Associate Research Scientist Gastrointestinal Lab, Texas A&M University College Station, TX, United States
Abstract: Background – Indoxyl sulfate (IS) and p-cresol sulfate (PCS) are uremic toxins produced by bacterial breakdown of tryptophan and tyrosine in the gastrointestinal tract followed by sulfotransferase conversions in the liver and gut mucosa. They have been associated with chronic kidney disease (CKD) severity and progression in dogs and cats. Objective – Analytically validate an assay for quantitation of IS and PCS in canine and feline serum.Animals – Excess serum samples submitted for routine diagnostic evaluation from dogs and cats, and serum from CKD cats, were used.Methods – The liquid chromatography tandem mass spectrometry assay was validated according to FDA guidelines for bioanalytical method validation. Isotopically labelled internal standards were utilized for each analyte. Results – The working range of the assay for IS and PCS was 1 - 40,000 ng/ml for both, covering the common biological range in healthy and CKD animals. The assay was precise and accurate (CV% < 12%). Extracted samples were stable on reinjection up to 72 hours (CV% < 10.2%). As little as 10 µl of serum was able to be used with accurate quantitation. Dilution and spiking recovery had observed to expected ratios between 80 - 120% apart from one low spike sample at 130%. Lipemia did not affect measurement in canine serum. Conclusions and clinical importance – The validated assay provides a wide working range. The low sample volume requirement of 10 µl increases the feasibility of routine IS and PCS measurement in dogs and cats with CKD.
