Selection of optimal sample preparation technique is very important while analysing soil samples for PAHs residues.
PAHs are lipophilic compounds that show high affinity to organic matter and their detection in soil requires efficient extraction techniques to release strongly sorbed contaminants from the soil material. Soxhlet , LLE or SPE methods for soil sample analysis typically involve evaporation step that would result in low recovery of most volatile PAHs, like naphthalene.
Quick, Easy, Cheap, Effective , Rugged and Safe (QuEChERS) multiresidue sample preparation procedure adopted from Association of Analytical Communities (AOAC) was used in this work.
The proposed method QuEChERS uses single step buffered acetonitrile extraction while simultaneously salting water out with MgSo4 to induce liquid-liquid partitioning. After removing an aliquot from an organic layer, for further clean-up, a dispersive SPE is done using a combination of primary secondary amine (PSA) sorbent to remove organic acids from other components and anhydrous MgSo4 to reduce the remaining water in the extract. Other sorbents such as graphitized carbon black – to remove pigments and sterol or C18 to remove lipids and waxes can be added. The sample prep steps used in this work is described on fig. 1
Fig.1 The QuEChERS sample preparation steps using VetexQ kit
GC-MS analysis of PAH’s is rather demanding because of Similar chemical structures. There are some challenges one need to overcome, they are well described elsewhere. To name a few: Difficult chromatographic separations; difficult to differentiate on mass (same masses with little and identical fragmentation); High boiling compounds leading to high GC column elution temperatures, long analysis times and injector discrimination effects etc. The GC-MSD experimental workflow is shown in fig. 2, whereas method parameters are listed below.
GC method parameters:
Column: RTX-5MS (30m x 0.25, 250)
T° Oven= 80°C (0.0 min), 30°C/min – 250°C (0.0 min) 5.0°C/min – 310°C (4.0 min)
T° Inlet= 250°C Constant Flow mode, 1 ml/min flow ; Split mode=20:1 ;
T° Interface = 300 ° C
MSD data acquisition parameters:
- SCAN: for setting up SIM method Mass Range: 50 – 400 amu (Average = 2) Solvent Delay: 120 sec
- SIM – for quantitation, SIM: selection of monitored ions is automated/facilitated by «SIM Wizard®» algorithm Dwell Time: is set automatically by «iDwell®Time» algorithm
Fig.2 The GC-MSD experimental workflow.
First, one must run a mix of standards containing all your target compounds at 100ppb each in full SCAN mode in order to optimize chromatographic separation parameters. That is the starting point for the Q-Tek SIM Wizard® to lead you to optimal SIM method that would be then ideal for quantitation of all the targets.
Please, see more details in Technical note: Q-Tek SIM Wizard® productivity tool
TIC peak autointegration and simultaneous extraction of clean spectra for each compound is done initially. Autointegration algorithm confidently defines peaks and makes effective spectral library search even on co-eluting peaks.
As soon as clean spectra have been extracted from the target peaks the ion masses automatically selected by the algorithm as the best SIM-candidates are flagged on the compound spectrum. Results are shown in Table 1.
Table 1. Retention times and monitored ions for all PAH targets in the mix.
| № | Compounds | RT, min | Quant Ion | Qual Ion 1 | Qual Ion 2 |
| 1 | Naphthalene | 3.06 | 128 | 127 | 102 |
| 2 | 2-Methylnaphthalene | 3.56 | 141 | 115 | 142 |
| 3 | 1-Methylnaphthalene | 3.64 | 141 | 115 | 142 |
| 4 | Acenaphthylene | 4.27 | 152 | 151 | 126 |
