Sample Preparation - Extraction and Clean-Up

The technologies currently avaUable for sample preparation were discussed in Section 4.3 a recent review (Hopfgartner 2003a) provides additional information particularly concerning the robots currently available to assist in high throughput analyses. The present section describes a practical approach to appropriate deployment 

1 10 dilution of the resulting supernatant is suggested as a good starting point for optimization. 

Other aspects that should be checked throughout development include matrix interferences (Section 9.6.3), contamination (either general laboratory sources or arising within the sample preparation process itself) and carryover (Section 9.7) all of these can be evaluated by appropriate use of extracts of analyte-free control matrix. 

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As previously discussed (Section 9.4.7b), extracted control matrix is used to assess the selectivity for the method under development. The selectivity of the method is a function of the sample preparation (extraction and clean-up), chromatography and mass spectrometry conditions that are used for the method. Assuming that the control matrix is representative of the sample matrix to be analyzed, and that method blanks have been used to demonstrate that the method is free of exogenous interferences due to solvents, or to containers or other apparatus (a source of interferences that is often overlooked in the method development process), an extracted blank is used to demonstrate that the method has sufficient selectivity for the intended analytical purpose. When interferences at the expected retention time of the analyte being quantified are detected, modifications to the sample preparation and chromatography (and sometimes the ions monitored hy the method) can be made to improve the selectivity of the method. Recall (Section 9.4.7b) that only re-analysis of incurred samples can reveal interferences resulting from metahoUtes or degradates of the analyte with either or both of the analyte and SIS. 

It is equally important that the sample preparation (extraction and clean-up) of the batch must be done carefully in accordance with the procedmes established by the validated method and any prevailing SOPs. The analysts conducting these tasks must be trained on the applicable SOPs and this training must be documented in their training records. Study samples must be analyzed within known analyte stabihty constraints for the matrix and the processed extracts. When sufficient stability has not been established at the time of sample reporting (e.g. long term freezer stability may be on-going at the time of analysis), this must be clearly documented within the study report. 

The Gilson Aspec automatic sample preparation system is a fully automated system for solid-phase extraction on disposable columns and online HPLC analysis. The Aspec system offers total automation and total control of the entire sample preparation process including clean-up and concentration. In addition, Aspec can automatically inject prepared samples into on-line HPLC systems. 

Advisable for methods including a complex sample preparation procedure like extraction and clean-up... 

Figure 5.1.2 Matrix solid-phase dispersion (MSPD) extraction as a micro-preparative extraction technique for an on-flow LC-NMR-MS screening. Since the latter requires only sample amounts in the 0.5-2 mg range, the sample preparation can be achieved by fast small-scale extraction procedures, such as MSPD. This is a sample preparation technique that combines both sample homogenisation and extraction of compounds of interest in one single step starting from the intact sample material. Thus, it simplifies the extraction and clean-up steps, reduces the sample manipulation and is much faster than conventional techniques. It is therefore very well suited for a rough separation of extracts into classes of compounds of similar polarities, which can then be submitted to LC-NMR-MS analysis...

Preparation of standard, sample extraction and clean up Limit of detection (LOD), limit of quantitation (LOQ), and recovery (Rec) References... 

Limited data were located regarding methods for detecting the hydrocarbon components of gasoline in biota (bivalve mollusks) (Dimock et al. 1980). The methods used were GC (detector not reported) and GC/MS. Sample preparation included tissue digestion, extraction and clean-up, and solvent exchange to hexane. Recovery was poor (-60%). Sensitivity and precision were not reported. 

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