Dibutyl phthalate degradation by Paenarthrobacter ureafaciens PB10 through downstream product myristic acid and its bioremediation potential in contaminated soil.

Dibutyl phthalate (DBP) is a widely used plasticizer to make plastic flexible and long-lasting. It is easily accessible in a broad spectrum of environments as a result of the rising level of plastic pollution. This compound is considered a top-priority toxicant and persistent organic pollutant by international environmental agencies for its endocrine disruptive and carcinogenic propensities. To mitigate the DBP in the soil, one DBP-degrading bacterial strain was isolated from a plastic-polluted landfill and identified as Paenarthrobacter ureafaciens PB10 by 16S rRNA gene sequence-based homology. The strain was found to develop a distinct transparent halo zone around grown colonies on an agar plate supplemented with DBP. The addition of yeast extract (100 mg/L) as a nutrient source accelerated cell biomass production and DBP degradation rate; however, the presence of glucose suppressed DBP degradation by the PB10 strain without affecting its ability to proliferate. The strain PB10 was efficient in eliminating DBP under various pH conditions (5.0-8.0). Maximum cell growth and degradation of 99.49% at 300 mg/L DBP were achieved in 72 h at the optimized mineral salt medium (MS) conditions of pH 7.0 and 32 °C. Despite that, when the concentration of DBP rose to 3000 mg/L, the DBP depletion rate was measured at 79.34% in 72 h. Some novel intermediate metabolites, like myristic acid, hexadecanoic acid, stearic acid, and the methyl derivative of 4-hydroxyphenyl acetate, along with monobutyl phthalate and phthalic acid, were detected in the downstream degradation process of DBP through GC-MS profiling. Furthermore, in synchronization with native soil microbes, this PB10 strain successfully removed a notable amount of DBP (up to 54.11%) from contaminated soil under microcosm study after 10 d. Thus, PB10 has effective DBP removal ability and is considered a potential candidate for bioremediation in DBP-contaminated sites.

Complete degradation of di-n-butyl phthalate by Glutamicibacter sp. strain 0426 with a novel pathway.

Di-n-butyl phthalate (DBP) is widely used as plasticizer that has potential carcinogenic, teratogenic, and endocrine effects. In the present study, an efficient DBP-degrading bacterial strain 0426 was isolated and identified as a Glutamicibacter sp. strain 0426. It can utilize DBP as the sole source of carbon and energy and completely degraded 300 mg/L of DBP within 12 h. The optimal conditions (pH 6.9 and 31.7 °C) for DBP degradation were determined by response surface methodology and DBP degradation well fitted with the first-order kinetics. Bioaugmentation of contaminated soil with strain 0426 enhanced DBP (1 mg/g soil) degradation, indicating the application potential of strain 0426 for environment DBP removal. Strain 0426 harbors a distinctive DBP hydrolysis mechanism with two parallel benzoate metabolic pathways, which may account for the remarkable performance of DBP degradation. Sequences alignment has shown that an alpha/beta fold hydrolase (WP_083586847.1) contained a conserved catalytic triad and pentapeptide motif (GX1SX2G), of which function is similar to phthalic acid ester (PAEs) hydrolases and lipases that can efficiently catalyze hydrolysis of water-insoluble substrates. Furthermore, phthalic acid was converted to benzoate by decarboxylation, which entered into two different pathways: one is the protocatechuic acid pathway under the role of pca cluster, and the other is the catechol pathway. This study demonstrates a novel DBP degradation pathway, which broadens our understanding of the mechanisms of PAE biodegradation.

Combined exposure to multiwalled carbon nanotubes and dibutyl phthalates aggravated airway inflammation in rats.