| 5 | Acenaphthene | 4.42 | 154 | 153 | 152 |
| 6 | Fluorene | 4.82 | 165 | 166 | 139 |
| 7 | Phenanthrene | 5.61 | 178 | 176 | 152 |
| 8 | Anthracene | 5.64 | 178 | 176 | 152 |
| 9 | Fluoranthene | 6.72 | 202 | 101 | 200 |
| 10 | Pyrene | 6.99 | 202 | 101 | 200 |
| 11 | Benz[a]anthracene | 8.83 | 228.1 | 114 | 226.1 |
| 12 | Chrysene | 8.91 | 228.1 | 226.1 | 113 |
| 13 | Benzo[b]fluoranthene | 11.24 | 250.1 | 126.1 | 252.1 |
| 14 | Benzo[k]fluoranthene | 11.29 | 250.1 | 126.1 | 252.1 |
| 15 | Benzo[a]pyrene | 12.03 | 250.1 | 126.1 | 252.1 |
| 16 | Indeno[1,2,3-cd]pyrene | 15.00 | 276.1 | 138 | 274.1 |
| 17 | Dibenz[a,h]anthracene | 15.09 | 278.1 | 139.1 | 279.1 |
| 18 | Benzo[ghi]perylene | 15.68 | 276.1 | 138 | 274.1 |
Then, Q-Tek SIM Wizard® will split the TIC into time segments to ensure required number of points per a mass peak during the SIM acquisition, thus, finalizing the SIM method parameters. Each time-segment will contain the least number of targets, as to increase method sensitivity resulting from longer dwell-time for each monitored compound in the given time-segment (see fig. 3)
Fig3. Extracted ions chromatogram for mix of 18 PAH standards (100 ppb each): 1 – Naphthalene; 2 – 2-Methylnaphthalene; 3 – 1-Methylnaphthalene; 4 – Acenaphthylene; 5 – Acenaphthene; 6 – Fluorene; 7 – Phenanthrene; 8 – Anthracene; 9 – Fluoranthene; 10 – Pyrene; 11 – Benz[a]anthracene; 12 – Chrysene; 13 – Benzo[b]fluoranthene; 14 – Benzo[k]fluoranthene; 15 – Benzo[a]pyrene; 16 – Indeno[1,2,3-cd]pyrene; 17 – Dibenz[a,h]anthracene; 18 – Benzo[ghi]perylene.
The Q-Tek GC-MSD was then calibrated using 7 points: at 1-2-5-10-20-50-100 ppb level. The calibration curves in the range had good linearity (calibration coefficient values, R2>0.98-0.99) for all 18 compounds of the mix.
Further to evaluate the method for recovery and reproducibility (RSD) the blank soil sample was spiked with PAH standard mix at 2ppb each of PAH compound and run as unknown sample. The Extracted Ions Chromatogram and resulting table are shown below.
Fig. 4 Extracted Ion Chromatogram of PAH mix in Soil sample (PAHs spiked @ 2 ppb)
Table 2. Recovery and relative standard deviation (RSDs) values for target PAH compounds at 2ppb spike.
| № | Compound | Recoveries, %
(@ 2 ppb) |
RSD,
% |
||
| Sample 1 | Sample 2 | Sample 3 | |||
| 1 | Naphthalene | 111.5 | 112.5 | 121.5 | 4.2 |
| 2 | 2-Methylnaphthalene | 105.6 | 118.6 | 131.6 | 8.7 |
| 3 | 1-Methylnaphthalene | 97.8 | 116.3 | 116.3 | 8.2 |
| 4 | Acenaphthylene | 98.4 | 104.9 | 114.9 | 5.9 |
| 5 | Acenaphthene | 101.6 | 104.1 | 113.1 | 4.6 |
| 6 | Fluorene | 114.8 | 120.8 | 142.8 | 11.1 |
| 7 | Phenanthrene | 102.7 | 108.7 | 115.7 | 4.4 |
| 8 | Anthracene | 100.8 | 104.8 | 113.3 | 4.7 |
| 9 | Fluoranthene | 88.5 | 94.0 | 101.5 | 4.6 |
| 10 | Pyrene | 81.5 | 88.0 | 91.5 | 3.7 |
| 11 | Benz[a]anthracene | 70.3 | 74.8 | 84.3 | 5.2 |
| 12 | Chrysene | 67.1 | 72.6 | 78.6 | 3.9 |
| 13 | Benzo[b]fluoranthene | 113.7 | 107.6 | 101.4 | 4.1 |
| 14 | Benzo[k]fluoranthene | 112.4 | 104.0 | 99.2 | 4.8 |
| 15 | Benzo[a]pyrene | 107.1 | 106.4 | 96.8 | 4.4 |
| 16 | Indeno[1,2,3-cd]pyrene | 70.4 | 86.1 | 90.0 | 7.9 |
| 17 | Dibenz[a,h]anthracene | 109.3 | 124.2 | 124.2 | 6.6 |
| 18 | Benzo[ghi]perylene | 57.4 | 75.8 | 71.9 | 7.3 |
Conclusion: The results of the work demonstrate capability of Q-Tek GC-MS to confidently detect PAH compounds in soil at 2ppb level or below.
The proprietary Q-Tek SIM Wizard® productivity tool substantially simplifies the development of optimized SIM method for quantitation of PAH targets at a much shorter time and requires less operator skills.
