Automated SPE of water samples according to EN16691 for PAHs

Application note scope

The purpose of this application note is to outline an automated extraction method utilizing the Atlantic® DVB SPE Disk and the Biotage automated and manual SPE solutions for the extraction of PAHs in different water matrices. The first section will highlight the use of the Biotage® Horizon 5000 fully automated extraction system and the method used for this application. Additionally, there will be an Application Modification section that will highlight the use of the Biotage® VacMasterTM Disk for this application. 

Introduction

Polycyclic aromatic hydrocarbons (PAHs) are found worldwide and emitted from several sources including fossil fuel, coal and shale oil derivatives, coke production and burning
wood for home heating, generally arising from incomplete combustion.^[1] Food is considered an important source of human exposure, although smoking may provide a larger contribution for those who smoke.^[2] Characterization of PAHs and their concentration is of interest in maintaining public health.

PAH contamination is often airborne and can contaminate remote land and water resources. A number of PAH compounds have been determined to cause cancer by the U.S. Department of Health and Human Services (HHS) and include benz(a) anthracene, benzo(b)fluoranthene, benzo(a)pyrene, dibenz(a,h)anthracene, and indeno(1,2,3-c,d)pyrene.^[3] Others are considered probable carcinogenic agents to humans or probable agents, by US and International Agencies, giving cause for concern.

The European Union Water Directive restricts the amount of PAH in drinking water to less than 0.1 µg/L for benzo(b) fluoranthene, benzo(k)fluoranthene, benzo(ghi)perylene, and indeno(1,2,3-cd)pyrene. Benzo(a)pyrene has its own limit of 0.01 µg/L.^[4]

Therefore, the method to determine PAHs in water should be accurate, precise and sensitive to measure low concentrations in various volumes of water. Method EN 16691 is a recently developed method that uses solid phase extraction to isolate organic compounds from 1 L of water using a divinylbenzene (DVB) solid phase extraction disk. This application note will outline the method for the extraction of PAHs from water using an automated system with DVB disks.

Instrumentation

• Biotage Instruments:
  
  • Biotage® Horizon 5000 Automated Extraction System
  • DryVap™* Concentrator System
  • DryDisk® Separation Membranes
  • Atlantic® DVB SPE Disk
• Agilent
  
  • 6890 GC
  • 5973 Mass Selective Detector

*The DryVap™ system has been discontinued. We recommend using the TurboVap® evaporation systems for achieving equivalent results.

Method summary

1. Adjust 1 L aqueous sample to pH 2 with HCl, cap the bottle and mix.
2. Spike a known amount of the appropriate PAH standard solution into any LCS or MS samples.
3. Attach the sample bottle to the Water Inlet Valve using cap adapters as necessary.
4. Load the disk holder with the disk and place onto the extractor.
5. Place a clean collection vessel onto the extractor.
6. Load the sample bottle onto the Biotage® HorizonTM 5000.
7. Start the extraction method in table 1 and collect the sample extract.
8. Cap and label extract as PAH.
9. Dry and Concentrate the extract on the DryVap™ Concentrator System using the parameters from table 2.
10. Analyze sample extracts on GC/MS utilizing the conditions listed in table 3.

Table 1. Biotage® Horizon 5000 extraction method.

Table 2. DryVap™ conditions.

Table 3. GC/MS conditions.

References

1. http://www.ncbi.nlm.nih.gov/books/NBK217758/ (accessed June 1, 2016).
2. Scientific Opinion of the Panel on Contaminants in the Food Chain on a request from the European Commission on Polycyclic Aromatic Hydrocarbons in Food. The EFSA Journal (2008) 724, 1–114.
3. http://www.atsdr.cdc.gov/csem/csem.asp?csem=13&po=11, (accessed June 1, 2016).
4. Official Journal of the European Communities, http://eur-lex.europa.eu/legal-content/EN/TXT/PDF/?uri=CELEX:31998L0083&from=EN (accessed June 1, 2016).

