Potency Factors Research Articles

Application of a Framework for Grouping and Mixtures Toxicity Assessment of PFAS: A Closer Examination of Dose-Additivity Approaches.

Environmental occurrence and biomonitoring data for per- and polyfluoroalkyl substances (PFAS) demonstrate that humans are exposed to mixtures of PFAS. This article presents a new and systematic analysis of available PFAS toxicity study data using a tiered mixtures risk assessment framework consistent with United States and international mixtures guidance. The lines of evidence presented herein include a critique of whole mixture toxicity studies and analysis of dose-response models based on data from subchronic oral toxicity studies in rats. Based on available data to-date, concentration addition and relative potency factor methods are found to be inappropriate due to differences among sensitive effects and target organ potencies and noncongruent dose-response curves for the same effect endpoints from studies using the same species and protocols. Perfluorooctanoic acid and perfluorooctane sulfonic acid lack a single mode of action or molecular initiating event and our evaluation herein shows they also have noncongruent dose-response curves. Dose-response curves for long-chain perfluoroalkyl sulfonic acids (PFSAs) also significantly differ in shapes of the curves from short-chain PFSAs and perfluoroalkyl carboxylic acids evaluated, and additional differences are apparent when curves are evaluated based on internal or administered dose. Following well-established guidance, the hazard index method applied to perfluoroalkyl carboxylic acids and PFSAs grouped separately is the most appropriate approach for conducting a screening level risk assessment for nonpolymeric PFAS mixtures, given the current state-of-the science. A clear presentation of assumptions, uncertainties, and data gaps is needed before dose-additivity methods, including hazard index , are used to support risk management decisions. Adverse outcome pathway(s) and mode(s) of action information for perfluorooctanoic acid and perfluorooctane sulfonic acid and for other nonpolymer PFAS are key data gaps precluding more robust mixtures methods. These findings can guide the prioritization of future studies on single chemical and whole mixture toxicity studies.

Neonicotinoid insecticide and their metabolite residues in fruit juices: Implications for dietary intake in China

Neonicotinoid insecticides (NEOs) have become the most widely used insecticides worldwide, and they are ubiquitous in food (i.e., fruit juices). In the present study, occurrence of seven NEOs and four metabolites (m-NEOs) in 400 fruit juice samples were investigated. NEOs and m-NEOs were frequently detected (65%–86%) in fruit juice samples. The median residues of NEOs and m-NEOs were ranged from 0.06 ng/mL to 0.94 ng/mL. Seasonal variations in NEOs and m-NEOs in fruit juices were found, indicating that the target analyte residues during the dry season were remarkably higher than those of residues during wet season. The relative potency factor (RPF) method was used to integrate individual NEOs into a single metric [imidacloprid (IMIRPF)] representing the intakes of IMI equivalent to total NEOs for each fruit juice sample. The estimated daily intake (EDI) of total NEOs for the general Chinese population was obtained. The median IMIRPF for total fruit juices was 13.4 ng/g, and the median EDI of NEOs was 18.2 ng/kg bw/day for the general population. Although the EDIs in this study were considerably lower than the acceptable daily intake (60 μg/kg bw/day, ADI), the dietary exposure risks for total NEOs should not be ignored because of the increasing usage of NEOs and their ubiquitous presence in fruit juices in China. To the best of our knowledge, this report was the first time to document residues of NEO and m-NEO in fruit juice samples collected from China.

Predicting the Activation of the Androgen Receptor by Mixtures of Ligands Using Generalized Concentration Addition.

Concentration/dose addition is widely used for compounds that act by similar mechanisms. But it cannot make predictions for mixtures of full and partial agonists for effect levels above that of the least efficacious component. As partial agonists are common, we developed generalized concentration addition, which has been successfully applied to systems in which ligands compete for a single binding site. Here, we applied a pharmacodynamic model for a homodimer receptor system with 2 binding sites, the androgen receptor, that acts according to the classic homodimer activation model: Each cytoplasmic monomer protein binds ligand, undergoes a conformational change that relieves inhibition of dimerization, and binds to DNA response elements as a dimer. We generated individual dose-response data for full (dihydroxytestosterone, BMS564929) and partial (TFM-4AS-1) agonists and a competitive antagonist (MDV3100) using reporter data generated in the MDA-kb2 cell line. We used the Schild method to estimate the binding affinity of MDV3100. Data for individual compounds fit the homodimer pharmacodynamic model well. In the presence of a full agonist, the partial agonist had agonistic effects at low effect levels and antagonistic effects at high levels, as predicted by pharmacological theory. The generalized concentration addition model fits the empirical mixtures data-full/full agonist, full/partial agonist, and full agonist/antagonist-as well or better than relative potency factors or effect summation. The ability of generalized concentration addition to predict the activity of mixtures of different types of androgen receptor ligands is important as a number of environmental compounds act as partial androgen receptor agonists or antagonists.

