主题：【第二届原创作品大赛】基于ICH Q2R的分析方法验证经典案例（英文）

3. Validation of the analytical procedure for test of the residue of XXXXXX

3.1 Summary of the validation results

The validation characteristics, validation results and acceptance criteria are summarized in the following table.




3.2 Validation results for test of the residue of XXXXXX in YYYYYY

System suitability

Following the analytical procedure, inject the reference solution for six times and get the chromatograms. Calculate symmetry factor of the peak due to XXXXXX, resolution between the peaks due to XXXXXX and the adjacent peak, and RSD of the peak areas of the six injections. The results are listed in the following table.


The results are well within the acceptance criteria.

Specificity

Spiked test solution: Transfer about 2.500g of the sample, accurately weighed, to a 25-ml volumetric flask, dissolve in and dilute with the Reference stock solution to volume, and mix.
Test solution: Prepare the Test solution according to the Analytical procedure.
Reference solution: Prepare the Reference solution according to the Analytical procedure.
Blank solution: Prepare the Blank solution according to the Analytical procedure.

Following the analytical procedure, inject the Blank solution,  Test solution,  Spiked test solution and Reference solution into the chromatograph separately. The retention times and relative retention times of representative peaks in the chromatogram obtained with the Spiked test solution are presented in the following table.



Limit of Quantitation (LOQ)

Prepare the Reference stock Solution according to the Analytical Procedure. Dilute the solution with DMF to obtain a certain concentration. Transfer 5.0ml of such dilution and 1.0ml of purified water to a 20-ml headspace vial and mix well. Inject the solution. The results are presented in the following table.The signal-to-noise ratio should be not less than 10/1.



Limit of Detection (LOD)

Prepare the Reference stock Solution according to the Analytical Procedure. Dilute the solution with DMF to obtain a certain concentration. Transfer 5.0ml of such dilution and 1.0ml of purified water to a 20-ml headspace vial and mix well. Inject the solution. The results are presented in the following table. The signal-to-noise ratio should be not less than 3/1.



Linearity

Reference stock Solution: Transfer about 1.000g of XXXXXX(HPLC),accurately weighed,to a 100-ml volumetric flask, dilute with DMF to volume, and mix. Dilute 1.0ml of this solution to 100-ml volumetric flask with DMF, Dilute 1.0ml of this solution to 50-ml volumetric flask with DMF.

Reference solution: Dilute the Reference stock Solution to produce solutions with a relative concentration of 20%, 40%, 70%, 100%, 120% of the residual limit of XXXXXX. Transfer 5.0ml of such dilutions and 1.0ml of purified water to a 20-ml headspace vial and mix well, 3 preparations in parallel.

Inject the solutions and LOQ solution into the chromatograph separately and record the chromatogram. The results are listed in the following table.

Note: If the concertration of LOQ is larger than that of 20% level, Replace the 20% level with the LOQ.


Calculate the regression equation and draw the diagram as below:



The above results show that the analytical method has a good linearity for XXXXXX at the concentration range of 0.042ug/ml to 0.24ug/ml, corresponding to 0.42ppm to 2.4ppm of XXXXXX in YYYYYY .

Precision

System precision

Prepare the Reference Solution according to the Analytical Procedure. Perform six injections. Record the area of the peak of XXXXXX in the chromatogram obtained from each injection. The peak areas are listed in the table below. Calculate the relative standard deviation and 95% confidence interval as below.



Method precision

Reference stock solution: Transfer about 1.000g of XXXXXX(HPLC),accurately weighed,to a 100-ml volumetric flask, dilute with DMF to volume, and mix. Dilute 1.0ml of this solution to 100-ml volumetric flask with DMF, Dilute 1.0ml of this solution to 100-ml volumetric flask with DMF.

Spiked test solution: Transfer 2.500g of the sample, accurately weighed, to a 25-ml volumetric flask, add 5.0ml of Reference stock solution,dissolve in and dilute with DMF to volume, and mix. Transfer 5.0ml of such solution to a 20-ml headspace vial, add 1.0ml of water, apply stopper ,cap, and mix.

Prepare the Blank solution, test solution  and Reference Solution according to the Analytical Procedure.Perform six preparations of Spiked test Solution. each preparation inject once. Record the area of the peak of XXXXXX in the chromatogram obtained from each injection. Concurrently, inject the test solution and Reference solution three times separately. Record the chromatograms. The peak areas and calculated contents of XXXXXX in Spiked test Solution are listed in the tables below. The chromatograms are presented in the annex of the response dossier. Calculate the relative standard deviation and 95% confidence interval.



Interm-precision

Spiked test solution : Prepare the Spiked test solution according to method precision.

Prepare the Blank solution, Reference Solution  and test solution according to the Analytical Procedure.

Perform six preparations of Spiked test Solution. each preparation inject once. Record the area of the peak of XXXXXX in the chromatogram obtained from each injection. Concurrently, inject the test solution and Reference solution three times separately. Record the chromatograms. The peak areas and calculated contents of XXXXXX in Spiked test Solution are listed in the tables below. Calculate the relative standard deviation and 95% confidence interval.

     
Accuracy


Perform three preparations for each solution and inject each preparation once.
Inject the blank solution, Test solution, the series concentration of the Spiked test solution and the corresponding reference solution separately. The recovery percent should be within 90%~110%. The results are listed in following table.
Recovery percent of XXXXXX
The peak areas of XXXXXX are listed in the table below, and the recovery rate of XXXXXX is calculated according to the following formula:


Conclusion: The recovery results for XXXXXX are well with the acceptance criteria 90.0% -110.0%.

Robustness

Prepare the Reference Solution according to the Analytical Procedure.

Test parameters are deliberately changed to assess robustness of the method. Peak area and retention time of XXXXXX is chosen to evaluate the effect which results from the test condition variation