Development of a novel method for the simultaneous detection of trimethylamine N-oxide and creatinine in the saliva of patients with chronic kidney disease – Its utility in saliva as an alternative to blood

Chronic kidney disease (CKD) is associated with increased levels of creatinine and other uremic toxins (UTs), which impaired kidneys cannot filtrate. Typically, CKD is diagnosed by calculating the estimated glomerular filtration rate using serum creatinine or cystatin C levels. In pursuit of more sensitive and reliable biomarkers of kidney dysfunction, scientific attention has turned towards other UTs, such as trimethylamine N-oxide (TMAO), successfully quantified in standard matrices, blood and urine. However, less invasive monitoring of kidney function can be performed using an alternative diagnostic biofluid, saliva, which has been shown to contain clinically relevant concentrations of renal function markers. Accurate quantitative estimation of serum biomarkers using saliva measurements can only be achieved provided that there is a tight saliva-serum correlation for the analyte of interest. Therefore, we aimed to verify the correlation between saliva and serum levels of TMAO in CKD patients using newly developed and validated quantitative liquid chromatography coupled to mass spectrometry (LC-MS) method for simultaneous detection of TMAO, and creatinine – the conventional marker of renal impairment. Secondly, we applied this method to quantify TMAO and creatinine levels in the resting saliva of CKD patients collected with a standardised method involving swab-based collectors. A good linear correlation was obtained between the concentration of creatinine in serum and resting saliva of CKD patients (r = 0.72, p = 0.029) and even better in the case of TMAO (r = 0.81, p = 0.008). The analysed validation criteria were fulfilled. No significant influence of the type of swab in the Salivette® device on creatinine and TMAO concentrations in saliva was detected. Our study indicates that saliva can be successfully used in the non-invasive monitoring of renal failure in CKD by measuring salivary TMAO concentrations.