Samples and sample preparation for biomedical HPLC

The bioanalyst can be required to analyse most biofluids although the most common are urine and the aqueous phase of blood, i.e. plasma or serum. Other samples may be cell and tissue extracts, synovial fluid, cerebrospinal fluid (CSF) and saliva. In the case of urine and CSF with their very low protein content it might be possible to directly inject the sample into an HPLC column. With most silica-based packing materials, direct injection of blood proteins will rapidly lead to column deterioration. HPLC columns are expensive and their efficiency is easily lost so correct preparation of samples will not only improve column life but also improve the results. At its simplest it is only necessary to remove particulate matter from samples to prevent clogging of the column and frits. Modern HPLC packings are very susceptible to contamination by proteins, fats and other macromolecules from biological samples and it is necessary to remove these (except of course for protein analysis). 

Extraction techniques should fulfil the following criteria to be ideal. 

It should release all of the analyte(s) of interest from the sample including any that is protein bound. 

It should precipitate all proteins in order to avoid detrimental chromatographic effects. 

It should neither degrade the analytes chemically nor permit biochemical degradation. It should not degrade polymeric species and thereby increase the concentration of monomers. 

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