Autosampler stability

AU stability in matrix and in reconstituted sample extracts (postpreparative stability or autosampler stability) should be assessed at least in three replicates and at two QC, that is, LQC and HQC concentration levels. 

Postpreparative Stability (Autosampler Stability) Stability of processed samples should be determined at the autosampler temperature that will be used during study sample analysis and for a duration of time equal to or greater than the batch size expected to be used for routine study sample analyses. If sample extracts are to be stored (e.g., in a refrigerator or freezer) before analysis, then the stability of the analyte in the reconstituted sample extract under these conditions should be investigated. In postpreparative stability assessment, the mean concentration value obtained from each QC concentration level after storage in autosampler is compared with the respective nominal concentration value. The mean bias (%) between the values should be within +15%. If the stability is demonstrated, then an LC-MS/MS run can be restarted after an unexpected interruption as long as system suitability has been established and documented. 

The nature of the autosampler stability issue is undetermined, especially in a qualitative analysis. It is possible that the analytes adsorb to plastic or crystallize under these reconstitution conditions. 

P 53] Before operation, a start-up time of about 10 min was applied to stabilize pressure in the chip micro reactor ([R 6]) [20]. As a result, a stable flow pattern was achieved. The reactant solutions were filled into vials. Slugs from the reactant solutions were introduced sequentially into the micro chip reactor with the autosampler and propelled through the chip with methanol as driving solvent. The flow rates were set to 1 pi min The slug volume was reduced to 2.5 pi. 

Cool on-column >250 pm column (i.d.) 1 ppm (FID) Reduced thermal degradation and discrimination Wide range of analyte concentrations High sample capacity (LVI) Autosamplers Direct quantification Excellent precision Control of operational conditions (initial oven temperature) Optimisation required Not applicable for polar solvents Column contamination by dirty matrices Poor long term stability... 

Stability — Samples remain stable for at least 468 days when frozen at -20°C. They are stable for at least five simulated freeze-and-thaw cycles and approximately 28 hr at room temperature. The analyte is viable for at least 6 days in a reconstitution solution stored in the autosampler (temperature set point at 10°C). A dried-down batch (sample process stopped at dry-down step) was stable at least 5 days in a refrigerator (temperature varied from 4 to 8°C). A stock solution of paricalcitol is stable for at least 11 months. Stock solution of internal standard is stable about 4.5 months under refrigeration. 

A system is typically comprised of multiple instrument components. Therefore, there is usually an individual IQ for each of these instruments and for any corresponding instrument control/data-handling software. The typical instrument components making up an HPLC system include a binary or quaternary HPLC pump, an autosampler supporting multiple vials or microtiter plates (autosamplers often include cooled Peltier trays for sample stability), a column oven, and a UV-Vis or photodiode array (PDA) detector. 

It is recommended that OQ test the following on an HPLC system flow accuracy, pump compositional accuracy, pressure pulsations, column oven temperature accuracy/stability, detector noise/drift and wavelength accuracy, autosampler injection precision and carryover. 

A key requirement is that the autosampler needs to be temperature stabilized so that the viscosity of the buffer remains consistent over time. Changes in sample or buffer temperature will directly affect the sample injected along with the migration time. Temperature stabilization can be accomplished in a variety of ways including oven and a peltier cooler. 

Postpreparative Stability. The stability of processed samples, including the resident time in the autosampler, should be determined. The stability of the drug and the internal standard should be assessed over the anticipated run time for the batch size in validation samples by determining concentrations on the basis of original calibration standards. Reinjection reproducibility should be evaluated to determine if an analytical run could be reanalyzed in the case of instrument failure. 

As a consequence of the limited stability of the OPA derivatives, the derivatization procedure should be carried out in an intelligent autosampler such as the Gilson 231XL sampler, equipped with a temperature-controlled sample holder (Gilson 832). 

For analytical sample preparation, measurement of the final pH of the sample solution (excipients, API(s), and sample solvent mixed together) will be helpful in the development of any analytical procedure. If an API is known to be stable in acidic pH (pH 1-2), then an analytical chemist will try to utilize a certain sample solvent that has a pH in the required range. However, when a dosage form is dissolved in a sample solvent, the excipients present in the formulation (and even the API) will change the pH of the solution. The final pH of the solution must be measured in order to determine the optimal pH of sample solution to achieve longest solution stability. This is particularly important for a long sequence of injections on autosamplers for analysis, so solutions do not need to be made daily. 

It is necessary to study stability in solution in the solvent used to prepare sample solutions for injection in order to establish that the sample solution composition, especially the analyte concentration, does not change in the time elapsed between the preparation of the solution and its analysis by HPLC. This is a problem for only a few types of compound (e.g. penicillins in aqueous solution) when the sample solution is analysed immediately after the preparation of the sample solution to be injected. The determination of stability in solution is more of an issue when sample solutions are prepared and then analysed during the course of a long autosampler run. While the acceptance criteria for stabUity in solution may be expressed in rather bland terms by making a statement such as, e.g. the analyte was sufficiently stable in solution in the solvent used for preparing sample solutions for reliable analysis to be carried out , in practice it has to be shown that within the limits of experimental error, the result of the sample solution analysis by the HPLC method is the same for injections at the time for which stability is being validated as for injections immediately subsequent to the sample solution preparation. While this may be done by a subjective assessment of results with confidence limits, strictly speaking a statistical method known as the Student s t-test should be used. 

Instability cannot only occur in the sample matrix, but also in prepared samples. It is therefore important to also test the stability of an analyte in the prepared samples under conditions of analysis (e.g. autosampler conditions for the expected maximum time of an analytical run). One should also test the stability in prepared samples under storage conditions, e.g. refrigerator, in case prepared samples have to be stored prior to analysis [4-6,9,10]. 

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