Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Optimization of a method for collecting infant and toddler urine for non-target analysis using cotton pads and commercially available disposable diapers



Urine is an abundant and useful medium for measuring biomarkers related to chemical exposures in infants and children. Identification of novel biomarkers is greatly enhanced with non-targeted analysis (NTA), a powerful methodology for broad chemical analysis of environmental and biological specimens. However, collecting urine in non-toilet trained children presents many challenges, and contamination from specimen collection can impact NTA results.


We optimized a caregiver-driven method for collecting urine from infants and children using cotton pads and commercially available disposable diapers for NTA and demonstrate its applicability to various children biomonitoring studies.


Experiments were first performed to evaluate the effects of processing method (i.e., centrifuge vs. syringe), storage temperature, and diaper brand on recovery of urine absorbed to cotton pads. Caregivers of 11 children (<2 years) used and retained diapers (with cotton pads) to collect their child’s urine for 24 h. Specimens were analyzed via a NTA method implementing an exclusion list of ions related to contamination from collection materials.


Centrifuging cotton pads through a small-pore membrane, compared to a manual syringe method, and storing diapers at 4 °C, compared to room temperature, resulted in larger volumes of recovered sample. This method was successfully implemented to recover urine from cotton pads collected in the field; between 5–9 diapers were collected per child in 24 h, and the total mean volume of urine recovered was 44.7 (range 26.7–71.1) mL. NTA yielded a list of compounds present in urine and/or stool that may hold promise as biomarkers of chemical exposures from a variety of sources.

Impact Statement

Infant and children urine is a valuable matrix for studies of the early life exposome, in that numerous biological markers of exposure and outcome can be derived from a single analysis. Depending on the nature of the exposure study, it may be the case that a simple collection method that can be facilitated by caregivers of young children is desirable, especially when time-integrated samples or large volumes of urine are needed. We describe the process for development and results of an optimized method for urine collection and analysis using commercially available diapers and non-target analysis.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type



Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Summary of analyses for selecting brand of hypoallergenic disposable diaper for non-targeted analysis of children’s urine.
Fig. 2: Laboratory testing of diapers to assess specimen recovery.
Fig. 3
Fig. 4: Principal component analysis of participant specimens analyzed via LC-HRMS.

Data availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.


  1. Cohen Hubal EA, Sheldon LS, Burke JM, McCurdy TR, Berry MR, Rigas ML, et al. Children’s exposure assessment: a review of factors influencing children’s exposure, and the data available to characterize and assess that exposure. Environ Health Perspect. 2000;108:475–86.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Moya J, Phillips L. A review of soil and dust ingestion studies for children. J Expo Sci Environ Epidemiol. 2014;24:545–54.

    Article  PubMed  Google Scholar 

  3. U.S. EPA. Exposure factors handbook chapter 5 (Update): soil and dust ingestion. Washington, DC, U.S. EPA Office of research and development; 2017.

  4. Binder S, Sokal D, Maughan D. Estimating soil ingestion: the use of tracer elements in estimating the amount of soil ingested by young children. Arch Environ Health. 1986;41:341–5.

    Article  CAS  PubMed  Google Scholar 

  5. Calabrese EJ, Stanek EJ. Resolving intertracer inconsistencies in soil ingestion estimation. Environ Health Perspect. 1995;103:454–7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Panagopoulos Abrahamsson D, Sobus JR, Ulrich EM, Isaacs K, Moschet C, Young TM, et al. A quest to identify suitable organic tracers for estimating children’s dust ingestion rates. J Expo Sci Environ Epidemiol. 2020;31:70–81.

  7. Herreros ML, Gili P, del Valle R, Barrios A, Pacheco M, Sánchez A. Urine collection methods for infants under 3 months of age in clinical practice. Pediatr Nephrol. 2021;36:3899–904.

    Article  PubMed  Google Scholar 

  8. Hu Y, Beach J, Raymer J, Gardner M. Disposable diaper to collect urine samples from young children for pyrethroid pesticide studies. J Expo Anal Environ Epidemiol. 2004;14:378–84.

    Article  CAS  PubMed  Google Scholar 

  9. Ueyama J, Aoi A, Ueda Y, Oya N, Sugiura Y, Ito Y, et al. Biomonitoring method for neonicotinoid insecticides in urine of non-toilet-trained children using LC-MS/MS. Food Addit Contam Part A, Chem, Anal, control, Expo Risk Assess. 2020;37:304–15.

    Article  CAS  PubMed  Google Scholar 

  10. Oerlemans A, van Dael MFP, Vermeulen RCH, Russel FGM, Scheepers PTJ. Urine collection methods for non-toilet-trained children in biological monitoring studies: validation of a disposable diaper for characterization of tebuconazole exposure. Toxicol Lett. 2018;298:201–6.

    Article  CAS  PubMed  Google Scholar 

  11. Shen M, Zhang C, Yi X, Guo J, Xu S, Huang Z, et al. Association of multi-metals exposure with intelligence quotient score of children: a prospective cohort study. Environ Int. 2021;155:106692.

