of endocrine disruptor bisphenol A in drinking water was investigated.A stainless completely mixed reactor was employed to carry out the degradation experiments by means of a batch model.With an initial concentration of 11.0 mg/L,the removal efficiencies of BPA(bisphenol A)could be measured up to 70%,82%,and 90%when the dosages of ozone were 1,1.5,and 2 mg/L,respectively.The impacts on BPA degradation under the con-ditions of different ozone dosages,water background values,BPA initial concentrations,and ozone adding time were ana-lyzed.The results showed that ozone dosage plays a dominant role during the process of BPA degradation,while the impact of the contact time could be ignored.UV wavelength scan-ning was used to confirm that the by-products were produced,which could be absorbed at UV254.The value of UV254 was observed to have changed during the ozonation process.Based on the change of UV254,it could be concluded that BPA is not completely degraded at low ozone dosage,while shorter adding time of total ozone dosage,high ozone dosage,and improvement of dissolved ozone concentration greatly contribute to the extent of BPA degradation.The effects of applied H_(2)O_(2) dose in ozone oxidation of BPA were also examined in this study.The O_(3)-H_(2)O_(2) processes proved to have similar effects on the degradation of BPA by ozone oxidation.

Artificial intelligence to link environmental endocrine disruptors(EEDs)with bone diseases

Our endocrine system is not only complex,but is also enormously sensitive to the imbalances caused by the environmental stressors,extreme weather situation,and other geographical factors.The endocrine disruptions are associated with the bone diseases.Osteoporosis is a bone disorder that occurs when bone mineral density and bone mass decrease.It affects women and men of all races and ethnic groups,causing bone weakness and the risk of fractures.Environmental stresses are referred to physical,chemical,and biological factors that can impact species productivity.This research aims to examine the impact of environmental stresses on bone diseases like osteoporosis and low bone mass(LBM)in the United States(US).For this purpose,we use an artificial neural network model to evaluate the correlation between the data.A multilayer neural network model is constructed using the Levenberg–Marquardt training algorithm,and its performance is evaluated by mean absolute error and coefficient of correlation.The data of osteoporosis and LBM cases in the US are divided into three groups,including gender group,age group,and race/ethnicity group.Each group shows a positive correlation with environmental stresses and thus the endocrinology.

Endocrine disruptors in teleosts:Evaluating environmental risks and biomarkers

Endocrine disruptors(EDs)are synthetic or natural chemical molecules occurring in environment that have the potential to impart adverse effects on homeostasis of endocrine axis leading to neurological,developmental,immunological and reproductive disarray at organismal level.A wide range of structurally diverse EDs such as,sex-steroid hormone mimics,pesticides and fertilizers,prevail in the environment originating from waste of industries,pharmaceutics,sewage treatment plants and agriculture.In addition,some metals,such as Cu,Hg and Zn,have endocrine disrupting potency in their metallic as well as synthesized nano-particulate forms.There is an increasing concern in research for the plausible threat posed by EDs that can disrupt the endocrine system in aquatic fauna as these compounds are frequently discharged or run-off into water stream.Fishes are well known bio-indicators to understand toxicity of EDs as they are vulnerable to endocrine disruption.Furthermore,EDs have the potential to affect fish-feeding higher vertebrates including mammals and subsequently human,as they make their way up on the food web pyramid due to biomagnification.In light of this,several observations suggesting adverse effects of EDs and the mechanism contributing to endocrine disruption in fish are discussed extensively in this review.This article highlights the necessity to choose a credible model for assessing the toxic effects exerted by EDs.Furthermore,the toxic effects of EDs will be comprehensively reviewed with reference to sexual plasticity,neuroendocrine mechanisms,thyroid and immune modulation,gonadal development and maturation as well as changes in transcriptome/genome profile using fish models to imply ED-induced aquatic pollution in a larger perspective.For decades now,studies on EDs have challenged traditional concepts in toxicology to develop new molecular markers to improve methodologies and to assess the ecological risks associated with field conditions.In this regard,it is imperative to highlight the development of modern diagnostic tools including biosensors to monitor the inadvertent usage of EDs and the resultant environmental risks.Lastly,current limitations in knowledge along with future research perspectives in the field are also highlighted in this article.

