Development and application of a headspace solid-phase microextraction and gas chromatography/mass spectrometry method for the determination of dimethylsulfide emitted by eight marine phytoplankton species

Headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry (GC-MS) has been developed, optimized, and applied to investigate the dimethylsulfide (DMS) emissions from eight marine phytoplankton species, namely, Calcidiscus leptoporus, Emiliania huxleyi, Phaeodactylum tricornutum, Chaetoceros neogracilis, Dunaliella tertiolecta, Synechococcus, Prochlorococcus, and Trichodesmium. Four SPME fiber coatings (PDMS, PDMS-DVB, PDMS-CAR, and CW-DVB) were tested for linearity and limit of detection. Key parameters such as equilibrium and extraction times, desorption temperature and time, and headspace volume were optimized to make extraction as efficient as possible using the PDMS-DVB fiber coating. This fiber enabled the characterization of DMS in seawater under 0.005 nM levels and within 1 min exposure time. Among the different algae groups, the two coccolithophorids, C. leptoporus and E. huxleyi, were the strongest emitters of DMS. Within the cyanobacteria, Prochlorococcus and Trichodesmium expressed no DMS emission, whereas Synechococcus showed very low DMS emission. The DMS emission of C. leptoporus, however, was several orders of magnitude higher than all other algae, including E. huxleyi, which is known to be a prolific emitter of DMS. *Corresponding author: [email protected] Acknowledgments We would like to thank Ian Probert for subletting us his isolated Calcidiscus leptoporus culture. This work was completed as part of the OOMPH project (Specific Targeted Research Project [STREP] in the Global Change and Ecosystems Sub-Priority). SUSTDEV-2004-3.I.2.1. Project Number 018419. Limnol. Oceanogr.: Methods 4, 2006, 374-381 © 2006, by the American Society of Limnology and Oceanography, Inc. LIMNOLOGY and OCEANOGRAPHY: METHODS Yassaa et al. DMS analysis using SPME-GC-MS 375 erable potential for marine research. Invented in 1989 by Pawliszyn and coworkers (Belardi and Pawliszyn 1989; Arthur and Pawliszyn 1990), SPME is an innovative, solvent-free technology that is fast, economical, sensitive, and versatile. The fiber coating removes the compounds from the sample by absorption in the case of liquid coatings or adsorption in the case of solid coatings. After the sample has been acquired, the SPME fiber is then inserted directly into the GC for desorption and analysis. Two basic types of extractions can be performed using SPME: direct and headspace extraction (Lord and Pawliszyn 2000). In the direct extraction mode, the coated fiber is inserted into the sample medium and the analytes are transported directly to the extraction phase. To facilitate rapid extraction, some level of agitation is required to enhance transport of the analytes from the bulk of the solution to the vicinity of the fiber. For gaseous samples, natural convection and diffusion in the medium is sufficient to facilitate rapid equilibration. For aqueous matrices, more efficient agitation techniques, such as fast sample flow, rapid fiber or vial movement, stirring, or sonication are required. These actions are undertaken to reduce the effect caused by the depletion zone, which occurs close to the fiber as a result of fluid shielding and slow diffusion of analytes in liquid media. In the headspace mode, the analytes need to be transported through a layer of air before they can reach the coating. This approach also has the advantage of protecting the fiber coating from damage by high molecular-weight species and other nonvolatile contaminants present in the liquid sample matrix, such as humic materials or proteins. This headspace mode also allows modification of the matrix, such as a change of the pH, without damaging the fiber. Concentrations derived from the same vial at equilibrium using direct and headspace sampling are identical, as long as sample and gaseous headspace volumes are the same and that calibration is performed in the same mode. This is a result of the fact that the equilibrium concentration in each phase is independent of fiber location in the sample/headspace system. The choice of sampling mode has a significant impact on extraction kinetics. In fact, the equilibration times for volatile components are shorter in the headspace SPME mode than for direct extraction under similar agitation conditions. When the fiber coating is in the headspace, the analytes are removed from the headspace first, followed by indirect extraction from the matrix. Therefore, volatile analytes are extracted faster than semi-volatile components since they are at a higher concentration in the headspace, which contributes to faster mass transport rates through the headspace. In this work, headspace-SPME has been optimized and applied to the determination of DMS emission from eight phytoplankton species, namely, Calcidiscus leptoporus (strain AC365), Emiliania huxleyi CCMP 371, Phaeodactylum tricornutum, Chaetoceros neogracilis CCMP1318, Dunaliella tertiolecta, Synechococcus RCC 40, Prochlorococcus RCC 158, and Trichodesmium IMS 101. Four SPME fiber coatings, 100 μm polydimethylsiloxane (PDMS), 65 μm polydimethylsiloxanedivinylbenzene (PDMS-DVB), 65 μm carbowax-divinylbenzene (CW-DVB), and 75 μm PDMS-Carboxen (PDMS-CAR), were tested by comparing the linearity and limit of detection. Key parameters such as equilibrium and extraction times, desorption temperature and time, and headspace volumes were all optimized in this work. Materials and procedures Chemicals and standard solutions—DMS (≥99% purity) was purchased from Sigma-Aldrich (Taufkirchen, Germany). A DMS solution of 50 μM was prepared in chromatography-grade deionized water (Merck), and dilutions were made immediately afterward. Aliquots of 20 mL of each DMS solution (ranging from 0.005 nM to 50 μM) were dispensed and sealed in 40 mL vial with PTFE/Silicone septum (Supelco) and immediately subjected to SPME extraction. Calibration of SPME fiber with gaseous DMS—To determine how much DMS is uptaken by an SPME fiber, calibrations need to be performed with representatively small amounts of DMS. A practical way to achieve both this and facilitate multiple dilutions was to make the calibration in gas phase. Therefore, a gravimetrically prepared standard gas bottle (ApelRiemer Environmental) containing the compounds listed in Table 1 has been used for the calibration of SPME fiber. Three mass flow controllers were used to dilute the calibration gas with purified synthetic air in the range of 0.1 to 510 ppbv. The Table 1. Compounds and their mixing ratios containing in the calibration gas bottle Compounds Mixing ratios (ppb)