Determination of proline in honey: Comparison between official methods, optimization and validation of the analytical methodology

The study compares official spectrophotometric methods for the determination of proline content in honey – those of the International Honey Commission (IHC) and the Association of Official Analytical Chemists (AOAC) – with the original Ough method. Results show that the extra time-consuming treatment stages added by the IHC method with respect to the Ough method are pointless. We demonstrate that the AOACs method proves to be the best in terms of accuracy and time saving. The optimized waiting time for the absorbance recording is set at 35 min from the removal of reaction tubes from the boiling bath used in the sample treatment. The optimized method was validated in the matrix: linearity up to 1800 mg L-1, limit of detection 20 mg L-1 , limit of quantification 61 mg L-1. The method was applied to 43 unifloral honey samples from the Marche region, Italy.