Previous work has shown that mice exposed to dibutyl phthalate (DBP) adsorbed onto multi-walled carbon nanotubes (MWCNTs), via tail vein injection, displayed black lesions in their lungs. To investigate the mechanism causing this toxicity in the lung tissue, we performed an experiment with rats, exposing them to DBP adsorbed onto MWCNTs via a tail vein injection for 14 days. The results revealed pulmonary edema and greyish-black lung tissue in the MWCNTs and the MWCNTs + DBP combined exposure groups. In the combined exposure group there was evident alveolar fragmentation and adhesion, and lung tissue sections showed significant levels of black particles. Sections of the non-cartilaginous region of the trachea had significant folding of the pseudostratified ciliated columnar epithelium and marked thickening of the submucosa. In broncho alveolar lavage fluid, the number of leukocytes (WBC), lymphocytes (Lym), neutrophils (Neu), and eosinophils (Eos), as well as levels of immunoglobulin E (IgE), interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-α), and interleukin 1ß (IL-1ß) were all significantly higher. TNF-α, IL-6, signal transducer and activator of transcription 3 (STAT3), and α-smooth muscle actin (α-SMA) mRNA expression were all elevated in the lung tissue. The combined exposure group, which had considerable airway remodeling, had a greater degree of tracheal constriction and luminal narrowing, according to the results of the α-SMA immunofluorescence assay. According to these experimental findings, the exposure to both MWCNTs and DBP seemed to have a synergistic effect and exacerbated rats' impaired respiratory function that resulted from exposure to MWCNTs alone.

Enhanced removal of dibutyl phthalate in a laccase-mediator system: Optimized process parameters, kinetics, and environmental impact.

The persistence and recalcitrance of endocrine-disrupting chemicals (EDCs) in the environment have raised momentous concerns due to their carcinogenic, teratogenic, genotoxic, and cytotoxic effects on humans, animals, and plants. Unarguably, dibutyl phthalate (DBP) is one of the most ubiquitous EDCs because of its bioavailability in water, soil, and atmosphere. This study aims to investigate the efficiency of Agaricus bisporus laccase in the degradation of dibutyl phthalate (DBP) in laccase-mediator system. Here, enhanced removal efficiency was recorded during DBP degradation in laccase-mediator systems than in reaction medium containing laccase only. About 98.85% of 30 mg L-1 DBP was efficiently removed in a medium containing 1.3 U mL-1, 0.045 mM Syringaldehyde (SYR) at incubation temperature 30 aC and pH 5 within 24 h. This finding was further corroborated by the synergistic interplay of the optimal parameters in the laccase-SYR system done using response surface methodology (Box-Behnken Design). Furthermore, the addition of 1.5 mM of metal ions in the laccase-SYR system further promoted the enhanced removal of DBP in the following order: Cr3+> Pb2+> Ca2+> Al3+>Zn2+ > Cu2+. A significant decrease in DBP degradation was observed at higher concentrations of metal ions above 1.5 mM due to the inhibition of laccase active sites. The coefficient of correlation (R2 = 0.9885) recorded in the Lineweaver bulk plot affirmed that the removal efficiencies are highly dependent on DBP concentration in the laccase-SYR system. The Gas-Chromatography Mass Spectrometry (GC-MS) analyses affirmed that the ortho-cleavage due to hydrolysis of DBP in the reaction system led to the formation of two metabolic degradation products (MBP and PA). The phytotoxicity assessment affirmed the detoxified status of DBP after treatment with significant improvement (90 and 91%) in the growth of Lens culinaris and Sorghum bicolor. This is the first report on DBP degradation in the laccase-SYR reaction system, underscoring the unique, eco-friendly, economical, and promising alternative to known conventional methods.

Combined cytotoxicity of phthalate esters on HepG2 cells: A comprehensive analysis of transcriptomics and metabolomics.

Phthalate esters (PAEs), widely used as plasticizers, may pose a potential environmental and human hazard. The aim of this study was to compare the cytotoxicity of di(2-ethylhexyl) phthalates (DEHP) and dibutyl phthalate (DBP)) after their exposure to HepG2 cells alone or in combination. HepG2 cells treated with individual/combined DEHP and DBP at a dose of 10-2 M for 24 h were selected for metabolome and transcriptome analysis. The results demonstrated that exposure to the mixtures of DEHP and DBP caused enhanced or reduced toxic effects regarding 8 pathways with 1065 downregulated genes and 643 upregulated genes, in comparison with those of single chemicals. The combined toxicity of mixture revealed both synergistic and antagonistic interactions between DEHP and DBP. Besides, combined exposure to DEHP and DBP promoted TCA cycle, pyrimidine, and purine metabolism, while an antagonistic effect on fatty acid derangement should require further investigation. To summarize, our results suggest that DEHP exposed alone or combined with DBP caused a variety of metabolic disorders, and the type of combination effects varied among metabolic pathways.

Gestational dibutyl phthalate exposure impairs primordial folliculogenesis in mice through autophagy activation and NOTCH2 signal interruption.