Levels and inhalation health risk of neonicotinoid insecticides in fine particulate matter (PM2.5) in urban and rural areas of China

Neonicotinoid insecticide (NEO) concentrations in ambient fine particulate matter (PM2.5) and daily exposure via inhalation were investigated during spring and fall in an urban area in Beijing and in urban and rural areas of Zhengzhou, Henan Province, China. Four NEOs, including imidacloprid, acetamiprid, thiamethoxam and clothianidin, were assessed using a QuEChERS (quick, easy, cheap, effective, rugged, safe) extraction procedure coupled to liquid chromatography–tandem mass spectrometry. Of 64 PM2.5 samples, 100% contained at least two NEOs (imidacloprid and acetamiprid). Imidacloprid was detected at the highest levels, ranging from 4.33 to 1.13 × 102 pg m−3. A relative potency factor method that considered different toxicities was used to integrate the four NEO concentrations. The total NEO concentrations in air in the Zhengzhou rural area (mean: 80.86 pg m−3) were higher than those in urban areas. Differences between seasons were not significant (p > 0.05). The highest value for the total average daily dose via inhalation of four NEOs (ADDinh,total), 91.0 pg kg−1 day−1, was found in rural children <6 years old. The ADDinh,total of rural residents was significantly higher than that of urban residents when there was no intensive pesticide application. Although the ADDinh,total values were below the current chronic reference dose, when possible joint toxicity and the increasing use of NEOs are considered, a potential health risk via inhalation is evident. We believe this study is the first to characterize NEO levels in fine particulate matter and to evaluate inhalation exposure in urban and rural residents under nonoccupational scenarios in China. It will enhance our understanding of exposure to NEOs and provide a basis for risk management decisions.

Derivation of clinical-grade mesenchymal stromal cells from umbilical cord under chemically defined culture condition – platform for future clinical application

The use of animal serum in culture medium brings safety concerns and batch-to-batch variability, and thus may restrict the clinical use of ex vivo expanded mesenchymal stromal cells (MSCs). Clinically compliant MSCs should be developed in adherence to serum-free, xeno-free and chemically defined medium (S&XFM-CD). In this study, we develop a S&XFM-CD by replacing all serum components with synthetic alternatives for the derivation of clinical-grade umbilical cord-derived MSCs (UCMSCs). The critical aspects including characterization, safety concerns, potency and exogenous factors contamination risk of UCMSCs in S&XFM-CD are compared with serum-containing medium (SCM). UCMSCs in S&XFM-CD retain fibroblastic-like morphology and immunophenotype of MSCs, and exhibit superior clone efficiency, proliferation capacity, and osteogenic and chondrogenic differentiation potential compared with SCM. Moreover, UCMSCs in S&XFM-CD retain similar immunosuppressive potential, and exhibit superior secretion levels of bFGF, PDGF-BB and IGF-1 compared with SCM. In addition, UCMSCs in S&XFM-CD do not undergo transformation, preserve the normal karyotypes and genomic stability, and are less prone to senescence process after long-term in vitro culture, which conforms to the current guidance of international and national evaluation standard. The S&XFM-CD developed here may serve as a GMP-grade production platform of UCMSCs for future clinical application.

Cumulative dietary risk assessment overarching different regulatory silos using a margin of exposure approach: A case study with three chemical silos

Risk assessment of chemicals occurring in our diet is commonly performed for single chemicals without considering exposure to other chemicals. We performed a case study on risk assessment of combined dietary exposure to chemicals from different regulatory silos, i.e. pesticides (PPRs), persistent organic pollutants (POPs) and food additives (FAs). Chemicals were grouped into the cumulative assessment group (CAG) liver steatosis using a component-based approach. Based on literature, the CAG included 144 PPRs, 49 POPS and 7 FAs for which concentration data were available.For each silo, chronic combined dietary exposure was assessed for adults and children of nine European countries following the most commonly used exposure methodologies in Europe and by using a relative potency factor approach. For risk characterization, a Margin of Exposure (MOE) was calculated. To overarch the risk across silos, a normalised combined margin of exposure (nMOET) approach was proposed.This case study demonstrated that risk assessment of combined exposure to chemicals can be performed within regulatory silos. It also highlighted important differences in the conservatism of exposure scenarios, the derivation of point of departures and the subsequent acceptable MOEs between the silos. To overarch the risk despite these differences, a nMOET approach can be used.

