Study population and procedures

From October 2012 to January 2013, ninety-two children between the ages of 3–5 years old were recruited from 28 preschool classrooms in two geographic areas of Oregon, USA. Written informed consent was obtained from the parent and/or legal guardian for all participants and from all preschool teachers prior to engaging in any study activity. Child assent was indicated by their engagement with the materials and/or project staff.

Researchers visited participants’ homes where parents completed a series of structured questionnaires to capture socio-demographic information (e.g. household income, parental education levels, race, etc.) and the home learning environment. During this visit, each child was given a silicone passive wristband sampler to wear around his/her wrist or ankle. This sampler was used to assess the child’s exposure to flame retardants. Parents were requested to have their child wear the wristband continuously for 7 days, although it could be taken off at night and placed next to the child’s bed on a table if preferred. After 7 days, parents were instructed to seal the wristband in the polytetrafluoroethylene (PTFE) bag, note the number of days the child actually wore the wristband on the chain of custody tag, and place it in the mail. The concentration of flame retardants measured in the silicone passive wristband sampler were also reported back to the participants. Children’s social behaviors were assessed using the Social Skills Improvement System - Rating Scales by their teacher in the preschools they were attending.

Of the 92 wristbands distributed to the children, 77 were returned for analysis. Of those, five samples were excluded due to parent report of substantial deviance from the protocol (e.g. never worn by the child, lost at school for several weeks, or went through the laundry). Additionally, three parents chose not to answer questions on the socio-demographic questionnaire, which left a final sample size of 69 children with complete data that were included in the final analyses.

Ethical statement

All research activities were approved by Oregon State University’s Institutional Review Board. All parents gave informed written consent and children gave assent before partaking in any research activity. Results from the chemical results from the wristbands were returned to the parents. Due to the novelty of the exposure assessment methodology there were no other populations that could serve as a comparison for this study. Subsequently, parents were told which flame retardants were detected in their children’s sample and where their child’s exposure ranked within this group of children (e.g. in the lowest 25th percentile, in the 25th to 75th percentile, or highest 25th percentile). Parents were also given resources created by the Agency for Toxic Substances and Disease Registry where they could learn more about flame retardants and resources created by the Oregon Environmental Council’s Eco-Health Homes Checkup Kit where they could learn about how to reduce exposure to pollutants in their home including flame retardants.

Social behavior assessment

The teacher form of the Social Skills Improvement System Rating Scale (SSIS-RS) was used to measure children’s social behavior in preschool classrooms. The SSIS-RS is a standardized assessment of social skills and problem behaviors for children ages 3–18 years, has strong psychometric properties, and measures both normative and clinically-relevant variation [36]. Each item asks teachers to indicate the frequency of children’s behaviors from 0 (never) to 3 (always). We examined seven subscales representing positive behaviors: Communication, Cooperation, Assertion, Responsibility, Empathy, Engagement, and Self-Control; and four subscales representing behavior problem domains: Externalizing, Bullying, Hyperactivity/Inattention, and Internalizing. Sample items of positive behaviors include “follows classroom rules” and “expresses feelings when wronged” and “makes a compromise during a conflict.” Sample behavior problem items include “disobeys rules or requests”, “Is aggressive toward people or objects”, and “acts without thinking”.

Our preliminary analyses indicated similar associations between flame retardants and the three subscales of externalizing behavior problems (externalizing, hyperactivity/inattention, and bullying). Subsequently, we utilized an aggregate of these three subscales in the final analyses. Preliminary analyses suggested, however, that the subscales of positive behaviors were associated differently to PBDEs and OPFRs. Thus, each of the individual subscales of positive behaviors were examined in the final analyses. Internal consistencies for the subscales ranged from .81 for assertion to .93 for self-control and the externalizing aggregate.

Flame retardant exposure

The methodology and results for the detection of flame retardants in silicone passive sampling devices utilized in this study have been published elsewhere [34]. Briefly, silicone wristbands were purchased from a commercial retailer (24hourwristbands.com) and prepared following the method described in O’Connell et al. [39]. The solvent-cleaned wristbands were then packaged in clean, air-tight PTFE bags along with a chain of custody tag and given to parents along with instructions to have the child wear the wristband at all time for 7 days either as a bracelet or anklet. Parents were instructed to place the wristband on a table near the child’s bed if he/she did not want to wear the wristband while sleeping. Parents were then requested to re-seal the wristband in its PTFE bag, fill in the chain of custody tag with the number of days the wristband was worn by the child, and mail it back to Oregon State University using the provided pre-paid business envelope. The wristbands were then extracted and analyzed for 41 different flame retardant compounds using gas chromatography mass spectrophotometry.