Female reproductive lifespan is largely determined by the size of the primordial follicle pool, which is established in early life. Dibutyl phthalate (DBP), a popular plasticiser, is a known environmental endocrine disruptor that poses a potential threat to reproductive health. However, DBP impact on early oogenesis has been rarely reported. In this study, maternal exposure to DBP in gestation disrupted germ-cell cyst breakdown and primordial follicle assembly in foetal ovary, impairing female fertility in adulthood. Subsequently, altered autophagic flux with autophagosome accumulation was observed in DBP-exposed ovaries carrying CAG-RFP-EGFP-LC3 reporter genes, whereas autophagy inhibition by 3-methyladenine attenuated the impact of DBP on primordial folliculogenesis. Moreover, DBP exposure reduced the expression of NOTCH2 intracellular domain (NICD2) and decreased interactions between NICD2 and Beclin-l. NICD2 was observed within the autophagosomes in DBP-exposed ovaries. Furthermore, NICD2 overexpression partially restored primordial folliculogenesis. Furthermore, melatonin significantly relieved oxidative stress, decreased autophagy, and restored NOTCH2 signalling, consequently reversing the effect on folliculogenesis. Therefore, this study demonstrated that gestational DBP exposure disrupts primordial folliculogenesis by inducing autophagy, which targets NOTCH2 signalling, and this impact has long-term consequences on fertility in adulthood, strengthening the potential contribution of environmental chemicals to the development of ovarian dysfunctional diseases.

Interaction between Phthalate Ester and Rice Plants: Novel Transformation Pathways and Metabolic-Network Perturbations.

Our understanding is limited concerning the interaction mechanism between widespread phthalate esters and staple crops, which have strong implications for human exposure. Therefore, this study was aimed at illuminating the transformation pathways of di-n-butyl phthalate (DnBP) in rice using an untargeted screening method. UPLC-QTOF-MS identified 16 intermediate transformation products formed through hydroxylation, hydrolysis, and oxidation in phase I metabolism and further by conjugation with amino acids, glutathione, and carbohydrates in phase II metabolism. Mono-2-hydroxy-n-butyl phthalate-l-aspartic acid (MHBP-asp) and mono-2-hydroxy-n-butyl phthalate-d-alanyl-ß-d-glucoside (MHBP-ala-glu) products were observed for the first time. The proteomic analysis demonstrated that DnBP upregulated the expression of rice proteins associated with transporter activity, antioxidant synthesis, and oxidative stress response and downregulated that of proteins involved in photosynthesis, photorespiration, chlorophyll binding, and mono-oxygenase activity. Molecular docking revealed that DnBP can affect protein molecular activity via pi-sigma, pi-alkyl, and pi-pi interactions or by forming carbon-hydrogen bonds. The metabolomic analysis showed that key metabolic pathways including citrate cycle, biosynthesis of aminoacyl-tRNA, and metabolism of amino acids, sphingolipids, carbohydrates, nucleotides, and glutathione were activated in rice plants exposed to DnBP and its primary metabolite mono-n-butyl phthalate (MnBP). Furthermore, exposure to 80 ng/mL MnBP significantly perturbed the metabolic profile and molecular function in plants, with downregulation of the levels of beta-alanine (0.56-fold), cytosine (0.48-fold), thymine (0.62-fold), uracil (0.48-fold), glucose (0.59-fold), and glucose-1-phosphate (0.33-fold), as well as upregulation of the levels of l-glutamic acid (2.97-fold), l-cystine (2.69-fold), and phytosphingosine (38.38-fold). Therefore, the degradation intermediates of DnBP pose a potentially risk to plant metabolism and raise concerns for crop safety related to plasticizer pollution.

Combined exposure to benzo(a)pyrene and dibutyl phthalate aggravates pro-inflammatory macrophage polarization in spleen via pyroptosis involving cathepsin B.