Development of an allogeneic stem cell product "ELIXCYTE" for the treatment of osteoarthritis and chronic kidney disease

Background & Aim ELIXCYTE is an allogeneic adipose stem cell product and is currently undergoing two human clinical trials. In the clinical trial of ELIXCYTE for osteoarthritis (NCT02784964), the patient enrollment of phase-I/IIa have been completed and the results of a 24-weeks follow-up have been analyzed. For chronic kidney disease (CKD) treatment (NCT02933827), the phase-II clinical trial is ongoing. In addition of clinical trial development, we established suitable mechanism of actions (MOAs) platforms to describe the pharmacology and the potency factors of ELIXCYTE. And in the process development of cell manufacturing technology, we have also made great breakthrough of mass and automatic production of ELIXCYTE by 3D bioreactor systems. Methods, Results & Conclusion Based on the pharmacological mechanisms of ELIXCYTE obtained from our pre-clinical studies, for osteoarthritis treatment, our stem cell products show good immune regulation, anti-inflammatory properties, as well as the characteristics of promoting chondrocyte proliferation and protecting chondrocytes. For CKD treatment, we proposed that ELIXCYTE can achieve the effect of maintain and repair renal function through its ability to regulate inflammation, angiogenesis, anti-fibrosis, and antioxidation. The results of clinical trials show that ELIXCYTE has good safety in the treatment of arthritis and kidney disease, and both show the potential and trends for positive response and efficacy. For osteoarthritis treatment, the trend of pain relief was shown in current data. For CKD treatment, current data show positive response of renal function index (eGFR) in CKD patients. In the breakthrough of cell manufacturing technologies, we have successfully developed the use of bioreactors for cell production in 50-L scale and the whole semi-automatic manufacturing processes have been established. ELIXCYTE is an allogeneic adipose stem cell product and is currently undergoing two human clinical trials. In the clinical trial of ELIXCYTE for osteoarthritis (NCT02784964), the patient enrollment of phase-I/IIa have been completed and the results of a 24-weeks follow-up have been analyzed. For chronic kidney disease (CKD) treatment (NCT02933827), the phase-II clinical trial is ongoing. In addition of clinical trial development, we established suitable mechanism of actions (MOAs) platforms to describe the pharmacology and the potency factors of ELIXCYTE. And in the process development of cell manufacturing technology, we have also made great breakthrough of mass and automatic production of ELIXCYTE by 3D bioreactor systems. Based on the pharmacological mechanisms of ELIXCYTE obtained from our pre-clinical studies, for osteoarthritis treatment, our stem cell products show good immune regulation, anti-inflammatory properties, as well as the characteristics of promoting chondrocyte proliferation and protecting chondrocytes. For CKD treatment, we proposed that ELIXCYTE can achieve the effect of maintain and repair renal function through its ability to regulate inflammation, angiogenesis, anti-fibrosis, and antioxidation. The results of clinical trials show that ELIXCYTE has good safety in the treatment of arthritis and kidney disease, and both show the potential and trends for positive response and efficacy. For osteoarthritis treatment, the trend of pain relief was shown in current data. For CKD treatment, current data show positive response of renal function index (eGFR) in CKD patients. In the breakthrough of cell manufacturing technologies, we have successfully developed the use of bioreactors for cell production in 50-L scale and the whole semi-automatic manufacturing processes have been established.

Perfluoroalkyl substances and anthropomorphic measures in children (ages 3–11 years), NHANES 2013–2014.

BackgroundPerfluoroalkyl acids (PFAAs) are man-made compounds that are persistent in the environment and highly bioaccumulative in the body. Humans are exposed to a mixture of these substances, and the effects of these mixtures may be different than the effects noted for individual compounds. Prenatal exposure to PFAAs has been associated with decreased birth weight. The objective of the present study is to evaluate concurrent serum PFAA levels, as single compounds and as mixtures, in relation to anthropomorphic measures in children. MethodsUsing multivariate linear regression, we evaluated the association between single or PFAA mixtures and with height-for-age (HAZ), weight-for-age (WAZ), and BMI (BMIZ) z-scores in children (ages 3–11 years) participants of the National Health and Nutrition Examination Survey (NHANES) 2013–2014. Analyses were also stratified by sex. The PFAA mixture was based on relative potency factors express in terms of PFOA equivalency (CmixRPFi) or as molar sum of the PFAA congeners (∑molPFAA). ResultsThere was a statistically significant association of PFHxS and PFOS with decreased HAZ in boys. The significantly decreased HAZ in boys was also found when the PFAAs were analyzed as mixtures: CmixRPFi (β = −0.33; 95%CI: 0.63, −0.04) or ΣmolPFAAs (β = −0.30; 95%CI: 0.56, −0.04). In boys, PFHxS was also associated with decreased WAZ and BMIZ. The only statistically significant association found in girls was between decreased HAZ and PFHxS. ConclusionsWe found sex differences in the association between concurrent serum PFAA levels and anthropomorphic measures in children 3–11 years old. PFAA levels, as single congeners or as mixture concentrations were associated with decreased height-for-age z-score in boys.

