Operating Instructions RF-2000
51
6.1 Outline of System Validation
6.1.1 Outline
For the holistic validation of HPLC system, chromatographic analysis is performed
under the analytical conditions specified by the manufacturer. The system status can
be judged based on the obtained results. This is because the failure of the LC system
may depend on the analytical conditions. Holistic validation procedure described in
this manual is the standard for checking the status of the LC system, serving as the
basis of inspection.
In routine operation, the operators must perform a system suitability test under the
pre-determined analytical conditions. If any problem occurs under such conditions,
perform holistic validation first described in this manual. If the result of holistic
validation meets the acceptance criteria, the LC system itself is working properly and
the cause of the problem is related to the analytical method itself. On the other hand,
if the results do not meet the acceptance criteria for holistic validation, it implies that
there is a problem in the LC system itself and modular validation should be
performed for diagnosis of each LC module.
The repeatability (relative standard deviation) of retention time, peak area, and peak
height is measured to check whether the values meet the acceptance criteria.
6.2 Repeatability Test of Chromatographic Data (Isocratic LC)
6.2.1 Purpose
The purpose of this test is to confirm that the chromatographic data can be obtained
with good repeatability for the LC system to be inspected. An HPLC System for this
inspection consists of the pump, detector, column oven, autosampler and data
system.
6.2.2 Preparation for Inspection
(1) Prepare the following parts and reagents:
(a) Isocratic LC system
(b) Mobile phase (acetonitrile/water = 4/1 (v/v)
* Use acetonitrile and water of HPLC grade.
(c) Column
(4.6mm ID X 150mm)
(d) Sample
* Mixture of two components (acridine and anthracene)
[Method for sample preparation]
Place 10mg of acridine and 10mg of anthracene in the volumetric flask of
100ml capacity. Add 100ml acetonitrile and shake it to dissolve. Transfer 1ml
portion of the sample solution into the 10ml volumetric flask and add
acetonitrile, resulting in a total volume of 10ml.
(e) Water (HPLC grade or equivalent)
(f) Isopropyl alcohol
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