Dried Milk Spots: A viable approach for assessing the chemical exposome in mothers and their infants by targeted LC-MS/MS

Breast milk is a key matrix for assessing early-life exposure. Dried milk spots (DMS) and microsampling devices are convenient low-volume sampling alternatives. Here, a sample preparation protocol and LC-MS/MS meth-od for (semi-)quantitively assessing >200 xenobiotics in DMS were optimized and thoroughly evaluated. Two extraction solutions were compared. Both approaches performed similarly, with about 50% of analytes falling within the assigned acceptance range for matrix effects (60-140%), and about 80% fulfilling the proposed recovery criteria (42-134%). In a proof-of-principle study, the method was applied to a pooled Austrian milk sample as well as to the NIST standard reference material SRM 1954 (pooled breast milk from US donors). A total of 30 exposure compounds were detected in SRM 1954, 22 of which were also found in the Austrian pooled sample. Compounds were mostly detected at very-low trace levels and included air pollutants (co-tinine), plastics-related chemicals (phthalates, bisphenols), flame retardants (TBBPA, TCBPA), perfluoroalkyl substances (PFOA, PFOS), personal care products ingredients (parabens) and pharmaceuticals (acetamino-phen, fluconazole). The stability of analytes was assessed in DMS at -20, 4, 18 and 37°C for up to two months. No significant changes were observed during storage at -20°C regardless of storage time, while short-term stability was confirmed for approximately 80% of all tested exposure chemicals even at more elevated tem-peratures. A comparison between DMS and Mitra volumetric absorptive microsampling (VAMS) devices showed comparable performance but differences in background contamination. The developed assay is fit-for-purpose enabling broad exposome-type population studies for investigating early-life exposure patterns.