Instruction manual

7.6 System Validation
7-41
RF-20A/20Axs
7
7.6.2 Validation of a Gradient LC System
 Objective
An analysis is performed on the LC system being validated and the retention time and peak area are obtained
for each peak. The data is then examined to check for reproducibility. Reproducible data validates the system.
Generally, the system being validated consists of a minimum of the following components: a pump, column
oven, autosampler, detector, system controller and data processor.
 Equipment Required for Validation
Part Name Description
Mobile phase
A: Water
B: Methanol
A / B = 10 % / 90 %
* Both the water and the methanol should be HPLC grade.
Column
Shim-pack VP-ODS (Part No. 228-34937-91), LUNA C18 (2) (Part No. 00F-4252-E0) or equivalent
ODS column
Particle size: 5 μm
Column dimension: I.D. 4.6 mm × length 150 mm
Sample
1 mg/L solution of anthracene in acetonitrile (part No.: 228-32996-15)
<Preparation Procedure>
1. Weigh 20 mg of anthracene and place it in a 200-mL measuring flask.
2. Dissolve the anthracene in acetonitrile, and bring the total volume up to 200 mL.
3. Measure out 1 mL of the solution prepared in step 2 and place it in a 100-mL measuring flask.
4. Add acetonitrile to the solution in the measuring flask in step 3 to bring the total volume up to
100 mL.
Methanol HPLC grade, or equivalent
2-propanol HPLC grade, or equivalent