Our aim is to develop the best and cheapest methods for steviol glycoside (SVgly) analysis so that even small laboratories as well as food inspectors can easily check the purity of the steviol glycosides. The internal standard (IS) method is the best method for the analysis of samples. The 19-O-β-D-galactopyranosyl-13-O-β-D-glucopyranosyl-steviol was used as an IS, that was previously synthesised and the method was validated. The calibration curves can be easily reproduced by other laboratories and the analyses are very much simplified. The possible errors are much reduced by the IS method compared to the external standard method. The inter-laboratory relative standard deviation (RSD) for the analysis of all the SVglys present was about 1.75%, which is much better than what can be obtained by an external standard method. This value might still decrease after improvement of peak resolution and peak integration techniques in some laboratories. Indeed, the RSD found with the results of the best scoring laboratories was only 0.8%, but to be objective, we had to include all the results obtained. Our method made it possible to inject 5 times more of the same sample, resulting in a more accurate measurement of the small peaks, enhancing overall accuracy without the need of making new calibration curves. In the final protocol that was developed, all possible errors were excluded except for two: the drying and weighing of the analyte. In some laboratories, peak resolution and integration should be improved to increase the precision and accuracy of the analysis. A good chemist and/or company has to be found that can synthesise the IS in large quantities and can make calibration mixtures as well as vials with pre-weighed amounts of IS. This way, the only labour to be done for a SVgly analysis is the drying and weighing of an analyte.
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