28 May / June 2014
Table 3. Compounds monitored by online SPE LC/MS, method parameters, and instrument limits of detection.
Compound
Acephate Allethrin Ametryn Atraton Atrazine
Atrazine Desethyl
Retention Time (min)
4.36
12.43 9.55 8.22 9.72 7.61
Atrazine-desisopropyl 6.52 Azoxystrobin Benzotriazole Bioresmethrin Bloc (Fenarimol) Carbaryl
Carbendazim DEET
Etofenprox Fenamiphos Fluoxastrobin Fluridone Flutolanil
Formasulfuron
Halosulfuron-methyl Imidacloprid Iprodione_a Iprodione_b Metalaxyl Metoprolol Oxadiazon Pramoxine Prometron Propanmide Quinclorac
10.38 6.6
13.24 10.3 9.32 6.01 9.79
13.55 11.25 10.95 10.31 10.75 9.41
11.23 6.89
11.26 11.26 9.81 7.34
12.44 9.65 9.11
10.81 8.33
Thiencarbazone-methyl 8.67 Thiophanate-methyl 8.88 Tramadol
Precursor Mass
184
303.2 228.1 212.2 216.1 188.1 174.1 404.1 120.1 339.2 331.2 202
192.1 192.1 394
304.1 459.1 330.1 324
453.1 435.1 256 330 332
280.2 268.2 345.1 294.2 226.1 256 242 391 343
7.25 264.2
stream. Since the matrix consists of a broad spectrum of physiochemical properties, some components of the matrix will not be retained during this stage and will go directly to waste, effectively cleaning the sample or extracting it from the bulk matrix.
The next stage is the elution step where the components retained after the initial loading step, are eluted from the column with a high elutropic mobile phase. At this point the valve is switched so that the eluent stream now goes to the detector rather than to waste, either via an analytical column or directly. The use of a second column allows for either greater separation of the analyte(s)
Product Mass 1
143 135 186 170 174 146 132 372 65
171 268 145 160 119 177 217 427 309 262 183 182 209 245 247 220 116 303 128 142 173 161 359 151 58
Product Mass 2
95
220 96
100 104 104 104 329 92
293 311 127 132 91
135 234 188 310 242 272 139 175 - -
160 191 220 100 170 209 224 230 311 246
LOD (ng/L)
0.24 7.8
0.12 0.12 0.12 0.12 0.24 0.12 7.8
62.5 0.24 0.12 0.12 0.98 3.9
0.12 0.5
0.12 0.06 0.12 0.12 0.06
15.63 31.25 0.06 0.24 3.9
0.12 0.12 0.12 7.8 3.9
0.24 0.06
and any remaining matrix components. The loading pump is used in this step to flush out the autosampler and associated tubing reducing the re-equilibration time and also reducing carryover.
The final phase is to re-equilibrate the system to be ready for the next sample injection. The valve is re-positioned to its original position, with the mobile phase composition being reset to its starting conditions.
On the actual system the valving looks a little more complicated due to the use of two injection ports to allow the user to swap between LC and SPE-LC, with the LC only injector valve also being used by the SPE-LC
configuration.
A 1 mL injection was loaded onto a Thermo Scientific™ Hypersil GOLD aQ™ column (20 x 2.1 mm, 12 µm particle size) and separated on a Thermo Scientific™ Accucore™ aQ analytical column (100 x 2.1 mm, 2.6 µm particle size) by gradient elution with methanol/water mobile phase. The MS data was acquired in selected reaction monitoring (SRM) mode on a Thermo Scientific™ TSQ Quantiva™ MS equipped with an H-ESI interface. Quantitative analysis was performed using Thermo Scientific™ TraceFinder™ software version 3.1.
Results and Discussion
HRMS Screening and Non-Targeted Identification
Representative HRMS chromatograms of SPE extracts subjected to non-targeted screening for the identification of organic pollutants and selection of target compounds for quantitative analysis are shown in Figure 3.
The non-targeted identification of fluridone in Pond 43 by EFS database screening and spectral library searching in ExactFinder software is demonstrated in Figure 4. Panel A shows an EFS database match for fluridone with a goodness of fit score of 0.93 between a modelled chromatographic peak and the observed peak. Panel B compares a modelled mass spectrum for the proposed pseudomolecular ion [C19
H14 NO]M+H F3 and
the averaged full-scan observed data with excellent mass accuracy (-0.31 ppm) at the mono-isotopic peak and a 100% isotope pattern score. In Panel C, library searching of the observed HRAM CID MS2 spectrum returns a match to the EFS library entry for fluridone with a score of 70%. Although in the examples shown there is a good match, it should be noted that in general EI and CI generated MS spectra tend to have more consistency, and thus databases generated using these sources are better.
The complete list of compounds identified by non-targeted screening and the samples in which they were found are listed in Table 2.
Targeted Quantification by Online SPE LC/MS
Based on the results of the non-targeted screening, knowledge of chemical usage on the island, and readily available reference standards, an online SPE LC/MS method was developed to quantify the occurrence
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