Application of a combined aggregate exposure pathway and adverse outcome pathway (AEP-AOP) approach to inform a cumulative risk assessment: A case study with phthalates

Advancements in measurement and modeling capabilities are providing unprecedented access to estimates of chemical exposure and bioactivity. With this influx of new data, there is a need for frameworks that help organize and disseminate information on chemical hazard and exposure in a manner that is accessible and transparent. A case study approach was used to demonstrate integration of the Adverse Outcome Pathway (AOP) and Aggregate Exposure Pathway (AEP) frameworks to support cumulative risk assessment of co-exposure to two phthalate esters that are ubiquitous in the environment and that are associated with disruption of male sexual development in the rat: di(2-ethylhexyl) phthalate (DEHP) and di-n-butyl phthalate (DnBP). A putative AOP was developed to guide selection of an in vitro assay for derivation of bioactivity values for DEHP and DnBP and their metabolites. AEPs for DEHP and DnBP were used to extract key exposure data as inputs for a physiologically based pharmacokinetic (PBPK) model to predict internal metabolite concentrations. These metabolite concentrations were then combined using in vitro-based relative potency factors for comparison with an internal dose metric, resulting in an estimated margin of safety of ~13,000. This case study provides an adaptable workflow for integrating exposure and toxicity data by coupling AEP and AOP frameworks and using in vitro and in silico methodologies for cumulative risk assessment.

An adverse outcome pathway-based approach to assess steatotic mixture effects of hepatotoxic pesticides in vitro

Exposure to complex chemical mixtures requires a tiered strategy for efficient mixture risk assessment. As a part of the EuroMix project we developed an adverse outcome pathway (AOP)-based assay toolbox to investigate the combined effects of the liver steatosis-inducing compounds imazalil, thiacloprid, and clothianidin in human HepaRG hepatocarcinoma cells. Compound-specific relative potency factors were determined using a benchmark dose approach. Equipotent mixtures were tested for nuclear receptor activation, gene and protein expression, and triglyceride accumulation, according to the molecular initiating events and key events proposed in the steatosis AOP. All three compounds affected the activity of nuclear receptors, but not key genes/proteins as proposed. Triglyceride accumulation was observed with three different methods. Mixture effects were in agreement with the assumption of dose additivity for all the combinations and endpoints tested. Compound-specific RPFs remained similar over the different endpoints studied downstream the AOP. Therefore, it might be possible to reduce testing to a smaller battery of key tests. The results demonstrate the suitability of our in vitro assay toolbox, integrated within an AOP framework and combined with the RPF approach, for the analysis of steatotic effects of chemical mixtures. However, mRNA results suggest that the steatosis AOP still needs improvement.

Risk assessment of aflatoxins in food.

EFSA was asked to deliver a scientific opinion on the risks to public health related to the presence of aflatoxins in food. The risk assessment was confined to aflatoxin B1 (AFB1), AFB2, AFG1, AFG2 and AFM1. More than 200,000 analytical results on the occurrence of aflatoxins were used in the evaluation. Grains and grain‐based products made the largest contribution to the mean chronic dietary exposure to AFB1 in all age classes, while ‘liquid milk’ and ‘fermented milk products’ were the main contributors to the AFM1 mean exposure. Aflatoxins are genotoxic and AFB1 can cause hepatocellular carcinomas (HCCs) in humans. The CONTAM Panel selected a benchmark dose lower confidence limit (BMDL) for a benchmark response of 10% of 0.4 μg/kg body weight (bw) per day for the incidence of HCC in male rats following AFB1 exposure to be used in a margin of exposure (MOE) approach. The calculation of a BMDL from the human data was not appropriate; instead, the cancer potencies estimated by the Joint FAO/WHO Expert Committee on Food Additives in 2016 were used. For AFM1, a potency factor of 0.1 relative to AFB1 was used. For AFG1, AFB2 and AFG2, the in vivo data are not sufficient to derive potency factors and equal potency to AFB1 was assumed as in previous assessments. MOE values for AFB1 exposure ranged from 5,000 to 29 and for AFM1 from 100,000 to 508. The calculated MOEs are below 10,000 for AFB1 and also for AFM1 where some surveys, particularly for the younger age groups, have an MOE below 10,000. This raises a health concern. The estimated cancer risks in humans following exposure to AFB1 and AFM1 are in‐line with the conclusion drawn from the MOEs. The conclusions also apply to the combined exposure to all five aflatoxins.

