- Simultaneous full scan and time programmed SIM. The central feature of our method is the use of full scan together (simultaneously) with time programmed SIM for selected main targeted pesticides. SIM provides sufficient instrument sensitivity (better than MS-MS), and it can be time programmed the same as parent ions in MS-MS, while the full scan enables universal analysis hence should ensure against false negatives. Full scan can further serve for both library identification and for the provision of few more ions for additional confirmation in case of need (if sufficient instrument sensitivity is provided). More importantly, full scan can serve for reconstructed (RSIM) search of unlimited number of pesticides via their molecular ions and one additional isotopomer (same molecule, different elemental isotopes, such as M+1 etc.) and/or high mass fragment ion per pesticide. Accordingly, truly universal analysis is achieved. We note that suitable software for such multi RSIMs data analysis is provided by the Agilent Chemstation software and an improved version of it is the Agilent DRS software. On the other hand, the time programmed SIM on the molecular ions and their isotopomers (and/or one high mass fragment in case of need) provides the needed instrument sensitivity for trace level pesticide analysis (<10 ppb) while the method selectivity is largely increased on the molecular ions as discussed in details in our previous blog note and as briefly described below.
- Improved Selectivity against Matrix Interference. We found that matrix interference is exponentially reduced with mass at a rate of about twenty every 100 amu (J. Chromatog. A. 974, 185-212 (2002)). Thus, SIM on the molecular ion is about as selective against matrix interference as MS-MS on a fragment ion. Since most pesticides are analyzed by MS-MS on their major fragment ions as parent MS-MS ions in standard EI, we claim that the 5975-SMB GC-MS with Cold EI is as selective as GC-MS-MS with standard EI. In addition, the instrumental SIM sensitivity is superior to that of MS-MS since the later suffers from four types of signal losses compared to a single quadrupole MS in SIM mode: 1) Q3 losses of a factor of 3-4 with standard tune resolution; 2) CID losses due to dissociation to a few ion masses which is about x3 loss factor but sometimes much higher; 3) Q2 losses due to ineffective retaining of fast fragment ions; 4) Q2 collimation losses at the Q2 entrance and exit lenses which do not exist in a single quadruple instrument (about x2-3). In total, MS-MS suffers from about an order of magnitude lower daughter ions signal than SIM signal on the parent ions. Obviously MS-MS is sometimes operated on parent ions that are molecular ions and in that case it is more selective than SIM. However, these cases are usually with easy to analyze pesticides while the harder to analyze pesticides that represent the bottleneck of the analysis method are typically with weak or no molecular ion in their standard EI MS.
- Extended range of pesticides amenable for analysis. The use of either PTV injector, or standard injector at a lower injector temperature with pulsed high flow rate splitless injection and the combination of short column, high column flow rate and the fly-through ion source of Cold EI enable the analysis of an extended range of pesticides including many of those that are considered difficult to analyze and those that currently require LC-MS for their analysis.
- No peak tailing. The full elimination of ion source peak tailing in the Cold EI fly-through ion source improves the signal of pesticides that are more difficult to analyze. Furthermore, the use of flow programming with very high flow rate at the end of the run serves to effectively clean the column from matrix residue hence to suppress column related peak tailing.
- Enhanced molecular ions and selectivity. The enhancement of the molecular ions in Cold EI in combination with the elimination of ion source peak tailing results in lower pesticides LOD, particularly for those pesticides that are difficult to analyze or that require LC-MS.
- Isotope ratio based identification. The ions that are selected for SIM for a given pesticide include the molecular ion and an additional one major isotopomer ion (such as M+1, M+2 etc.) and only if considered essential one additional high mass fragment. The use of our Tal-Aviv Molecule Identifier (TAMI) software provides a machine generated isotopomers ratio matching factors which in combination with having peaks with the same elution times results in effective sample identification.
- Fast screening analysis. The use of a short column and high column flow rate enables column temperature programming at 25ºC/min right from the start hence chromatography time of 10 minutes which is three times faster than commonly employed. If needed, even 50ºC/min can be used and at the end of the run the column flow rate is either reversed (back-flush) or increased to 30 ml/min for effective column cleaning from everything that was introduced into the column by the injector.
- Complementary method to LC-MS-MS. Currently, pesticides are analyzed by both GC-MS and LC-MS-MS. Since LC-MS is operated with a target based method of LC-MS-MS it is only logical that the GC-MS method will be universal with an instrument that can analyze extended range of pesticides and thus can also serve for the confirmation of LC-MS findings.
We evaluated the relative sensitivity and limits of detection of the 5975-SMB GC-MS with Cold EI. We found that for easy to analyze pesticides such as diazinon the TIC peak signal to noise ratio is about the same in both systems. However, in RSIM on the molecular ion which is the most selective ion in the MS Cold EI benefits from enhanced molecular ion which for diazinon is a factor of 4.2. As the pesticide becomes harder to analyze the Cold EI sensitivity gain is increased due to its uniform response. An example can be seen in Figure 3 below, in which we compare the analysis of Prochloraz with the 5975-SMB and a Shimadzu QP 2010 Plus system. For a "fare" comparison we did not use the molecular ion m/z=375.1 as it was fully missing in Standard EI (unlike in Cold EI). However, as shown, the Prochloraz RSIM trace on its high mass fragment m/z=308 badly tails in standard EI and as a results it lost about a factor of 7.5 in its TIC signal compared with Cold EI. The software calculated S/N (after file conversion of the Shimadzu data file into Chemstation file (using the MASSTransit file conversion software) showed that Cold EI was superior to Standard EI by a factor of over 20 as the noise level in Cold EI was very low.
Finally, in Figure 5 we further demonstrate the effectiveness of our method in the analysis of an incurred difficult to analyze pesticide such as Iprodion in Strawberry at ~70 ng/g concentration with the 5975-SMB GC-MS with Cold EI, in simultaneous full scan and two ions SIM mode. This Iprodion residue was undetected by standard GC-MS. Note the very high S/N in the SIM trace of 11 pesticides (22 ions in the group). In the SIM trace on either the isolated molecular ion m/z-329 or high mass fragment m/z=314 the RSIM trace was clean. Furthermore, while Iprodion was not identified by the NIST library due to matrix interference, IAA with the TAMI software provided a matching factor of 961 (in the full scan) thus Iprodion was positively identified.