INTRODUCTION
The A. E. Lalonde AMS Laboratory (AEL-AMS) has been in operation since 2014, providing routine radiocarbon (14C) analysis of a wide range of organic and inorganic materials (Crann et al. Reference Crann, Murseli, St-Jean, Zhao, Clark and Kieser2017). The radiocarbon content of dissolved inorganic carbon (DIC) and dissolved organic carbon (DOC) in freshwater systems, as well as CO2 and CH4 in gas mixtures, provides insights on carbon cycling dynamics and source apportionment in environmental systems (Lachinet et al. Reference Lachinet, Lawson and Sloat2012; Mann et al. Reference Mann, Eglinton, McIntyre, Zimov, Davydova, Vonk, Holmes and Spencer2015; Grinter et al. Reference Grinter, Lacelle, Baranova, Murseli and Clark2018; Wang et al. Reference Wang, Lafrenière, Lamoureux, Simpson, Gélinas and Simpson2018). To meet increasing demand, a method development program has been undertaken to improve sample preparation techniques for greater precision and to accelerate processing of water (DIC, DOC) and gas (CO2, CH4) samples at AEL-AMS, particularly for dissolved organic carbon (DOC) in water. Wet chemical oxidation has commonly been employed for stable isotope analysis of DO13C in waters (St-Jean Reference St-Jean2003; Osburn and St-Jean Reference Osburn and St-Jean2007; Lang et al. Reference Lang, Bernasconi and Früh-Green2012; Zhou et al. Reference Zhou, Guo, Lu, Mao, Zhen and Wang2015, etc.), and more recently has been applied to 14C (Leonard et al. Reference Leonard, Castle, Burr, Lange and Thomas2013, Lang et al. Reference Lang, McIntyre, Bernasconi, Früh-Green, Voss, Eglinton and Wacker2016). Other methods such as UV oxidation (Beaupre et al. Reference Beaupre, Druffel and Griffin2007; Druffel et al. Reference Druffel, Griffin, Walker, Coppola and Glynn2013), evaporation and lyophilization (Burr et al. Reference Burr, Thomas, Reins, Jeffrey, Courtney, Jull and Lange2001; Neff et al. Reference Neff, Finlay, Zimov, Davydov, Carrasco and Schuur2006) and solid phase extraction techniques (Sickman et al. Reference Sickman, DiGiorgio, Davisson, Lucero and Bergamaschi2010; Hendry and Wassenaar Reference Hendry and Wassenaar2005) are limited by complicated analytical routines and long processing times, where high temperature catalytic oxidation (HTCO) methods generally report high analytical blanks (Hedges et al. Reference Hedges, Bergamaschi and Benner1993; Benner and Strom Reference Benner and Strom1993). Focused efforts have been on developing a robust DOC extraction method by wet oxidation for freshwater samples designed specifically for high throughput. Extraction protocols for gas samples (CO2 and CH4) at AEL-AMS are also discussed.
METHODS
Samples are assigned a lab identifier number (UOC-#) and processed according to media codes based on the analysis of interest (Table 1). Appropriate reference materials are processed with each batch of unknowns, also summarized in Table 1. Concentration information (ppmC) is requested at the time of submission to determine the appropriate volume of material required to yield ∼1.0 mgC for 14C analysis.
Table 1 Standard reference material for AEL-AMS material codes.
* Dissolved in 20 mL of ultra pure Milli-Q water.
1 Independently verified by direct combustion (organics) or hydrolysis (inorganics).
2 Consensus value.
DIC and DOC
Definitions of what constitutes DIC and DOC can vary depending on the user and field of study; typical designations of what represents carbon species in aqueous systems can be found in St-Jean (Reference St-Jean2003). At the AEL-AMS lab, water samples are requested to be received pre-filtered with baked (500ºC for 3 hr) 0.7-µm GF/F filters or pre-rinsed 0.45-µm nylon or polypropylene filters, depending on the application; these filters were tested to have slightly lower blank contributions (<1 pMC) compared with nitrocellulose (>1.5 pMC). Round-bottom borosilicate Wheaton® bottles (Cat. No. 219439) are used as the reaction bottles for both DIC (by acidification) and DOC (by wet chemical oxidation) extractions. All bottles are pre-cleaned (10% HCl, MilliQ rinse), pre-baked (500ºC for 3 hr) before each use, and are reusable. Bottles are capped with a pre-cleaned (10% HCl, sonicate in MilliQ, dried at 50ºC overnight) butyl septum closure (33-430, Wheaton Cat. No. 240680). The appropriate bottle size (125 mL, 250 mL, 500 mL or 1000 mL) is selected based on the volume of water required (depending on concentration) to yield ∼1.0 mgC, and to leave a minimum of 1/3 headspace volume so as to not overpressure the reaction bottle during the extraction.
