3
phase extraction step performed through a C 18 cartridge. The recovered methanol
solution containing both OLP and d 3 -OLP is then injected directly in the APCI
source and analyzed in SRM, monitoring the appropriate fragmentation. The key
points of this approach are:
(i). the use of a suitable labeled internal standard:
Regarding this point, it is important (even in the perspective of a possible wide
application of the method) to easily obtain the labeled molecule by simple synthetic
procedures. The method describes the synthesis of d 3 -OLP starting from demethyoleuropein (previously recovered and purified by a polar extract of olives) and CD 2 N 2
(Fig. 1.1).
It is important to achieve the highest grade of isotopic purity of the labelled internal standard in order to avoid inaccurate quantification of the analyte. Figure 1.2
shows the comparison between the high resolution signals from protonated OLP
and d 3 -OLP. It can be seen that the isotopic purity of the internal standard is
above 97%.
(ii). The selection of the appropriate fragmentation reaction:
The fragmentation pattern of oleuropein is rather straightforward (Fig. 1.3). The
molecule is composed of three moieties: the glucose ring, whose loss generate the
ions at m/z 361, 379 (364, 382 for d 3 -OLP), the hydroxytyrosol portion which generates the ion at m/z 137, and the elenolic acid which is the core of the molecule. The
most responsive ion in both analyte and internal standard MS/MS spectra is the
tyrosyl ion (m/z 137) which is particularly stable due to its aromatic character. The
fragmentation energy may be optimized to maximize the signal intensity.
(iii). the determination of analytical parameters.
In this type of approach the most important analytical parameters to test are of
course the accuracy, the limit of quantitation (LOQ) and the limit of detection
(LOD). These aspects may be verified by using an artificially fortified matrix which
Fig. 1.1 Synthesis of
d3-Oleuropein
1 Evaluation of Quality and Safety of Foods by Tandem Mass Spectrometry
phase extraction step performed through a C 18 cartridge. The recovered methanol
solution containing both OLP and d 3 -OLP is then injected directly in the APCI
source and analyzed in SRM, monitoring the appropriate fragmentation. The key
points of this approach are:
(i). the use of a suitable labeled internal standard:
Regarding this point, it is important (even in the perspective of a possible wide
application of the method) to easily obtain the labeled molecule by simple synthetic
procedures. The method describes the synthesis of d 3 -OLP starting from demethyoleuropein (previously recovered and purified by a polar extract of olives) and CD 2 N 2
(Fig. 1.1).
It is important to achieve the highest grade of isotopic purity of the labelled internal standard in order to avoid inaccurate quantification of the analyte. Figure 1.2
shows the comparison between the high resolution signals from protonated OLP
and d 3 -OLP. It can be seen that the isotopic purity of the internal standard is
above 97%.
(ii). The selection of the appropriate fragmentation reaction:
The fragmentation pattern of oleuropein is rather straightforward (Fig. 1.3). The
molecule is composed of three moieties: the glucose ring, whose loss generate the
ions at m/z 361, 379 (364, 382 for d 3 -OLP), the hydroxytyrosol portion which generates the ion at m/z 137, and the elenolic acid which is the core of the molecule. The
most responsive ion in both analyte and internal standard MS/MS spectra is the
tyrosyl ion (m/z 137) which is particularly stable due to its aromatic character. The
fragmentation energy may be optimized to maximize the signal intensity.
(iii). the determination of analytical parameters.
In this type of approach the most important analytical parameters to test are of
course the accuracy, the limit of quantitation (LOQ) and the limit of detection
(LOD). These aspects may be verified by using an artificially fortified matrix which
Fig. 1.1 Synthesis of
d3-Oleuropein
1 Evaluation of Quality and Safety of Foods by Tandem Mass Spectrometry