Comparative analysis of creatinine and osmolality as urine normalization strategies in targeted metabolomics for the differential diagnosis of asthma and COPD.

Introduction: Urine is an ideal matrix for metabolomics investigation due to its non-invasive nature of collection and its rich metabolite content. Despite the advancements in mass spectrometry and H-NMR platforms in urine metabolomics, the statistical analysis of the generated data is challenged with the need to adjust for the hydration status of the person. Normalization to creatinine or osmolality values are the most adopted strategies, however, each technique has its challenges that can hinder its wider application. We have been developing targeted urine metabolomic methods to differentiate two important respiratory diseases, namely asthma and chronic obstructive pulmonary disease (COPD).

Objective: To assess whether the statistical model of separation of diseases using targeted metabolomic data would be improved by normalization to osmolality instead of creatinine.

Methods: The concentration of 32 metabolites was previously measured by two liquid chromatography-tandem mass spectrometry methods in 51 human urine samples with either asthma (n = 25) or COPD (n = 26). The data was normalized to creatinine or osmolality. Statistical analysis of the normalized values in each disease was performed using partial least square discriminant analysis (PLS-DA). Models of separation of diseases were compared.

Results: We found that normalization to creatinine or osmolality did not significantly change the PLS-DA models of separation (RQ = 0.919, 0.705 vs RQ = 0.929, 0.671, respectively). The metabolites of importance in the models remained similar for both normalization methods.

Conclusion: Our findings suggest that targeted urine metabolomic data can be normalized for hydration using creatinine or osmolality with no significant impact on the diagnostic accuracy of the model.