Simultaneous Determination of Ten Endocrine Hormone Disrupters in Water Using SPE/LC-(ESI)MS-MS

The aim of this study was developed and validated an analytical method based on liquid chromatography and tandem mass spectrometry after solid phase extraction to monitorizing ten endocrine hormone disrupters in Lisbon drinking water system. Natural and synthetic hormones (17-β-estradiol, ethinylestradiol, estriol, estrone, progesterone, mestranol and diethylstilbestrol) and some industrial products (4-n-nonylphenol, 4-tert-octylphenol and bisphenol A) were studied. Mass spectrometer detection parameters were optimized, such as the best conditions for the precursor ion formation, namely cone voltage, when applying negative and positive electrospray ionization, and also collision energy for MRM1 and MRM2 transitions. The best conditions of the solid phase extraction (SPE) using Waters Oasis HLB (6 mL, 200 mg) and Isolute C18 (EC) (6 ml, 1000 mg) were also optimized. The method was validated through the application of several statistical tests and the uncertainty estimation of the analytical assay. This method showed a very good linear range for all the studied analytes with determination coefficients (r2) between 0.9962 and 0.9999 and coefficients of variation lower than 4%. There were no significant differences between recoveries obtained with the studied matrices, like groundwater, surface water and water for human consumption. In these matrices, the recovery values varied between 32 and 95%. The limits of method detection were between 0.28 and 22 ng/L. The validated method was applied for the analysis of water samples from the EPAL (Empresa Portuguesa das águas Livres, S.A.) water supply system including tap water, spring water, groundwater, and river water. Some target compounds (bisphenol A, progesterone, 4-tert-octylphenol, and 4-n-nonylphenol) were found in trace amounts in analysed waters.

Integrative rodent models for assessing male reproductive toxicity of environmental endocrine active substances

In the present review, we first summarize the main benefits, limitations and pitfalls of conventional in vivo approaches to assessing male reproductive structures and functions in rodents in cases of endocrine active substance （EAS） exposure from the postulate that they may provide data that can be extrapolated to humans. Then, we briefly present some integrated approaches in rodents we have recently developed at the organism level. We particularly focus on the possible effects and modes of action （MOA） of these substances at low doses and in mixtures, real-life conditions and at the organ level, deciphering the precise effects and MOA on the fetal testis. It can be considered that the in vivo experimental EAS exposure of rodents remains the first choice for studies and is a necessary tool （together with the epidemiological approach） for understanding the reproductive effects and MOA of EASs, provided the pitfalls and limitations of the rodent models are known and considered. We also provide some evidence that classical rodent models may be refined for studying the multiple consequences of EAS exposure, not only on the reproductive axis but also on various hormonally regulated organs and tissues, among which several are implicated in the complex process of mammalian reproduction. Such models constitute an interesting way of approaching human exposure conditions. Finally, we show that organotypic culture models are powerful complementary tools, especially when focusing on the MOA. All these approaches have contributed in a combinatorial manner to a better understanding of the impact of EAS exposure on human reproduction.

Plantain-based diet modulates atrazine-induced testicular toxicities in rats

Objective:To assess the potential of plantain-based diet in modulating testicular toxicities in rats exposed to atrazine.Methods:The plantain-based diet at 50%,25% and 12.5% were prepared from the basal diet by substituting the corn starch with plantain fruit pulp flour at different percentages.Wistar rats were fed plantain-based diet in varying concentrations ranging from 12.5% to 50% of the basal diet for 21 days before or after atrazine treatment in a two-phase experiment:preventive and therapeutic phases.The therapeutic model(n=35)had seven groups with 5 rats each,including the control,atrazine,atrazine recovery,atrazine plus plantain-based diet 50%,25%,12.5%,and atrazine plus quercetin groups.The preventive model(n=30)had six groups of 5 rats,consisting of the control,atrazine,50%,25%,12.5% plantain-based diet plus atrazine,and quercetin plus atrazine groups.Gonadal hormones(testosterone,luteinizing hormone and follicle-stimulating hormone),sperm parameters(sperm motility,viability,morphology and concentration),and testicular function indices(protein,cholesterol,glycogen,acid phosphatase,alkaline phosphatase and lactate dehydrogenase)were measured.Results:The gonadal hormones,sperm characteristics,and testicular function indices of the rat testis decreased significantly in the atrazine group alongside degeneration of the histoarchitecture.However,plantain-based diet restored the gonadal hormone concentrations,semen parameters,and testicular function indices in both the preventive and therapeutic models.Conclusions:Treatment with plantain-based diet protects against rat testicular toxicity caused by atrazine via the modulation of gonadal hormones,sperm quality,testicular function index as well as histoarchitecture of rat testes.