The MCRA toolbox of models and data to support chemical mixture risk assessment

A model and data toolbox is presented to assess risks from combined exposure to multiple chemicals using probabilistic methods. The Monte Carlo Risk Assessment (MCRA) toolbox, also known as the EuroMix toolbox, has more than 40 modules addressing all areas of risk assessment, and includes a data repository with data collected in the EuroMix project. This paper gives an introduction to the toolbox and illustrates its use with examples from the EuroMix project. The toolbox can be used for hazard identification, hazard characterisation, exposure assessment and risk characterisation. Examples for hazard identification are selection of substances relevant for a specific adverse outcome based on adverse outcome pathways and QSAR models. Examples for hazard characterisation are calculation of benchmark doses and relative potency factors with uncertainty from dose response data, and use of kinetic models to perform in vitro to in vivo extrapolation. Examples for exposure assessment are assessing cumulative exposure at external or internal level, where the latter option is needed when dietary and non-dietary routes have to be aggregated. Finally, risk characterisation is illustrated by calculation and display of the margin of exposure for single substances and for the cumulation, including uncertainties derived from exposure and hazard characterisation estimates.

Relative potency factor approach enables the use of in vitro information for estimation of human effect factors for nanoparticle toxicity in life-cycle impact assessment

The major theme of the NRC report “Toxicity Testing in the Twenty-first Century” is to replace animal testing by using alternative in vitro methods. Therefore, it can be expected that in the future in vivo data will be replaced with in vitro data. Hence, there is a need for new strategies to make use of the increasing amount of in vitro data when developing human toxicological effect factors (HEF) to characterize the impact category of human toxicity in life cycle assessment (LCA). Here, we present a new approach for deriving HEF for manufactured nanomaterials (MNMs) based on the combined use of in vitro toxicity data and a relative potency factor (RPF) approach. In vitro toxicity tests with nano-CuO, nano-Ag and nano-ZnO and their corresponding ions were performed on THP-1, CaCo-2 and Hep-G2 cell lines. The ratio of the here calculated EC50 of the ionic form and the nanoform corresponds to the Relative Potency Factor (RPF). Using this approach, HEFs (case/kgintake) for the aforementioned nanoparticles were obtained. Non-carcinogenic HEFs (case/kgintake) for exposure via ingestion of 5.9E-01, 7.5E-03 and 2.5 E-02 were calculated for nano-Ag, nano-CuO and nano-ZnO, respectively. The HEF values here proposed were compared with HEF values extrapolated from in vivo toxicity data reported in the literature. The here presented procedure is the most appropriate approximation currently available for using in vitro toxicity data on MNM for application in the field of LCIA.

Dietary exposure to neonicotinoid insecticides and health risks in the Chinese general population through two consecutive total diet studies

Neonicotinoid insecticides are ubiquitous in food and the environment due to their wide use. Growing evidence suggests the adverse effects of neonicotinoids in many species, including mammals. Some studies have reported the urinary concentrations of neonicotinoids in human biological monitoring, but the potential risks of neonicotinoids on human health based on long-term chronic exposure studies in any general population have been rarely tackled. In this study, the dietary exposure to neonicotinoids of the Chinese adult population was studied on the basis of composite dietary samples collected from the 5th (2009–2012) and 6th (2015–2018) Chinese total diet studies (TDS). Residue levels of ten neonicotinoids were determined in 528 composite dietary samples from 24 provinces in China. Most of the samples (53.3% and 70.5% in the 5th and 6th TDS, respectively) that we analyzed contained the multi-residue of neonicotinoids. Imidacloprid and acetamiprid were the most frequently detected neonicotinoids, and thiamethoxam and clothianidin were increasingly used and found in the 6th TDS. The estimated daily intake (EDI) for total neonicotinoids was calculated to evaluate health risk of the Chinese adult population based on a relative potency factor assessment method. The mean EDIs of total neonicotinoids in the 5th and 6th TDS respectively reached 598.95 and 710.38 ng/kg bw per day. Although the mean EDIs of total neonics in 6th TDS was relatively higher than that in 5th TDS, no statistical difference was observed (p > 0.05). Vegetables were the main source of dietary exposure, but exposure via cereals and beverages and water must also be addressed in China. Although the average exposure for total neonicotinoids was much lower than the current chronic reference dose (57 μg/kg bw per day), the dietary exposure risks of a general population for total neonicotinoids should not be overlooked due to the ubiquity of neonicotinoids in food and the environment.