Humans are often simultaneously exposed to benzo(a)pyrene (BaP) and dibutyl phthalate (DBP) through consumption of food and water. Yet, direct evidence of the link between BaP and DBP co-exposure and the risk of splenic injury is lacking. In the present study, we established the rats and primary splenic macrophages models to evaluate the effects of BaP or/and DBP exposure on spleen and underlying mechanisms. Compared to the single exposure or control groups, the co-exposure group showed more severe spleen damage and higher production of pro-inflammatory cytokines. Co-exposure to BaP and DBP resulted in a 1.79-fold, 2.11-fold and 1.9-fold increase in the M1 macrophage markers iNOS, NLRP3 (pyroptosis marker protein) and cathepsin B (CTSB), respectively, and a 0.8-fold decrease in the M2 macrophage marker Arg1 in vivo. The more prominent effects in perturbation of imbalance in M1/M2 polarization (iNOS, 2.25-fold; Arg1, 0.55-fold), pyroptosis (NLRP3, 1.43-fold), and excess CTSB (1.07-fold) in macrophages caused by BaP and DBP co-exposure in vitro were also found. Notably, MCC950 (the NLRP3-specific inhibitor) treatment attenuated the pro-inflammatory macrophage polarization and following pro-inflammatory cytokine production triggered by BaP and DBP co-exposure. Furthermore, CA-074Me (the CTSB-specific inhibitor) suppressed the macrophages pyroptosis, pro-inflammatory macrophage polarization, and secretion of pro-inflammatory cytokine induced by BaP and DBP co-exposure. In conclusion, this study indicates co-exposure to BaP and DBP poses a higher risk of spleen injury. Pro-inflammatory macrophage polarization regulated by pyroptosis involving CTSB underlies the spleen injury caused by BaP and DBP co-exposure.

Effects of preconception exposure to phthalates on mouse sperm capacitation parameters.

BACKGROUND: Phthalates have been linked to adverse male reproductive health, including poor sperm quality and embryo quality as well as a longer time to pregnancy (months of unprotected intercourse before conception occurs). The present study aimed to evaluate the effect of preconception exposure to two ubiquitous phthalate chemicals, di(2-ethylhexyl) phthalate (DEHP), di-n-butyl phthalate (DBP), and their mixture on sperm function, fertilization, and embryo development in mice. MATERIALS AND METHODS: Adult male C57BL/6J mice aged 8-9 weeks were exposed to di(2-ethylhexyl) phthalate, di-n-butyl phthalate, or their mixture (di-n-butyl phthalate + di(2-ethylhexyl) phthalate) at 2.5 mg/kg/day or vehicle for 40 days (equivalent to one spermatogenic cycle) via surgically implanted osmotic pumps. Caudal epididymal spermatozoa were extracted and analyzed for motility using computer-assisted sperm analyses. Sperm phosphorylation of protein kinase A substrates and tyrosine phosphorylation, markers of early and late capacitation events, respectively, were analyzed by Western blots. In vitro fertilization was used to evaluate the sperm fertilizing capacity. RESULTS: While the study did not reveal any significant differences in sperm motility and fertilization potential, abnormal sperm morphology was observed in all phthalate exposures, particularly in the phthalate mixture group. In addition, the study revealed significant differences in sperm concentration between control and exposed groups. Moreover, protein phosphorylation of protein kinase A substrates was decreased in the di(2-ethylhexyl) phthalate and mixture exposure groups, while no significant changes in protein tyrosine phosphorylation were observed in any of the groups. Assessment of the reproductive functionality did not reveal significant effects on in vitro fertilization and early embryo development rates but showed wide variability in the phthalate mixture group. CONCLUSION: Our findings suggest that preconception phthalate exposure affects sperm numbers and phosphorylation of protein kinase A substrates involved in capacitation. Future research is warranted to examine the associations between phthalate exposure and capacitation in human spermatozoa.

Persistent immunosuppressive effects of dibutyl phthalate exposure in adult male mice.

Increased exposure to manmade chemicals may be linked to an increase in immune-related diseases in humans and immune system dysfunction in wildlife. Phthalates are a group of endocrine-disrupting chemicals (EDCs) suspected to influence the immune system. The aim of this study was to characterize the persistent effects on leukocytes in the blood and spleen, as well as plasma cytokine and growth factor levels, one week after the end of five weeks of oral treatment with dibutyl phthalate (DBP; 10 or 100 mg/kg/d) in adult male mice. Flow cytometry analysis of the blood revealed that DBP exposure decreased the total leukocyte count, classical monocyte and T helper (Th) populations, whereas it increased the non-classical monocyte population compared to the vehicle control (corn oil). Immunofluorescence analysis of the spleen showed increased CD11b+Ly6G+ (marker of polymorphonuclear myeloid-derived suppressor cells; PMN-MDSCs), and CD43+staining (marker of non-classical monocytes), whereas CD3+ (marker of total T cells) and CD4+ (marker of Th cells) staining decreased. To investigate the mechanisms of action, levels of plasma cytokines and chemokines were measured using multiplexed immunoassays and other key factors were analyzed using western blotting. The observed increase in M-CSF levels and the activation of STAT3 may promote PMN-MDSC expansion and activity. Increased ARG1, NOX2 (gp91phox), and protein nitrotyrosine levels, as well as GCN2 and phosphor-eIRFα, suggest that oxidative stress and lymphocyte arrest drive the lymphocyte suppression caused by PMN-MDSCs. The plasma levels of IL-21 (promotes the differentiation of Th cells) and MCP-1 (regulates migration and infiltration of monocytes/macrophages) also decreased. These findings show that adult DBP exposure can cause persistent immunosuppressive effects, which may increase susceptibility to infections, cancers, and immune diseases, and decrease vaccine efficacy.

