The use of low calorie sweeteners (LCS) as substitutes for sugar in food and beverage products has increased as a strategy to reduce the energy density of the diet(Reference Zygler, Wasik and Namiesnik1). As food additives, LCS are assigned an acceptable daily intake (ADI) following examination of available safety and toxicological data. In Europe, member states are required to monitor consumption of LCS to ensure that the ADI is not being exceeded(2). Traditional methods of monitoring are prone to error and as LCS pass through the body largely unchanged(Reference Renwick3–Reference Guens, Buyse and Vankeirsbilck5), the opportunity may exist to use a biomarker approach to investigate levels of exposure to LCS. Although analytical methodologies have been published describing the simultaneous determination of LCS in foodstuffs(Reference Zygler, Wasik and Namiesnik1) as well as waste water(Reference Ordonez, Quintana and Rodil6), to date no such method has been described for use in human urine.
This project aimed to develop a novel method of simultaneously determining four LCS (acesulfame-K, saccharin, cyclamate, sucralose) and the excretory product of a relatively new sweetener, steviol glycosides (steviol glucuronide), in human urine. An LC-MS/MS method was developed to separate and quantify these LCS and steviol glucuronide. Sodium cyclamate d-11 (for cyclamate) and warfarin sodium (for acesulfame-k, saccharin, sucralose and steviol glucuronide) were used as internal standards. Method validation was carried out by assessing linearity and range, limits of detection (LOD), limits of quantification (LOQ), precision and accuracy. Preliminary validation data indicate linearity across a large dynamic range of 10–1000 ng/ml for all five LCS (Table). For each LCS, LOD was below 0·3 ng/ml while LOQ was below 0·7 ng/ml. Accuracy and precision were tested by analysing spiked urine samples at three concentrations within the dynamic range (ie.12·5, 562·5 and 930 ng/ml).
a Triplicate runs carried out at each concentration; values represent the range between concentrations. b Results of analyses undertaken over 3 consecutive days
This novel method represents an opportunity to implement a biomarker approach for objectively assessing dietary exposure to five LCS. Further work is underway to establish the level of sensitivity of the method within the expected range of LCS observed in free-living populations.
This work was supported by the National Institute for Public Health and the Environment (RIVM) in the Netherlands.