Griffithsin Retains Anti-HIV-1 Potency with Changes in gp120 Glycosylation and Complements Broadly Neutralizing Antibodies PGT121 and PGT126.

Griffithsin (Grft) is an antiviral lectin that has been shown to potently inhibit HIV-1 by binding high-mannose N-linked glycosylation sites on HIV-1 gp120. A key factor for Grft potency is glycosylation at N295 of gp120, which is directly adjacent to N332, a target glycan for an entire class of broadly neutralizing antibodies (bNAbs). Here, we unify previous work on the importance of other glycans to Grft potency against HIV-1 and Grft's role in mediating the conformational change of gp120 by mutating nearly every glycosylation site in gp120. In addition to a significant loss of Grft activity by the removal of glycosylation at N295, glycan absence at N332 or N448 was found to have moderate effects on Grft potency. Interestingly, in the absence of N295, Grft effectiveness could be improved by a mutation that results in the glycan at N448 shifting to N446, indicating that the importance of individual glycans may be related to their effect on glycosylation density. Grft's ability to alter the structure of gp120, exposing the CD4 binding site, correlated with the presence of glycosylation at N295 only in clade B strains, not clade C strains. We further demonstrate that Grft can rescue the activity of the bNAbs PGT121 and PGT126 in the event of a loss or a shift of glycosylation at N332, where the bNAbs suffer a drastic loss of potency. Despite targeting the same region, Grft in combination with PGT121 and PGT126 produced additive effects. This indicates that Grft could be an important combinational therapeutic.

Neonicotinoid insecticides in the drinking water system – Fate, transportation, and their contributions to the overall dietary risks

Neonicotinoids (Neonics) have become the most widely used insecticides around the world in recent years. Due to the hydrophilic character, neonics are emerging contaminants in drinking water. In this study, we aimed to characterize and quantify the fate and transport of neonics in the drinking water treatment system and their contributions to the overall dietary risks. Seven neonics in 97 surface and drinking water samples in the city of Hangzhou, China were analyzed. The relative potency factor method was adopted in order to calculate the total neonics concentrations. We then used the Monte Carlo simulation to calculate the chronic daily intake (CDI) of total neonics from water consumption. All 16 surface water samples collected from two rivers contained at least two neonics, and more than 93% of those contained 3 or more neonics. Imidacloprid was detected in all 16 surface water samples, followed by clothianidin and acetamiprid with average concentrations of 11.9, 7.6, 17.6 ng L-1, respectively. The drinking water treatment plants removed approximately 50% of neonics from surface water. However, 68 out of 71 tap water samples that we collected from the household faucets contained at least one neonic, with the highest average concentrations of 5.8 ng L-1 for acetamiprid. The maximum of CDIs of total neonics from water consumption for adult and children were 10.2 and 12.4 ng kg-1 d-1, respectively, which are significantly lower than the acceptable daily intake (ADI). The results presented here shown drinking water consumption only represented an insignificant portion of dietary risks of total neonics, mainly due to the modern drinking water treatment technologies that are capable of removing significant amount of neonics from drinking water. However, the ubiquity of neonics in the drinking water sources to kitchen faucets, should be a concern for public health.

Update of the risk assessment of di-butylphthalate (DBP), butyl-benzyl-phthalate (BBP), bis(2-ethylhexyl)phthalate (DEHP), di-isononylphthalate (DINP) and di-isodecylphthalate (DIDP) for use in food contact materials.

