Stability incurred sample

Additional Components Analyte recovery (extraction efficiency) Stock stability Matrix stability Post process stability (extract) Integrity of dilution Carryover and contamination Matrix effects Incurred sample reproducibility Incurred sample stability... 

Incurred Sample Stability (ISS) A proper evaluation of incurred sample stability (ISS) needs to be performed on the study samples from each species for GLP toxicology studies as well as on samples... 

Results from these samples should be recorded continuously, and the data are used to verify that the test works reliably. The choice of analytes to include in routine QC samples should follow the same rules as those selected for the initial or abridged validation exercise, that is, the worst-case analytes that are listed in the method scope or the most relevant analytes in a national control plan. Even if the use of spiked samples as QC is applicable, it is highly preferable to use incurred samples where possible. QC samples should be stored for a period determined by the laboratory according to stability data available for the analyte/matrix. The data obtained with the QC samples should be stored and remain traceable as long as the method is used in the laboratory. The results obtained from the QC samples should be used to supplement the initial and abridged validation data. 

Investigate the stability of the incurred sample both before and after sample preparation. This step is intended to check for the possibility that a metabo-hte can imdergo degradation or even reversion back to the original analyte during sample preparation and before LC-MS analysis. 

The stability of the analyte(s) in matrix during extended freezer storage should be determined by analyzing stored stability QC samples at appropriately selected time intervals and at temperatures that reflect the intended storage conditions and anticipated storage periods for study samples. For many applications, the compound is determined to be stable as long as the calculated concentration of the analyte is within 15 % of the nominal concentration, or the concentration that was established for the same batch of QCs when analyzed immediately after preparation (time zero). Freshly prepared QC samples, or QC samples prepared and stored within an established period of stability, should be used as analytical QCs in the same set that the stability QCs are analyzed to confirm run acceptance. If the analytical QC samples do not meet assay acceptance criteria, the run should be rejected and the stability interval should be repeated. Incurred samples or sample pools may also be analyzed for assessment of stability in a similar manner. 

The fundamental components of a bioanal5dical validation (and aU validations in general) are selectivity sensitivity accuracy precision range of reliable response and linearity and reproducibility. Other characteristics of the method that should be addressed are stability carryover and control of lab contamination and matrix effects, including interferences from metabolites in incurred samples or other co-extracted compounds, as well as ionization suppression. 

Further implications of differences between incurred bioanalytical samples and calibrators and QC samples prepared in accord with accepted practice (FDA 2001) have been discussed (Gallicano 2006). It was emphasized that if the imprecision of data obtained for incurred samples differs from that of spiked samples used in the original validation, this will lead to questioning the estimates of accuracy, LLOQ and stability for the incurred samples. If the incurred sample data are of low precision it will be necessary to analyze more samples to investigate these parameters. It was proposed (Gallicano 2006) that an LLOQ higher than the lowest standard concentration (LSC) should be considered if the incurred samples show more imprecision than spiked samples, because the main criteria for selection of the LSC is its precision on the basis of spiked samples . 

A second approach to determining freezer storage stability involves the reanalysis of incurred residues found in actual samples that are stored over time. Using this approach, soil from an actual field sample containing residues is periodically analyzed... 

To determine the stability of albendazole residues in milk upon storage at 18 C from 3 to 8 months, both incurred and spiked milk samples containing all three albendazole metabolites, namely albendazole sulfoxide, albendazole sulfone, and albendazole 2-aminosulphone, were used. Results showed that the 3 month storage did not produce any change in the concentration of the analytes... 

The effect of heating on aqueous solutions of ivermectin is difficult to study due to the very low solubility of the compound in water. Thus, the stability of ivermectin has been only investigated on incurred tissue samples. When a series of incurred pig and cattle muscle and liver samples and salmon muscle samples were subjected to various cooking processes, some loss of ivermectin was observed but the loss was associated with the leakage of the fat from the tissue samples... 

A number of studies have highlighted the degradation of residues during frozen storage, including P-lactam antibiotics in milk ampicillin in pig muscle chlortetracycline in incurred pig muscle, liver, and kidney " sulfamethazine in incurred pig muscle and bovine milk and gentamicin residues in egg. The EU validation criteria require that stability be determined for the analytes in matrix and in solution at various stages of the sample preparation process. Preferably, incurred tissue should be used otherwise... 

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