Optimization and validation of a method for the determination of the refractive index of milk serum based on the reaction between milk and copper(II) sulfate to detect milk dilutions

•Determination of the refractive index of milk serum by copper(II) sulfate method.•A method to detect milk dilutions was modernized and it is discussed in details.•The method was optimized and validated, making it simple, fast and robust.•Optimized method showed an accuracy of 0.4% and a quadrupled analytical throughput.

We report the use of a method to determine the refractive index of copper(II) serum (RICS) in milk as a tool to detect the fraudulent addition of water. This practice is highly profitable, unlawful, and difficult to deter. The method was optimized and validated and is simple, fast and robust. The optimized method yielded statistically equivalent results compared to the reference method with an accuracy of 0.4% and quadrupled analytical throughput. Trueness, precision (repeatability and intermediate precision) and ruggedness are determined to be satisfactory at a 95.45% confidence level. The expanded uncertainty of the measurement was ±0.38 °Zeiss at the 95.45% confidence level (k=3.30), corresponding to 1.03% of the minimum measurement expected in adequate samples (>37.00 °Zeiss).

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