The EFSA Panel on Food Contact Materials, Enzymes and Processing Aids (CEP Panel) was asked by the European Commission to update its 2005 risk assessments of di‐butylphthalate (DBP), butyl‐benzyl‐phthalate (BBP), bis(2‐ethylhexyl)phthalate (DEHP), di‐isononylphthalate (DINP) and di‐isodecylphthalate (DIDP), which are authorised for use in plastic food contact material (FCM). Dietary exposure estimates (mean and high (P95)) were obtained by combining literature occurrence data with consumption data from the EFSA Comprehensive Database. The highest exposure was found for DINP, ranging from 0.2 to 4.3 and from 0.4 to 7.0 μg/kg body weight (bw) per day for mean and high consumers, respectively. There was not enough information to draw conclusions on how much migration from plastic FCM contributes to dietary exposure to phthalates. The review of the toxicological data focused mainly on reproductive effects. The CEP Panel derived the same critical effects and individual tolerable daily intakes (TDIs) (mg/kg bw per day) as in 2005 for all the phthalates, i.e. reproductive effects for DBP (0.01), BBP (0.5), DEHP (0.05), and liver effects for DINP and DIDP (0.15 each). Based on a plausible common mechanism (i.e. reduction in fetal testosterone) underlying the reproductive effects of DEHP, DBP and BBP, the Panel considered it appropriate to establish a group‐TDI for these phthalates, taking DEHP as index compound as a basis for introducing relative potency factors. The Panel noted that DINP also affected fetal testosterone levels at doses around threefold higher than liver effects and therefore considered it conservative to include it within the group‐TDI which was established to be 50 μg/kg bw per day, expressed as DEHP equivalents. The aggregated dietary exposure for DBP, BBP, DEHP and DINP was estimated to be 0.9–7.2 and 1.6–11.7 μg/kg bw per day for mean and high consumers, respectively, thus contributing up to 23% of the group‐TDI in the worst‐case scenario. For DIDP, not included in the group‐TDI, dietary exposure was estimated to be always below 0.1 μg/kg bw per day and therefore far below the TDI of 150 μg/kg bw per day. This assessment covers European consumers of any age, including the most sensitive groups. Based on the limited scope of the mandate and the uncertainties identified, the Panel considered that the current assessment of the five phthalates, individually and collectively, should be on a temporary basis.

Validity of using a relative potency factor approach for the risk management of dioxin-like polychlorinated naphthalenes.

Here, we characterized the dioxin-like activities of 42 polychlorinated naphthalenes (PCNs) and 6 technical Halowax formulations by using the DR-CALUX (dioxin-responsive chemically activated luciferase expression) assay with rat hepatoma luciferase-expressing H4IIE cells. Of the 42 PCNs examined, 31 showed dioxin-like activities, for which the mass-based REP-EC5TCDD (potency relative to that of 2,3,7,8-tetrachlorodibenzo-p-dioxin based on the 5% effective concentration determined from the dose-response curve for 2,3,7,8-TCDD) ranged from 0.00000012 to 0.0051, indicating that some of the PCNs (e.g., 1,2,3,6,7,8-HxCN and 1,2,3,4,6,7-HxCN) had dioxin-like activities that were equal to or higher than the WHO-TEFs and the mass-based REP-EC5TCDD reported for dioxins such as octachlorodibenzo-p-dioxin, octachlorodibenzofuran, 3,3',4,4'-tetrachlorobiphenyl (PCB-77), 3,4,4',5-tetrachlorobiphenyl (PCB-81), and 3,3',4,4',5,5'-hexachlorobiphenyl (PCB-169). For PeCNs to OCN with high dioxin-like activities, REPs determined in previous studies were comparable to the REP values obtained in the present study. The TCDD-EQs (2,3,7,8-TCDD equivalents) obtained experimentally for the Halowax formulations decreased in the order HW1051 (37mg/kg)>HW1014 (30mg/kg)>HW1013 (5.6mg/kg)>HW1099 (2.9mg/kg)>HW1001 (0.60mg/kg)>HW1031 (<0.10mg/kg) and were comparable to the theoretical TCDD-EQs calculated by multiplying the concentration and REP of each PCN. In addition, the theoretical TCDD-EQs for PCNs in emission gases produced by thermal processes were below the Japanese emission standard of 0.1-10ng WHO-toxicity equivalent (TEQ)/m3N, and 3 to 4 orders of magnitude lower than the corresponding WHO-TEQ. Based on a comparison of theoretical and experimental TCDD-EQs, we found that our REP-based approach was suitable for the risk management of industrially produced and unintentionally generated dioxin-like PCNs. This approach will be particularly useful for the risk management of unintentionally generated PCNs in emission gases because the contribution of dioxin-like PCNs to the whole dioxin-like toxicity of emission gases can be elucidated.

USP Potency Adjusted Bovine Mucosal Heparins (BMH) Are Comparable to Porcine Mucosal Heparin (PMH) at Equivalent Levels

