Define the scope of the method to be validated and quantify the objectives
The accuracy profile method

As an illustration, we will use the validation study of a method for determining nicotinamide (a vitamin B 3 vitamin) in milk by HPLC, with quantification by external calibration.

• The natural content of nicotinamide in milk is between 0.6 and 1.0 mg L -¹ ; regulations authorize supplementation, ranging from 0.4 to 2.0 mg L -¹ . The chosen validation range is therefore between 0.4 and 4.0 mg L -¹ .

• There is no official reference for setting the acceptability limits of this method. However, the microbiological reference method has a high uncertainty of ± 40%. To demonstrate that the HPLC method can easily be an alternative to the reference method, acceptability limits have been set at just ± 15%.

• It was decided to set the β proportion at 90%. As we shall see, this means that, eventually, 1/20 of the – quality...