HBM4EU ICI EQUAS report Phthalates in urine round 4

Within the frame of the HBM4EU project, an EQUAS study was organised on the determination of 15 phthalate biomarkers in urine. This was the fourth ICI/EQUAS round for this substance group within the HBM4EU program. In total 33 laboratories were invited for this third round, of which 18 laboratories registered. Results were received from 17 laboratories, located in 12 EU countries and the USA. In November 2019, each participant received two burdened control materials of human urine, A and B (single tube each), containing phthalate biomarkers at various concentrations mostly in the range 1-13 µg/L and 2-22 µg/L, respectively. Homogeneity assessment showed that both materials were sufficiently homogeneous for ICI/EQUAS testing. Previously conducted stability tests had shown that the biomarkers are stable when stored in the freezer and no significant loss of the biomarkers occurs during the course of the EQUAS test. The proficiency of the laboratories was assessed through Z-scores calculated using the mean concentration as established by expert laboratories as assigned value, and a fixed fit-for-purpose relative target standard deviation (FFP-RSDR) of 25%. Assigned values and Z-scores could be determined for all 15 biomarkers in both urine sample A and B. The percentage of satisfactory Z-scores obtained for each biomarker ranged from 64% for MnPePin sample A, to 100% for MiBP and 5OH-MEHP in sample A, and 5oxo-MeHP in both samples. In most cases the satisfactory rate was 80-95%. There were no false positives or false negatives. The number of phthalate biomarkers covered by the different laboratories varied widely: from 3 to all 15 target biomarkers. Thirteen out of 17 labs covered ≥10 biomarkers. MEHP, 5OH-MEHP and 5oxoMEHP were the most frequently determined phthalate biomarkers, followed by MiBP, MnBP, MEP, MBzP, and 5cx-MEPP. cx-MiDP, OH-MiNP, OH-MiDP, and MnPeP were the least frequently determined biomarkers (included in the scope of 8-11 out of the 17 laboratories). Compared to previous rounds, the number of participating laboratories was lower. The overall laboratory performance was similar to that observed in round 3, indicating that the ICI/EQUAS exercises has resulted in a consolidated quality of phthalate biomarker analysis.