Bisphenol A (BPA) Contamination in Whole Milk Marketed in Different Packages

Bisphenol A (BPA), an important endocrine disruptor, is used in the manufacturing of various materials, including food packaging. Ingestion of contaminated foodstuffs is, in fact, the most relevant form of exposure to this substance. However, scarce data on the presence of this contaminant in milk, or whether different types of food packaging influence food contamination are available in Brazil. This study, therefore, aimed to evaluate the BPA contamination of whole milk (fluid and powder) samples packaged in different types of packaging (Tetra Pak?;PET: Poly (ethylene terephthalate;Metallic can (epoxy resin);Polyethylene (PE) and poly (vinylidene chloride) (PVDC);Laminated Film - Metallized Polyester-Polyethylene and glass) and marketed metropolitan region of Rio de Janeiro, Brazil. An analytical method for the BPA determination in milk was optimized for both fluid (pasteurized and ultra-high temperature) and powdered milk samples. A modified QuEChERS method was applied, and BPA determinations were conducted by ultra-performance liquid chromatography coupled with sequential mass spectrometry (HPLC-MS/MS). The validated method was then applied to 51 milk samples, where BPA was detected in five samples (9.8%) and quantified in two (3.8%).

Phthalate Exposure and Human Semen Quality in Shanghai:A Cross-sectional Study

Objective To monitor the level of phthalates in human semen samples and to analyze the relationship between phthalate levels and semen parameters. Methods Concentrations of three kinds of commonly used phthalates （di-ethyl phthalate, DEP; di-n-butyl phthalate, DBP; di-2-ethylhexyl phthalate, DEHP） were measured using reversed-phase HPLC. Semen parameters were measured by computer aided sperm analysis （CASA）. Results The three phthalates were detected in most of the biological samples, with median levels of 0.30 mg/L （0.08-1.32 mg/L） in semen specimens. There was a significant positive association between liquefied time of semen and phthalate concentrations of semen. The correlation coefficient was 0.456 for DEP, 0.475 for DBP, and 0.457 for DEHP, respectively. There was no significant difference between phthalate concentrations of semen and sperm density or livability, though the correlation coefficients were negative. Conclusion These results suggest that people who reside in Shanghai are exposed to phthalates, especially to DBP and DEHP. Although the level of phthalates is relatively mild, an association of phthalate levels and reduced quality of human semen has been shown in the present study.

Spatial distribution and deposition history of nonylphenol and bisphenol A in sediments from the Changjiang River(Yangtze River) Estuary and its adjacent East China Sea

Nonylphenol（NP） and bisphenol A（BPA） are endocrine disruptors causing harmful effects including feminization and carcinogenesis to various organisms,and consequently,their contamination in natural environment has received wide concerns.This study reports the distribution characteristics of NP and BPA in surface sediments and their deposition history based on a dated sediment core in the Changjiang River（Yangtze River） Estuary and its adjacent East China Sea.The contents of NP and BPA in surface sediments ranged from 1.56-35.8 and 0.72-13.2 ng/g（dry mass）,respectively,with high values recorded in the two mud zones,the Changjiang River Estuarine Mud Zone and the Zhejiang Coastal Mud Zone.High values in the Zhejiang Coastal Mud Zone suggest the possibility of long distance transport of both contaminants through the Changjiang riverine plume.The contents were not correlated with the distance from the pollution source,indicating other factors including particle deposition rate and sediment grain size obviously affecting the distribution pattern.NP was also detected in a sediment core at layers deposited from the year of 1971 to 2001 with contents of up to 20.9 ng/g（dry mass）.The deposition fluxes of NP varied from 0.68 to 17.9 ng/（cm^2 · a） with peaks and valleys reflecting the traces of economic development history in China during the previous three decades.BPA was detected at sediment layers deposited from 1973 to 2001 with contents of up to 3.66 ng/g.The fluxes of BPA varied from 0.62 to 3.13 ng/（cm^2 · a） showing a similar pattern as NP.The contents of NP and BPA also indicated potential risks on benthic organisms in the study area.

