A simple, rapid and sensitive HILIC LC-MS/MS method for simultaneous determination of 16 purine metabolites in plasma and urine

Purine intermediates play important roles in physiological function and participate in the kidney disorders, while a targeted quantification of the metabolic alterations in the purine metabolism in acute kidney injury (AKI) individuals has not been conducted. In the study, a novel, rapid and sensitive LC-MS method for simultaneous quantification of 16 purine metabolites was developed using hydrophilic interaction separation mode in human plasma and urine. The developed method was validated by using charcoal-stripped plasma and urine as blank matrix. The results showed that the method was good linear (R2 > 0.99) and the lower limit of quantification (LLOQ) ranged from 0.833 ng/mL to 800 ng/mL. The recovery and matrix effect were repeatable and stable. The intraday precision ranged from 0.7% to 12.7%, while the inter-day precision ranged from 1.6% to 18.5%. Most analytes were stable in the autosampler and could subject three freeze-thaw cycles. The method provided a wider coverage of purine metabolites and completed good separation of interfering compounds of nucleosides, deoxynucleosides and their corresponding nucleobases without derivatization, which was time-saving and labor-saving for the large-scale analysis. Furthermore, the method was successfully applied to plasma and urine samples of hospitalized patients without and with AKI. The results showed certain purine intermediates were up-regulated in plasma and down-regulated in urine of AKI inpatients, indicating that AKI stress may associate with inflammatory responses. The novel method can facilitate the quantitative analysis of purine metabolites in biological fluids, and exhibit great prospects in providing more information on the pathogenesis of AKI.