Fluorescent labeling and tracing of immobilized efficient degrading bacterium DNB-S1 and its remediation efficiency of DBP contaminated soil.

Dibutyl phthalate (DBP) is an organic pollutant frequently detected in soil, and is a reproductive poison that harms animals both before and after birth and has mutagenic, teratogenic, and carcinogenic effects. DBP removal from farmland has been the subject of extensive research in recent years. Efficient DBP degrading bacterial strains were screened in the laboratory. GFP (Green fluorescent protein) labeled degradation strain GFP-DNB-S1 was analyzed for its activity and dynamics. Using sodium alginate (SA) and nano-hydroxyapatite (n-HAP) as carrier materials and CaCl2 as a cross-linking agent, the immobilized microbial agent n-HAP/SA + DNB-S1 was prepared by embedding cross-linking immobilization technology to study the remediation effect of DBP contaminated soil. The best formation effect of immobilized materials (n-HAP/SA) was found when the SA to n-HAP ratio was 3:2. When compared to single SA immobilized bacteria, n-HAP/SA immobilized bacteria improved the surface roughness and porosity of the microspheres. After 70 days, LED light revealed that the immobilized bacteria's GFP green fluorescent protein expression was stable. At 70 days, the initial DBP concentration of 500 mg ∙ L-1 degraded at a rate of 69.9%. The degrading bacteria had no effect on DBP degradation before and after being labeled with GFP. The n-HAP/SA immobilized bacteria offered a better living environment for microorganisms due to their rougher surface and a greater number of pores. This protected the microorganisms and increased the efficiency of DBP degradation. When the concentration of DBP in contaminated soil was set to 20 mg ∙ kg-1 and the n-HAP/SA + DNB-S1 immobilized bacterial agent was applied to the soil, the rate of DBP degradation was determined to be 93.34%. The degradation process followed First-order degradation kinetics, which improved the physical and chemical properties of the soil as well as its fertility.

Phthalate plasticizer decreases the prion-like protein doppel essential for structural integrity and function of spermatozoa.

Di-n-butyl phthalate (DBP), a well-known endocrine disruptor, causes male reproductive dysfunction. To understand the underlying mechanisms, we performed histological, endocrinological, and biochemical analyses and assessed the expression of genes involved in spermatogenesis and sperm function according to OECD test guideline 407. Following 28 days of administration of the lowest observed adverse effect level dose of DBP to mice, no significant changes in body weight, testis and epididymis weights and histology, serum testosterone level, or testicular daily sperm production were found. Nonetheless, the motility of the epididymal sperm of the DBP group was significantly decreased together with an increase in the incidence of bent tails and abnormal heads. In the testes of the DBP group, lipid peroxidation (LPO) level was significantly increased and testicular Bcl-2 mRNA level was significantly decreased together with an increase in the Bax/Bcl-2 mRNA ratio. In the testes of the DBP group, levels of Prnd mRNA and protein and Pou4f1 mRNA, an activator of the Prnd promotor, were significantly decreased. Of note, prion-like protein doppel (PRND) was significantly decreased together with decreased PRND immunoreactivity in the head, midpiece, and tail of sperm. In the testes of the DBP group, levels of Sox9, Sgp1, and Sgp2 mRNA, which are functional Sertoli cell markers, were significantly decreased. Level of Amh mRNA, a Sertoli cell immaturity marker, was significantly increased together with that of Inha mRNA, suggesting deregulation of the brain-gonadal axis. Together, our findings suggest that DBP at present dosage may potentiate LPO generation and Sertoli cell immaturity via downregulation of Sox9 and disruption of the Pou4f1-Prnd gene network in post-meiotic germ cells without visible changes in spermatogenesis or testosterone level. This may result in structural and functional abnormalities in spermatozoa. Additionally, our findings suggest that assessment of the male reproductive toxicity of phthalate ester plasticizers based on conventional OECD test guidelines should be reconsidered.

