Agriculture Reference
In-Depth Information
9.5 Tissue Disruption
Disruption of tissue is normally achieved using a probe blender or through
enzymatic digestion with proteolytic enzymes such as subtilisin A [88].
Several tissue disruption apparatus are available, including probe blenders,
ultrasonic probes and stomachers. The ultrasonic probe uses pulsed, high
frequency sound waves to agitate and disperse cells. The Stomacher® is an
alternative apparatus which extracts residues from samples using crushing
action. Stomachers® offer an advantage over probe blending techniques
because they eliminate the risk of cross-contamination, as each sample is
contained in separate bags [88, 105].
9.6 Clean-up Procedures
Because of the complex nature of the sample matrices, a clean-up step is
required before the chromatographic determination. Sample clean-up
procedures include column chromatography, thin layer chromatography
(TLC), liquid-liquid extraction (LLE), solid phase extraction (SPE) and matrix
solid phase dispersion (MSPD) [94].
9.7 Nitrofurans
Two approaches can be adopted in nitrofuran analysis, specifically,
extraction of total (free and bound) or bound residues. The total residue
approach has been applied by several groups and offers advantages in terms of
speed as it eliminates the need for time-consuming washing of tissue [50, 55,
71, 117-119]. The metabolites (AHD, AOZ, AMOZ and SEM) are usually
released from tissue by acid hydrolysis and derivatised (overnight incubation
with HCl and 2-nitrobenzaldehyde (NBA)). After neutralisation, the
derivatized metabolites are extracted with ethyl acetate and undergo solid-
phase extraction prior to determination by LC-MS/MS. The nitrophenyl
derivatives are used because they are more stable than parent compounds and
also to increase their molecular mass and produce more characteristic ions best
suitable for the identification of the analytes. A particular disadvantage of
analysing total nitrofuran residues is that lower sensitivity can be achieved due
to matrix effects.
As an alternative, the bound residue approach can be adopted. This
involves the intensive washing of tissues to remove free residues and matrix
