A proof-of-concept study on molecular fluorescence spectroscopy (EEM) combined with chemometrics applied to screening for autism spectrum disorder

Autism Spectrum Disorder (ASD) is a heterogeneous set of neurodevelopmental conditions characterized by deficits in social interaction and communication. ASD diagnosis is exclusively clinical, and in the absence of validated biomarkers, early and minimally invasive screening methods become essential. This study investigated the use of excitation-emission matrix (EEM) molecular fluorescence spectroscopy, combined with chemometric approaches, for the purpose of discriminating between neurotypical (n = 18) and neurodivergent (n = 17) children from saliva samples.

The spectra were evaluated by PARAFAC-QDA, Tucker3-QDA, and nPLS-DA classification methods. All models showed accuracy above 80.0%, with nPLS-DA having the best performance, with an accuracy of 90.0%, precision and specificity of 100.0%, sensitivity of 80.0%, F1 score and Fss score of 88.9%, and G score of 89.4%. The most relevant spectral bands selected by the models indicated the contribution of tryptophan, vitamin B6 (pyridoxine), and compounds associated with protein modification, such as dityrosine and advanced oxidation protein products (AOPPs), suggesting potential salivary biomarkers.

The findings reinforce the potential of EEM fluorescence spectroscopy combined with second-order algorithms as a non-invasive tool to support ASD screening and monitoring. Future studies with larger cohorts are essential for clinical consolidation of this methodology. The authors acknowledge the support provided by the Post-Graduate Chemistry Program (PPGQ/UFRN) and the Analytical Center of the UFRN Institute of Chemistry (IQ/UFRN) for providing analytical instruments, to the Neuropediatrics Outpatient Clinic of the Onofre Lopes University Hospital (HUOL), the Integrated Clinic of the Directorate of Attention to the Health of the Server (DAS/UFRN), the Januário Cicco Maternity School (MEJC-UFRN), the Specialized Center for Child Rehabilitation (CERAI), the Early Childhood Education Center (NEI/UFRN), as well as the children and their families for their voluntary participation and collaboration.

T. de (A) Silva thanks the National Council for Scientific and Technological Development (CNPq) [process numbers: 173825/2023-0 and 178008/2024-9 (PDJ)] and the National Institute of Science and Technology in Molecular Sciences (INCT-CiMol, CNPq 406804/2022-2) for funding the project. Câmara would like to thank the coordenação de Aperfeiçoamento de Pessoal de Nível Superior by the Programa Institucional de Pós-Doutorado (CAPES/PIPD-PPGQ, Brazil), code 88887.082609/2024-00 for the financial support. Biological Chemistry and Chemometrics, Institute of Chemistry, Federal University of Rio Grande do Norte, Natal, Rio Grande do Norte, 59078-900, Brazil Hellyda K.

Silvestre, Ricardo F. dos Santos, Anne B. Lima Postgraduate Program in Health Sciences, Federal University of Rio Grande do Norte, Natal, Rio Grande do Norte, 59078-900, Brazil Shayanne M. F. de Araujo & Rodrigo Pegado Federal Institute of Education, Science and Technology of Rio Grande do Norte, Natal, Rio Grande do Norte, 59015-000, Brazil Center for Education, Science and Technology of the Inhamuns Region, State University of Ceará, Tauá, Ceará, 63660-000, Brazil Department of Physical Therapy, Health Sciences Center, Federal University of Rio Grande do Norte, Natal, Rio Grande do Norte, 59078-900, Brazil Edmond and Lily Safra International Institute of Neuroscience, Santos Dumont Institute, Macaíba, Rio Grande do Norte, 59288-999, Brazil Postgraduate Program in Physical Therapy, Federal University of Rio Grande do Norte, Natal, Rio Grande do Norte, 59078-900, Brazil The authors declare no competing interests.

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Below is the link to the electronic supplementary material. Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if you modified the licensed material.