Microbial interactions enhanced environmental fitness and expanded ecological niches under dibutyl phthalate and cadmium co-contamination.

Co-contamination of organic pollutants and heavy metals is universal in the natural environment. Dibutyl phthalate (DBP), a typical plasticizer, frequently coexists with cadmium (Cd) in nature. However, little attention has been given to the impacts of co-contamination by DBP and Cd on microbial communities or the responses of microbes. To address this, a microcosm experiment was conducted by supplying the exogenous DBP-degrading bacterium Glutamicibacter nicotianae ZM05 to investigate the interplay among DBP-Cd co-contamination, the exogenous DBP-degrading bacterium G. nicotianae ZM05, and indigenous microorganisms. To adapt to co-contamination stress, microbial communities adjust their diversity, interactions, and functions. The stability of the microbial community decreased under co-contamination, as evidenced by lower diversity, simpler network, and fewer ecological niches. Microbial interactions were strengthened, as evidenced by enriched pathways related to microbial communications. Meanwhile, interactions between microorganisms enhanced the environmental fitness of the exogenous DBP-degrading bacterium ZM05. Based on co-occurrence network prediction and coculture experiments, metabolic interactions between the non-DBP-degrading bacterium Cupriavidus metallidurans ZM16 and ZM05 were proven. Strain ZM16 utilized protocatechuic acid, a DBP downstream metabolite, to relieve acid inhibition and adsorbed Cd to relieve toxic stress. These findings help to explain the responses of bacterial and fungal communities to DBP-Cd co-contamination and provide new insights for the construction of degrading consortia for bioremediation.

Degradation of dibutyl phthalate by Paenarthrobacter sp. Shss isolated from Saravan landfill, Hyrcanian Forests, Iran.

Phthalic acid esters are predominantly used as plasticizers and are industrially produced on the million ton scale per year. They exhibit endocrine-disrupting, carcinogenic, teratogenic, and mutagenic effects on wildlife and humans. For this reason, biodegradation, the major process of phthalic acid ester elimination from the environment, is of global importance. Here, we studied bacterial phthalic acid ester degradation at Saravan landfill in Hyrcanian Forests, Iran, an active disposal site with 800 tons of solid waste input per day. A di-n-butyl phthalate degrading enrichment culture was established from which Paenarthrobacter sp. strain Shss was isolated. This strain efficiently degraded 1 g L-1 di-n-butyl phthalate within 15 h with a doubling time of 5 h. In addition, dimethyl phthalate, diethyl phthalate, mono butyl phthalate, and phthalic acid where degraded to CO2, whereas diethyl hexyl phthalate did not serve as a substrate. During the biodegradation of di-n-butyl phthalate, mono-n-butyl phthalate was identified in culture supernatants by ultra-performance liquid chromatography coupled to electrospray ionization quadrupole time-of-flight mass spectrometry. In vitro assays identified two cellular esterase activities that converted di-n-butyl phthalate to mono-n-butyl phthalate, and the latter to phthalic acid, respectively. Our findings identified Paenarthrobacter sp. Shss amongst the most efficient phthalic acid esters degrading bacteria known, that possibly plays an important role in di-n-butyl phthalate elimination at a highly phthalic acid esters contaminated landfill.

Prenatal exposure to di-n-butyl phthalate induces erectile dysfunction in male adult rats.

Di-n-butyl phthalate (DBP) is a widely used plasticizer and an environmental endocrine-disrupting compound. However, whether prenatal exposure to DBP can impair erectile function remains unknown. We conducted this study to investigate the potential effects of prenatal exposure to DBP on erectile function and the underlying mechanisms. A rat model of prenatal DBP exposure (12.5, 100 or 800 mg/kg/day by gavage during gestational days 13-21) was established. Prenatal DBP exposure significantly decreased penis/body weight ratio, myelin sheath thickness of cavernosum nerves and serum testosterone level in male rats at the age of 10 weeks. Furthermore, erectile dysfunction was detected in all DBP exposure groups, which exhibited substantial increases in transforming growth factor-ß1 (TGF-ß1) expression and decreases in the expression of alpha smooth muscle actin (α-SMA), neuronal and endothelial nitric oxide synthase (nNOS and eNOS). Additionally, the phospho-B-cell lymphoma 2 (Bcl-2)-associated death promoter (p-Bad)/Bad and phospho-the protein kinase B (p-AKT)/AKT ratios were remarkably lower, but the Bcl-2-associated X protein (Bax)/Bcl-2 ratio and caspase-3 were higher in DBP exposure groups than in the control group. Notably, prenatal exposure to DBP increase the risk of ED in male adult rats, even taking low dose of DBP (12.5 mg/kg/day). DBP exposure causing penile fibrosis, decreased testosterone level, and endothelial dysfunction may be responsible for ED by activating Akt/Bad/Bax/caspase-3 pathway and suppressing NOS/cGMP pathway in penis.