Biotage® VacMaster™ Disk method summary

1. Repeat the following steps for each active VacMaster Disk station.

2. Setup the VacMaster Disk manifolds ensuring all waste lines and vacuum lines are attached. Set the vacuum pump to -24”Hg.

3. Prepare the disk holder assembly (47mm): ensure the support screen is flat in the center of the disk holder. Place the Atlantic® DVB Disk on top of the support screen with the ripples of the disk on top and add any prefilters on top of the disk. Place the disk holder assembly on the VacMaster Disk ensuring there is a tight seal with the luer fitting.

4. If using the multifunnel, place onto the disk holder assembly. If not using the multifunnel, omit those directions throughout the method.

5. Condition the SPE Disk:

   1. Guide for each conditioning step in table 4 below:

      1. Measure appropriate volume of solvent into a gradu- ated cylinder and pour into the disk holder assembly.

      2. Using a Nalgene Wash Bottle (phthalate free), rinse the multifunnel and disk holder in a circle for about 3 seconds using the same SOLVENT (approximately 5 additional mL).

      3. SATURATE the disk for the time shown (in seconds). (Saturate means: quickly turn the knob to the proper waste destination and back to the “OFF” position. This brings the solvent into the disk media bed).

      4. SOAK the disk for the the time shown (in seconds).

      5. DRAIN to proper waste destination for the time shown (in SECONDS). Switch to the “OFF” position.

Table 4. Disk conditioning.

6. Load the sample:

a. For multifunnel: quickly and efficiently angle the bottle to rest on the multifunnel upside-down.

b. For no multifunnel: pour a portion of the sample into the disk holder.

c. Adjust the vacuum between -10”Hg and -15”Hg for sample load (please note, if the sample is flowing too slowly, the vacuum can be adjusted to a lower setting). Drain the sample to “AQUEOUS” waste. Continue to pour the sample into the disk holder ensuring the disk does not go dry or overflow for the duration of sample load.

7. Wash sample container:

a. Guide for each wash step in table 5 below:

i. Measure the appropriae VOLUME of REAGENT into agraduated cylinder, pour into the sample bottle, and swirl around. Pour the solvent in the sample bottle into the disk holder assembly.

ii. Using a Nalgene Wash Bottle (phthalate free), rinse the multifunnel and disk holder in a circle for about 3 seconds using the same REAGENT (approximately 5 additional mL).

iii. SATURATE the disk for the time shown (in seconds).

iv. SOAK the disk for the the time shown (in seconds).

v. DRAIN to “AQUEOUS” waste for the time shown (in SECONDS). Switch to the “OFF” position.

Table 5. Sample container wash.

8. Air dry the SPE Disk:

a. Return the vacuum to -24”Hg and continue to air dry the SPE disk to “AQUEOUS” waste for an additional 240 SECONDS. Switch to the “OFF” position.

b. Remove the sample bottle from the multifunnel if it was used.

9. Elute the SPE Disk: (Please note: the elutions will go into the collection flask inside the chamber, not to waste containers)

a. Place a clean 125 mL 24/40 tapered Erlenmeyer flask into the VacMaster Disk collection chamber. Place the cover on the chamber. Remove the disk holder assembly and place the disk holder assembly into the luer fitting on top of the collection chamber. Attach the luer fitting of the collection chamber assembly onto the manifold.

b. Guide for each elution step in table 6 below:

i. Measure the appropriate volume of solvent into a graduated cylinder, pour into the sample bottle, and swirl around. Pour the solvent in the sample bottle into the disk holder assembly.

ii. Using a Nalgene Wash Bottle (phthalate free), rinse the multifunnel and disk holder in a circle for about 3 seconds using the same SOLVENT (approximately 5 additional mL).

iii. SATURATE the disk for the time shown (in seconds) to “ORGANIC”.

iv. SOAK the disk for the the time shown (in seconds).

v. DRAIN to “ORGANIC” for the time shown (in SECONDS). Switch to the “OFF” position.

vi. Remove the chamber lid to release the vacuum from inside the chamber.

Table 6. Disk elution.

Literature number: AN110