This analysis focused on 11 compounds (PBDE-47, PBDE-99, PBDE-153, PBDE-154, PBDE-49, PBDE28 + 33, tris(1,3-dichloro-2-propyl) phosphate], TPP [e.g. triphenylphosphate], TCPP [e.g tris(1-chloro-2-propyl) phosphate], and TCEP [e.g. tris(2-chloroethyl) phosphate) which were detected in 60% or more of the wristbands. For compounds that were measured below the limits of detection, a value was assigned which was equivalent to the LOD divided by the square root of 2. The concentration of the chemical detected in the wristband was then divided by the number of days the wristband was reported to be worn which resulted in a unit of nanograms per gram silicone per day (ng/g-day). All the calibration standards were within 15% of the true value for all compounds on 12 separate days indicating good instrument performance. Information describing the limit of detection and quality control measurements for these samples has been described in detail elsewhere [34].

Congeners in the same class (e.g. 4 BFRs and 7 OPFRs) were highly correlated with each other (ρ spearman > 0.40). Thus, we created a sum score for the different flame retardant classes and used this as our exposure index. Subsequently, ƩPBDEs is the total amount of PBDE-47, PBDE-99, PBDE-153, PBDE-154, PBDE-49, and PBDE28 + 33; whereas ƩOPFRs is the total amount of TDCPP [e.g. tris(1,3-dichloro-2-propyl) phosphate], TPP [e.g. triphenylphosphate], TCPP [e.g tris(1-chloro-2-propyl) phosphate], and TCEP [e.g. tris(2-chloroethyl) phosphate].

Covariates

The primary caregiver of the child was asked to complete several structured questionnaires to collect information about socio-demographics and aspects of the home environment. Six variables were aggregated into a covariate to represent the family context: maternal education, paternal education, maternal employment, paternal employment, household income, and home learning environment. Parents reported their total years of education (e.g. 12 = completed high school; 16 = 4-yr college degree, etc.). Parents who reported that they were employed either part- or full-time were coded as “1”; others were coded as “0” for not employed. Annual household income was reported on a scale from 1 = less than 22,000 to 8 = 70,001 or more. The home learning environment was measured with 14 items from the Parenting Questionnaire [40] related to literacy and numeracy activities in the household (e.g. how often do you read to your child? How often do you encourage your child to do math-related activities?). Items were standardized and aggregated into a home learning composite (Cronbach’s alpha = 0.83). Values for all six variables were standardized and averaged (Cronbach’s alpha = 0.74). Previous research has documented similar internal reliabilities for the Parenting Questionnaire [40, 41].

Parents were also asked to self-report if their child had ever experienced any of the four following adverse experiences since birth: lived with an adult that had problems with alcohol or drugs or substance abuse, lived with an adult that was depressed or mentally ill, or attempted suicide, experienced violence or trauma (physical, psychological, or sexual abuse) or neglect, witnessed domestic violence. The total number of items children were reported to have experienced was summed into an adverse experience covariate (range: 0 to 4).

Statistical analysis

Descriptive statistics were used to explore the data and calculate averages, spearman correlations, and percentiles of selected characteristics. The distributions of ΣBDE and ΣOPFRs were right skewed and subsequently natural log transformed. We then analyzed the data in two steps. First, we conducted multiple regression analysis to test for linear associations between levels of lnΣBDE or lnΣOPFR and subscales of social behaviors using Mplus version 6.0 with full information maximum likelihood estimation (FIML) [42]. These models were adjusted for children’s age, gender, flame retardant exposure, and family context. While the two flame retardant exposure variables were modestly correlated (ρ = 0.24) including both exposures in the model did not inflate standard errors by more than 5%. We then utilized generalized additive models (GAM) to explore non-linear associations between lnΣBDE or lnΣOPFR and subscales of social behaviors using R version 3.3 [43]. The upper limit on the degrees of freedom was set at k-1 to relax any assumptions about the shape of the exposure-response curve. Finally, all models were run with and without the flame retardant exposure variable to examine the proportion of variance explained by the chemical exposure.