A single measurement of organophosphate flame retardant (OPFR) metabolites in a spot sample is often used in epidemiological studies to estimate individual exposures. Over seven consecutive days, we collected 661 spot samples, including 127 first morning voids (FMVs) and 123 simulated 24-h collections, from 20 healthy adults and analyzed for eight OPFR metabolites. Intraclass correlation coefficients (ICCs) were calculated to evaluate the variability of the analyzed metabolites. In spot samples group, serial measurements of OPFR metabolites showed poor reproducibility (0.0422 ≤ ICC ≤ 0.349), and the within-day variability was the main contributor of the total variability. The estimated ICCs based on different correction methods for urine dilution (i.e., specific gravity-adjusted, creatinine-adjusted, and creatinine as a covariate) were similar, but varied according to gender and body mass index. Uniformly low sensitivities (0.417–0.633) were observed when using a single FMV or spot sample to predict the 1-week highly (top 33.0%) exposed volunteers. Therefore, using a single urinary measurement to predict chronic exposure to OPFRs can lead to a high degree of classification errors. When multiple urine samples are collected, considering the sampling type, the time of collection, and demographic characteristics may provide a more complete approach to assess exposure to diverse OPFRs.
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