    Article  CAS  PubMed  Google Scholar 

  12. Farzan SF, Shahriar M, Kibriya MG, Jasmine F, Sarwar G, Slavkovic V, et al. Urinary arsenic and relative telomere length in 5–7 year old children in Bangladesh. Environ Int. 2021;156:106765.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Welch BM, Keil AP, Buckley JP, Calafat AM, Christenbury KE, Engel SM, et al. Associations between prenatal urinary biomarkers of phthalate exposure and preterm birth: a pooled study of 16 US cohorts. JAMA Pediatr. 2022;176:895–905.

    Article  PubMed  PubMed Central  Google Scholar 

  14. Lee EJ, Arbuckle TE. Urine-sampling methods for environmental chemicals in infants and young children. J Expo Sci Environ Epidemiol. 2009;19:625–33.

    Article  PubMed  Google Scholar 

  15. Andra SS, Austin C, Patel D, Dolios G, Awawda M, Arora M. Trends in the application of high-resolution mass spectrometry for human biomonitoring: an analytical primer to studying the environmental chemical space of the human exposome. Environ Int. 2017;100:32–61.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Jackson F, Georgakopoulou N, Kaluarachchi M, Kyriakides M, Andreas N, Przysiezna N, et al. Development of a pipeline for exploratory metabolic profiling of infant urine. J Proteome Res. 2016;15:3432–40.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Goodpaster AM, Ramadas EH, Kennedy MA. Potential effect of diaper and cotton ball contamination on NMR- and LC/MS-based metabonomics studies of urine from newborn babies. Anal Chem. 2011;83:896–902.

    Article  CAS  PubMed  Google Scholar 

  18. Makoś-Chełstowska P, Kurowska-Susdorf A, Płotka-Wasylka J. Environmental problems and health risks with disposable baby diapers: monitoring of toxic compounds by application of analytical techniques and need of education. TrAC Trends Anal Chem. 2021;143:116408.

    Article  Google Scholar 

  19. Broadhurst D, Goodacre R, Reinke SN, Kuligowski J, Wilson ID, Lewis MR, et al. Guidelines and considerations for the use of system suitability and quality control samples in mass spectrometry assays applied in untargeted clinical metabolomic studies. Metabolomics .2018;14:72.

    Article  PubMed  PubMed Central  Google Scholar 

  20. Schymanski EL, Jeon J, Gulde R, Fenner K, Ruff M, Singer HP, et al. Identifying small molecules via high resolution mass spectrometry: communicating confidence. Environ Sci Technol. 2014;48:2097–8.

    Article  CAS  PubMed  Google Scholar 

  21. INnovations to generate estimates of children’s soil/dust inTake (INGEST) 2022. Available from:

  22. Defossez E, Bourquin J, von Reuss S, Rasmann S, Glauser G. Eight key rules for successful data-dependent acquisition in mass spectrometry-based metabolomics. Mass Spectrom Rev. 2021;42:131–43.

  23. Knolhoff AM, Premo JH, Fisher CM. A proposed quality control standard mixture and its uses for evaluating nontargeted and suspect screening LC/HR-MS method performance. Anal Chem. 2021;93:1596–603.

    Article  CAS  PubMed  Google Scholar 

  24. Sobus JR, Grossman JN, Chao A, Singh R, Williams AJ, Grulke CM, et al. Using prepared mixtures of ToxCast chemicals to evaluate non-targeted analysis (NTA) method performance. Anal Bioanal Chem. 2019;411:835–51.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Rampono J, Kristensen JH, Hackett LP, Paech M, Kohan R, Ilett KF. Citalopram and demethylcitalopram in human milk; distribution, excretion and effects in breast fed infants. Br J Clin Pharm. 2000;50:263–8.

    Article  CAS  Google Scholar 

Download references


The authors thank all the caregivers who participated in this study. We thank Brian Caffo for his consultation on and assistance with statistical evaluations. We thank Aimee Bourey for her assistance with sample collection.


This project was supported by a grant from the US Environmental Protection Agency: Estimating Children’s Soil and Dust Ingestion Rates for Exposure Science EPA-G2020-STAR-D1. Matthew N. Newmeyer was supported by NIEHS Training grant (T32 ES 007141).

Author information

Authors and Affiliations



All authors assisted with the design of the study and sample collection. SNL and KEN recruited all participants. QL prepared and processed all biological samples for analysis. MNN conducted non-targeted analysis and data analysis. SNL and MNN wrote the first draft of the manuscript; all authors revised the manuscript. KEN, CP, and SNL acquired funding.

Corresponding authors

Correspondence to Carsten Prasse or Keeve E. Nachman.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplemental Information

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Lupolt, S.N., Newmeyer, M.N., Lyu, Q. et al. Optimization of a method for collecting infant and toddler urine for non-target analysis using cotton pads and commercially available disposable diapers. J Expo Sci Environ Epidemiol 33, 602–609 (2023).

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:


  • Urine biomarkers, non-target analysis
  • disposable diapers
  • children’s exposures


Quick links