The subject of PFAS is more urgent than ever. The latest European Drinking Water Directive with amendments regarding PFAS was implemented on January 12, 2021. Also in the United States, the corresponding US EPA 537.1 regulation for PFAS in drinking water was recently updated. day.
Today, the analytical procedures for the determination of PFAS are not yet as harmonized and regulated as the procedures for other substances, such as PCDD/Fs or PCBs. Not only the final determination of PFAS with LC/MS-MS methods is very demanding, but also the preparation of the samples, regardless of regional regulations. The samples are purified and enriched by solid phase extraction, eluted in a significantly lower volume in methanol and then concentrated to dryness without loss before being redissolved.
For standardized and automated sample preparation in PFAS analysis, LCTech’s Freestyle SPE system, in combination with the D-EVA vacuum concentrator, enables specially tailored automated concentration of substances.
In order to be able to achieve the determination limits required for the new MRL values, a sufficiently large sample volume must be applied. These large sample volumes lead to a very high workload and time for manual SPE, which is why automated sample processing, in addition to standardization, is also advantageous. Another major challenge is posed by the ubiquity of perfluorinated and polyfluorinated substances in the laboratory. Contamination of PFAS analytes in samples should be kept as low as possible to avoid false positive results. A well-known undesirable source of PFAS in samples is fluorocarbon materials, which are commonly used in laboratory equipment and materials, such as PTFE. These release small amounts of PFAS upon contact with the sample, resulting in a significant increase in background levels. Therefore, in the Freestyle SPE-PFAS system for automated processing of the SPE protocol, all fluorocarbon materials have been systematically minimized and replaced with polyethylene or polypropylene.
Environmental samples, such as soil and water, and biota samples, such as animal tissue or food, can be purified automatically with the Freestyle SPE-PFAS system. If sample volumes greater than 100ml are inserted, the system is upgraded with the XANA water module. This allows up to 24 1 L samples to be pumped from the rack through the SPE cartridge and dramatically increases sample throughput by processing up to 7 samples simultaneously.
Figure 1 shows the Freestyle-PFAS-XANA system with a capacity of up to 24 x 1 L bottles (in the figure, glass bottles) with D-EVA in use. To implement US EPA 537.1, samples were placed in 250 mL PE bottles and the eluate was collected in 15 mL or 50 mL FalcontubesTM. Without further sample transfer of the eluate, the samples in these containers are then inserted into the appropriate D-EVA rotor for gentle evaporation to dryness.
For evaluation, PFAS compounds from existing DIN 38407-42 and analytes from updated US EPA 537.1 were processed on a Freestyle PFAS XANA system and evidence of all formal validation requirements for the US EPA 537.1 have been provided. The results prove that all PFAS compounds are well recovered, no traces adhere to the system and at the same time no blank values are generated during processing on the robot. For this experiment, background PFAS values of neat solvents were compared to sample blank values and a selection shown in Table 1.
After SPE purification, the eluate of 8 mL of methanol is evaporated to dryness and almost dry and redissolved for analysis in methanol/water 96:4 (v/v) according to US EPA 537.1.
Even this seemingly trouble-free step poses challenges for PFAS testing labs. The adhesion of PFAS to glass walls makes most equipment typically used for concentration unsuitable. Therefore, the only remaining option would be the time-consuming and expensive supervised blowing with nitrogen, which should be avoided due to the unintended formation of aerosols.
LCTech provides an automated solution in a D-EVA device, which holds the SPE eluates in common FalcontubesTM and runs a controlled process. By design, D-EVA avoids losses, because it uses infrared light for heating, lowers the boiling temperature with a particularly low pressure gradient and prevents the formation of aerosols by its rotation. In addition, neutral PFAS having a particular tendency to be lost during concentration beyond the dry point, it is necessary to ensure that the protocol ends in a controlled manner over time. For this purpose, the D-EVA has a special sensor in a reference container, which reliably detects the reached dryness and stops the process. To this end, internal protocols have been developed with optimal rotation speed and associated termination temperature. When the target point is reached, the centrifugation chamber is vented and the infrared light used for temperature control is turned off so that no further direct heating power is applied to the sample containers. The “memory effect”, known from heating blocks and bain-maries, is thus avoided. In this way, even neutral PFAS can be concentrated very quickly and measured with relatively high recoveries compared to expensive and time-consuming nitrogen blowing.
The graph in Figure 2 shows the recovery rates of various PFASs related to US EPA 537.1. Where 8 mL of methanol was spiked and evaporated to the mid calibration level. The blowing method under N2 in a water bath is compared to the “methanol” evaporation method in D-EVA. The evaporated tubes are redissolved in 95/4 methanol/water plus internal standard and measured with HPLC-MS/MS.
By precisely applying US EPA 537.1, some residual water content in the eluate is sometimes impossible to avoid, which presents an additional challenge for concentration. However, even samples containing up to 25% water can be gently dried using D-EVA.
In the experiment, 8 samples were spiked with a surrogate standard solution, with a concentration in the middle calibration range, then mixed with different amounts of water and made up to 8 ml with methanol; all samples were evaporated with the D-EVA “Methanol” program. Any nearly dry sample is considered finished. When all samples are nearly dry, they are dissolved in 96:4% (v/v) methanol/water, spiked with the internal standard and analyzed by LC-MS/MS.
Due to the high sample throughput of the Freestyle-XANA, another primary goal of the D-EVA application is to evaporate faster than the methodology recommended by US-EPA 537.1, without the risk of losing analytes, while achieving the desired signal strength without any staff supervision. . In the respective rotors, 23 samples in 15 mL or 10 samples in 50 mL FalcontubesTM can be used in parallel. The fast program evaporates eluates to dryness in 11 minutes and the longer program, specially selected for water-contaminated samples, in 77 minutes. Thus, the two devices can be ideally combined in the laboratory and a workflow for a high sample throughput can be realized.
The analytical objectives of the applied method were achieved with good recovery rates and very low standard deviations without cross-contamination, thanks to reliable and robust automation. Likewise, the analytical goals of the DIN method were also achieved with high selectivity, trueness and precision. The use of the D-EVA automated vacuum centrifuge, which allows unattended parallel concentration of the collected PFAS eluate, enables gentle concentration, which achieves particularly high robustness.