Untargeted Metabolomic Profiling Using UHPLC-QTOF/MS Reveals Metabolic Alterations Associated with Autism

Abstract

Autism spectrum disorder (ASD) is a clinical spectrum of neurodevelopment disorder characterized by deficits in social communication and social interaction along with repetitive/stereotyped behaviors. The current diagnosis for autism relies entirely on clinical evaluation and has many limitations. In this study, we aim to elucidate the potential mechanism behind autism and establish a series of potential biomarkers for diagnosis. Here, we established an ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry- (UHPLC-QTOF/MS-) based metabonomic approach to discriminate the metabolic modifications between the cohort of autism patients and the healthy subjects. UHPLC-QTOF/MS analysis revealed that 24 of the identified potential biomarkers were primarily involved in amino acid or lipid metabolism and the tryptophan kynurenine pathway. The combination of nicotinamide, anthranilic acid, D-neopterin, and 7,8-dihydroneopterin allows for discrimination between ASD patients and controls, which were validated in an independent autism case-control cohort. The results indicated that UHPLC-QTOF/MS-based metabolomics is capable of rapidly profiling autism metabolites and is a promising technique for the discovery of potential biomarkers related to autism.

Figure 1

PCA scatter plot of quality controls (QCs) and controls injected during each batch analysis. (a) Scatter plot for autism patients and QCs are, respectively, colored with blue dots and red triangles. (b) Scatter plot for controls and QCs are, respectively, colored with green circles and red triangles.

Figure 2

(a) OPLS score plot models of autism patients vs. normal controls. R²X = 0.393, R²Y = 0.894. (b) Plot of R²Y and Q² from permutation tests in OPLS-DA models.

Figure 3

Diagnostic performances of the urinary metabolite parameters in ASD. (a) ROC curve analysis: nicotinamide, anthranilic acid (vitamin L1), D-neopterin, and 7,8-dihydroneopterin as biomarkers of ASD. (b) ROC curve cross-validation based on the combined metabolic biomarker panel could significantly discriminate autism from a healthy individual. AUC: area under the ROC curve; ROC: receiver operating characteristic.