Sorption of biodegradation end products of nonylphenol polyethoxylates onto activated sludge

Nonylphenol(NP), nonylphenoxy acetic acid(NP1EC), nonylphenol monoethoxy acetic acid(NP2EC), nonylphenol monoethoxylate(NP1EO) and nonylphenol diethoxylate(NP2EO) are biodegradation end products(BEPs) of nonionic surfactant nonylphenolpolyethoxylates (NPnEO). In this research, sorption of these compounds onto model activated sludge was characterized. Sorption equilibrium experiments showed that NP, NP1EO and NP2EO reached equilibrium in about 12 h, while equilibrium of NP1EC and NP2EC were reached earlier, in about 4 h. In sorption isotherm experiments, obtained equilibrium data at 28℃ fitted well to Freundlich sorption model for all investigated compounds. For NP1EC, in addition to Freundlich, equilibrium data also fitted well to Langmuir model. Linear sorption model was also tried, and equilibrium data of all NP, NP1EO, NP2EO and NP2EC except NP1EC fitted well to this model. Calculated Freundlich coefficient(K F) and linear sorption coefficient(K D) showed that sorption capacity of the investigated compounds were in order NP>NP2EO>NP1EO>NP1EC≈NP2EC. For NP, NP1EO and NP2EO, high values of calculated K F and K D indicated an easy uptake of these compounds from aqueous phase onto activated sludge. Whereas, NP1EC and NP2EC with low values of K F and K D absorbed weakly to activated sludge and tended to preferably remain in aqueous phase.

Gracilaria changii(Rhodophyta) alleviates bisphenol A-induced adverse reproductive abnormalities in mice

Objective:To evaluate the potential of Gracilaria changii extract in ameliorating the potential adverse effects of bisphenol A.Methods:The antioxidant capacity of Gracilaria changii extracted using different solvents(methanol,ethanol,and aqueous)was studied.The mice were administered by oral gavage with bisphenol A(60 mg/kg body weight)for 6 weeks with or without Gracilaria changii aqueous extract.Thereafter,the mice were either euthanized for histology and immunohistochemistry studies or mated to evaluate the pregnancy rate.Results:Gracilaria changii aqueous extract showed the highest antioxidant properties compared with extract using methanol and ethanol.The aqueous extract of Gracilaria changii improved the uterus index and uterine lipid peroxidation after bisphenol A exposure,although the uterine expressions of estrogen receptors and complement C3 were not improved.Histological evaluation of the uterus during the estrus stage has revealed that the extract could mitigate bisphenol A-induced adverse effects in the uterus as there was a lower percentage of mice showing abnormalities like decreased eosin staining in the myometrium,and decrease in the number of eosinophil and endometrial glands in the endometrium.Besides,Gracilaria changii aqueous extract improved the pregnancy rate of mice administered with bisphenol A.Conclusions:Gracilaria changii extract protects against bisphenol A-induced female reproductive abnormalities in mice which may be mediated via modulation of eosinophil migration,endometrial gland formation,and protein expressions associated with prostaglandins in the myometrium.

Potential genomic biomarkers of obesity and its comorbidities for phthalates and bisphenol A mixture:In silico toxicogenomic approach

This in silico toxicogenomic study aims to explore the relationship between phthalates and bisphenol A(BPA)co-exposure and obesity,as well as its comorbid conditions,in order to construct a possible set of genomic biomarkers.The Comparative Toxicogenomics Database(CTD;http://ctd.mdibl.org)was used as the main data mining tool,along with GeneMania(https://genemania.org),ToppGene Suite(https://toppgene.cchmc.org)and DisGeNET(http://www.disgenet.org).Among the phthalates,bis(2-ethylhexyl)phthalate(DEHP)and dibutyl phthalate(DBP)were chosen as the most frequently curated phthalates in CTD,which also share similar mechanisms of toxicity.DEHP,DBP and BPA interacted with 84,90 and 194 obesity-related genes/proteins,involved in 67,65 and 116 pathways,respectively.Among these,53 genes/proteins and 42 pathways were common to all three substances.31 genes/proteins had matching interactions for all three investigated substances,while more than half of these genes/proteins(56.49%)were in co-expression.7 of the common genes/proteins(6 relevant to humans:CCL2,IL6,LPL,PPARG,SERPINE1,and TNF)were identified in all the investigated obesity comorbidities,while PPARG and LPL were most closely linked to obesity.These genes/proteins could serve as a target for further in vitro and in vivo studies of molecular mechanisms of DEHP,DBP and BPA mixture obesogenic properties.Analysis reported here should be applicable to any mixture of environmental chemicals and any disease present in CTD.