Effects of Fe-Mn oxide-modified biochar composite applications on phthalate esters (PAEs) accumulation in wheat grains and grain quality under PAEs-polluted brown soil.

Phthalate esters (PAEs), such as dibutyl phthalate (DBP) and di-(2-ethylhexyl) phthalate (DEHP), are used extensively as additives and plasticizers, and have become ubiquitous in the environment. PAEs in the soil could have adverse effects on crop plants as well as humans via accumulations in food chain. Thus, it is important to explore strategies to reduce the bioavailability of phthalate esters. We investigated the effects of Fe-Mn oxide-modified biochar composite (FMBC) applications on the quality of wheat grown in DBP- and DEHP-polluted brown soil. The application of FMBC and biochar (BC) increased the wheat grain biomass by 9.71-223.01% and 5.40-120.15% in the DBP-polluted soil, and 10.52-186.21% and 4.50-99.53% in the DEHP-spiked soil in comparison to the controls. All FMBC treatments were better than the BC treatments, in terms of decreasing DBP and DEHP bioavailability for the wheat grains. The activities of the glutamine synthetase and glutamic-pyruvic transaminase in the flag leaves at the filling stage and of granule-bound starch synthase, soluble starch synthase, and adenosine diphosphate-glucose pyrophosphorylase in the grains at maturity increased significantly with increases in either the BC or FMBC applications. This, in turn, increased the starch, protein, and amino acid content in the wheat grains. Compared with the BC treatment, the FMBC amendment induced only slight increases in the aforementioned factors. This study offers novel insights into potential strategies for decreasing PAEs bioavailability in soil, with potential positive implications for crop quality and environmental health improvements.

Toxicokinetics of diisobutyl phthalate and its major metabolite, monoisobutyl phthalate, in rats: UPLC-ESI-MS/MS method development for the simultaneous determination of diisobutyl phthalate and its major metabolite, monoisobutyl phthalate, in rat plasma, urine, feces, and 11 various tissues collected from a toxicokinetic study.

The aim of this study was to explore the toxicokinetics of diisobutyl-phthalate (DiBP) and its major metabolite, monoisobutyl-phthalate (MiBP), by developing a UPLC-ESI-MS/MS method for simultaneously measuring DiBP and MiBP in rat plasma, urine, feces, and 11 different tissues. For the experiment, 0.1% (v/v) aqueous formic acid and acetonitrile mobile phase by gradient elution at a flow rate of 0.3 mL/min, equipped with a KINETEX core-shell C18-column (50 × 2.1 mm, 1.7 µm), was used to completely separate analytes. The mass transitions were m/z 279.1 â†’ 149.0 for DiBP, 221.0 â†’ 77.0 for MiBP, and 283.2 â†’ 153.0 for DiBP-d4 as an internal standard. The developed assay had lower limits of quantification of 0.01 ng/mL for DiBP and 0.1 ng/mL for MiBP at all biological matrices. Toxicokinetics of DiBP were characterized by extensive distribution, short half-life, and high clearance. DiBP was rapidly metabolized to MiBP, with MiBP levels consistently exceeding the DiBP levels. Distribution of MiBP to tissues was considerable. The developed analytical method satisfied international criteria and was successfully applied to toxicokinetic studies after oral and intravenous administration of DiBP to rats. Findings of this study may be useful for evaluating the external exposure and toxic potential of DiBP and its metabolite in risk assessment.

Development and Validation of an Analytical Method for Quantitation of Monobutylphthalate, a Metabolite of Di-n-Butylphthalate, in Rat Plasma, Amniotic Fluid, Fetuses and Pups by UPLC-MS/MS.