Introduction: Unfractionated heparin (UFH) remains to be the only parenteral anticoagulant used in the management of various thrombotic disorders such as deep vein thrombosis (DVT), pulmonary embolism (PE), and cardiovascular interventions. Most of the heparins used clinically are derived from porcine intestinal mucosa. There is likelihood of supply shortage of this important anticoagulant which is crucial for hemodialysis, cardiopulmonary bypass surgery and other vascular interventions. BMH are currently being developed for re-introduction for both medical and surgical indications. In contrast to the PMH, the active pharmaceutical ingredient (API) of BMH exhibit a somewhat weaker USP potency as cross-referenced against PMH. We hypothesized that at equivalent potencies as adjusted by using the USP reference, the BMH may exhibit comparable in vitro and in vivo effects. Therefore, in vitro and in vivo studies were used to compare the APIs of the bovine (140 U/mg) and the PMH (190 U/mg) to demonstrate their bioequivalence. Materials and Methods: API versions of PMH (190 U/mg) were obtained from Celsus Laboratories (Franklin, OH). API versions of BMH (140 U/mg) were obtained from KinMaster (Paso Fundo, Brazil). Each of these heparins was assayed for their molecular weight profile, AT affinity, USP potency, protamine and platelet factor 4 neutralization and anticoagulant/antiprotease profiles using standard laboratory methods. In the primate studies, potencies of each heparin were determined by amidolytic anti-Xa assay in relation to the USP heparin standard. Individual groups of primates (n=4) were administered 100 anti-Xa U/kg doses of bovine or porcine heparin via intravenous route. Blood samples were collected prior to dosing and at 15-, 30-, 60- and 120-minutes post-administration. Anti-Xa and anti-IIa activities were measured to determine circulating heparin concentrations using commercially available USP compliant kits (Aniara Diagnostica, West Chester, OH). These drug concentrations were used to determine pharmacokinetic parameters such as area under the curve (AUC), half-life (t1/2), clearance (Cl) and volume of distribution (Vd) using the PKSolver add-in for Excel. Results: BMH exhibited higher molecular weight profiles compared to PMH as determined by size exclusion chromatography (BMH (Mw) 18.6 ± 0.5 kDa and PMH 15.4 ± 0.4 kDa). BMH exhibited a potency of 140 U/mg and PMH had a potency of 195 U/mg. In the anticoagulant and antiprotease assays, the BMH exhibited lower functionality which was proportional to USP potency. In vitro, when the BMH was compared at a potency adjusted concentration with PMH, it showed identical calibration curves in the aPTT and anti-protease assays. However, in the protamine neutralization and platelet factor 4 studies, BMH required slightly higher amounts of the agents in contrast to PMH. The concentration vs. time curves for both heparins were almost superimposable. Peak drug levels of approximately 1.5 and 1.4 U/mL were measured using anti-Xa and anti-IIa assays, respectively. After 2 hours, circulating drug levels were decreased to approximately 0.4 U/mL for all heparins. Pharmacokinetic parameters calculated from plasma concentration-time curves indicated that both heparins behaved similarly. Mean half-life based on anti-Xa activity ranged from 54 ± 11 min for porcine heparin to 71 ± 18 min for bovine heparin. Slightly longer half-lives were observed using plasma concentrations determined using anti-IIa activity. Mean AUC values based on anti-Xa or anti-IIa activities were comparable for both heparins. Mean Vd (~60 ml/kg) and Cl (~0.75 ml/kg/min) were also comparable for both heparins. Conclusion: In vitro, BMH at adjusted biologic potency is comparable to PMH, however, it requires proportionally higher amount of protamine and platelet factor 4 due to the increased mass for adjusting to higher potency. In the non-human primates, USP cross-referenced anti-Xa potency adjusted based dosing results in comparable pharmacokinetic profiles for bovine and porcine heparins. Therefore, such dosing may provide uniform levels of anticoagulation for the parenteral indications for heparins. These observations warrant clinical validations in the specific indications. Disclosures No relevant conflicts of interest to declare.

Structure-dependent hepato-cytotoxic potencies of selected pyrrolizidine alkaloids in primary rat hepatocyte culture

Contamination of food, feed and herbal medicines with plants containing pyrrolizidine alkaloids (PA) leads to measurable amounts of PA in many products. Since a number of PA are hepatotoxic in humans and animals and hepato-carcinogenic in animal experiments, the assessment of the relative toxic potencies of widely occurring PA contaminants warrants detailed investigation. Here, we studied the hepato-cytotoxic potencies of a number of relevant PA congeners in rat hepatocytes in primary culture. It was found that cyclic and open di-esters were much more toxic than mono-esters. Furthermore, the hepatocellular levels of cytochrome P450-catalyzed 7-benzoxyresorufin O-dealkylase (BROD) activity decreasing over time in culture, played an important role for activation of PA into cytotoxic metabolites. With a highly toxic PA (lasiocarpine), inhibition of BROD activity with ketoconazole markedly reduced toxicity while this was not obvious with the less toxic congener lycopsamine. Depletion of cellular glutathione with buthionine sulfoximine had no significant influence on the effects of highly toxic PA whereas it slightly increased toxicity of less potent congeners. Overall, our data partially confirm previously published structure-dependent interim Relative Potency (iREP) factors although for echimidine and monocrotaline in particular, substantial deviations were found, possibly due to specific toxicokinetic properties of these congeners.