# How to prepare solutions of 80% to 150% in recovery test

Can any one tell me that how can i prepare solution of 80% to 150% briefly in recovery test in Assay method validation if my method of preparation is 100 mg to 100 ml —> 5 ml to 50

Dear, Recovery study done on finished product if available. If FP contain 100 mg of API, for 80% take equivalent wt of FP contain 80 mg of API make dilution as per method. For API no need to do recovery study. For impurity make stock solution and spicke as per validation protocol.

Based on the information you provided, it looks like your nominal sample concentration is 100 µg/mL.

So your target concentrationsat 80% and 150% levels should be 80 µg/mL and 150 µg/mL, respectively.

In order to prepare these samples, you will need to prepare one stock solution at a much higher concentration level than your nominal concentration stated in the method. For example, you can use the stock solution you initially prepared at 1000 µg/mL (where you transferred 100 mg into 100 mL) before doing secondary dilution.

So,
To prepare sample at 80% level: Pipet 8 mL of your 1000 µg/mL stock solution into a 100 mL volumetric flask and Q.S with your diluent (or alternatively, pipet 4 mL into 50 mL vol. flask)

To prepare sample at 150% level: Pipet 15 mL of your 1000 µg/mL stock solution into a 100 mL volumetric flask and Q.S with your diluent.

for 80% solution : 100 mg to 100 ml —> 4 ml to 50

for 150% solution : 100 mg to 100 ml —> 7.5 ml to 50

For assay where the sample preparation step is critical, ie. involved extraction or solubilization procedure, it is recommended to use samples that contains placebo and API in different ratio. Placebo is the mixture of components other than analyte(s).
In the topic starter case, the preparation procedure was 100 mg to 100 ml —> 5 ml to 50. 100 mg maybe refer to the amount of API. Let’s say the total weight of product equivalent to 100 mg is 200 mg (API-placebo ratio=1:1).
For 80% recovery samples we could make a mixture API-placebo in ratio 80:120.
For 150% recovery samples we could make a mixture API-placebo in ratio 150:50

For dissolution and related compound, spiking procedure could be used.