Phthalates have been used for decades as softening agents in the production of plastics, but in recent years have been extensively investigated for their potential hazards to human health and the environment. Di-n-butyl phthalate (DBP), with widespread exposure occurring through a variety of consumer products such as cosmetics and pesticides, is a suspected carcinogen and an endocrine system disruptor in both humans and laboratory animals. Its predominant metabolite is the monoester, monobutyl phthalate (MBP), which can serve as a marker of exposure. To support toxicological studies of DBP in pregnant and lactating rats and their offspring, a novel ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for quantitation of MBP in rat plasma, amniotic fluid, fetuses and whole pup samples. Plasma samples were extracted using a simple protein precipitation with acetonitrile. Extraction and delipidation of pup homogenate was performed using acetonitrile and then submerging the vials in liquid nitrogen. Extracts were analyzed by UPLC-MS/MS in the negative ion mode. The method was successfully validated over the concentration ranges 25-5,000 ng/mL in female Sprague Dawley (SD) rat plasma and 50-5,000 ng/g in SD pup homogenate. Matrix calibration curves were linear (r ≥ 0.99), and the percent relative error (%RE) values were ≤ ±15% for standards at all levels. Absolute recoveries were > 92% in both matrices. The limits of detection (LODs) were 6.9 ng/mL in plasma and 9.4 ng/g in pup homogenate. Acceptable intra- and interday accuracy and precision were demonstrated by mean %RE ≤ ±7.5 and relative standard deviation (%RSD) ≤ 10.1%. Extract stability was demonstrated for ~6 days at various temperatures and freeze-thaw stability was demonstrated after 3 cycles over 3 days. Secondary matrix evaluation was performed for MBP in amniotic fluid and pooled fetus homogenate (mean %RE ≤ ±11.5 and %RSD ≤ 13.7). These data demonstrate that this simple method is suitable for determination of MBP in plasma, amniotic fluid, fetus and pup samples from toxicological studies of DBP.

Plastic film mulching increased the accumulation and human health risks of phthalate esters in wheat grains.

Plastic film mulching is a common practice to increase crop yield in dryland, while the wide use of plastic film has resulted in ubiquitous phthalate esters (PAEs) releasing into the soil. PAEs in soil could be taken up and accumulated by dietary intake of food crops such as wheat, thus imposing health risks to residents. In the present study, samples from a long-term location-fixed field experiment were examined to clarify the accumulation of PAEs in soil and wheat, and to assess the human health risks from PAEs via dietary intake of wheat grain under plastic film mulching cultivation in dryland. Results showed that concentrations of PAEs in grains from mulching plots ranged from 4.1 to 12.6 mg kg-1, which were significantly higher than those in the control group. There was a positive correlation for the PAE concentrations between wheat grains and field soils. Concentrations of PAEs in the soil were in the range of 1.8-3.5 mg kg-1 for the mulching treatment, and 0.9-2.7 mg kg-1 for the control group. Di-n-butyl phthalate (DBP) and di-(2-ethylhexyl) phthalate (DEHP) were detected in all soil and grain samples, and DEHP was found to be the dominant PAE compound in grains. Based on DEHP concentrations in wheat grains, the values of carcinogenic risk for adults were higher than the recommended value 10-4. Results indicated that wheat grains from film mulching plots posed a considerable non-carcinogenic risk to residents, with children being the most sensitive resident group. Findings of this work call the attention to the potential pollution of grain crops growing in the plastic film mulching crop production systems.

Interactions between microplastics and phthalate esters as affected by microplastics characteristics and solution chemistry.

Microplastics have become a major concern in recent years as they can be recognized as the transport vectors for pollutants in environment. In this study, the sorption behavior of two phthalate esters (PAEs), including diethyl phthalate (DEP) and dibutyl phthalate (DBP), onto three types of microplastics (PVC: polyvinyl chloride, PE: polyethylene, and PS: polystyrene) was investigated. The sorption isotherms of both DEP and DBP on microplastics were highly linear, suggesting that the partition was the main sorption mechanism. The Kd values of DBP were much higher than those of DEP, demonstrating that hydrophobic interaction governed the partition mechanism. Sorption of the two PAEs on the three microplastics followed the order of PS > PE > PVC, indicating that chemical properties of microplastics played an important roles in their sorption behaviors. Solution pH and natural organic matter had no significant impact on PAEs sorption by microplastics. However, the presence of NaCl and CaCl2 enhanced the sorption of both DEP and DBP because of the salting-out effect. The findings of the present study may have significant implications for the fate and transport assessment of both PAEs and microplastics.