For DI14C analysis, the required volume of water is transferred via septum cap using a double-ended needle or by syringe into a pre-evacuated (<10–2 mbar) reaction bottle with 3–5 mL of 85% o-phosphoric acid to acidify the sample to pH <2. The sample bottle is then heated on a hot plate to 60°C for 1 hr to convert all dissolved inorganic species to CO2 into the headspace. The bottles are allowed to cool to room temperature prior to CO2 extraction.
For DO14C analysis, the water sample is transferred into an appropriate sized reaction bottle leaving at least 1/3 headspace to avoid overpressure. The sample is acidified to pH <2 with 85% o-phosphoric acid via open cap and sparged with He for 15 min. The reaction bottles are then capped and heated to 60°C for 1 hr. Following heating, the headspace is sparged a second time with He through the septa to remove any residual DIC fraction, and the sample is cooled to 4ºC. The DOC fraction is then oxidized to CO2 following a wet chemical oxidation (WCO) technique modelled after St-Jean (Reference St-Jean2003); Lang et al. (Reference Lang, Bernasconi and Früh-Green2012, Reference Lang, McIntyre, Bernasconi, Früh-Green, Voss, Eglinton and Wacker2016) and Zhou et al. (Reference Zhou, Guo, Lu, Mao, Zhen and Wang2015). A stock of oxidant solution is prepared by dissolving 400g Na2S2O8 (Sigma Aldrich >99% purity, Cat. No. 71890) in 1L boiling MilliQ water for 5 min to clean the oxidant and reduce the blank. A fixed volume of oxidant solution (8 mL) is added to the cold DIC sparged sample via the septa along with 1 mL of 0.5N AgNO3 (Fisher Scientific, Cat. No. S181) which acts as a catalyst to improve CO2 yield (Zhou et al. Reference Zhou, Guo, Lu, Mao, Zhen and Wang2015). The volume of oxidant remains constant as all samples (of varying ppmC concentrations) are portioned volumetrically to ultimately target a fixed amount of carbon (1 mgC). The amount of oxidant added is a minimum of 10× excess for what is needed to convert 1 mg of organic carbon to CO2, to account for (1) loss of oxidation power during heating of the oxidant solution, (2) recalcitrant organic carbon that is difficult to oxidize, and (3) dissolved ions in solution competing for the Na2S2O8. The bottle is heated to 95°C for 60 min on a hotplate with occasional gentle manual swirling, then allowed to cool to room temperature prior to CO2 extraction.
Once the sample is reacted, CO2 is extracted from the headspace on a low-flow He carrier extraction line (discussed below).
CX (CO2 Extraction and Purification)
CO2 is extracted from the headspace of the reaction bottle (for DIC/DOC) or a thick butyl rubber septum-fitted Wheaton® bottle (for CX) on a multipurpose extraction line. The line consists of stainless steel and PEEK tubing for the Helium (He) carrier flow-through section (Figure 1A) interfaced to a glass vacuum line for subsequent purification and trapping of CO2 in a breakseal (Figure 1B).
Figure 1 Schematic diagram of multipurpose line for extraction of waters (DIC/DOC) and CO2 (CX) samples, showing (A) flow-through section and (B) vacuum section. Numbers are described in the text.
The flow-through section (A) sweeps CO2 from the headspace of the reaction bottle using a He carrier gas, regulated at 200 mL/min (1). This section of the line consists of two 20-gauge stainless steel needles which are inserted through the septum of the reaction bottle: a 12-inch-long sparging needle (2) that bubbles He through the water sample, and a 6-inch exit needle (3) into the headspace which completes the flow conduit through the reaction bottle. The carrier section is equipped with a Nafion™ (Permapure MD-070-96P-2) gas dryer (4), designed to remove traces of water from the carrier (He) and extracted CO2, which are impermeable to the membrane and continue through to a stainless steel U-trap (5) packed with Ag wool (to increase surface area). This trap is immersed in liquid nitrogen (LN2), where CO2 freezes and He vents to atmosphere. The reaction bottle headspace is flushed by 10x its volume to ensure all CO2 is removed.
The flow section is interfaced to a vacuum line (B) via a three-way valve (6) with a port to vent and the other to vacuum. Once the extraction is complete, the stainless-steel U-trap (5) is isolated on both sides, and the flow of He is immediately shut off (NB: this is a critical step to prevent over pressuring the carrier section and damaging the Nafion™ membrane). Helium and other non-condensable gases are evacuated from the U-trap volume while the CO2 remains frozen at LN2 temperature. The vacuum is then isolated from the manifold, a –80°C ethanol bath is placed on the stainless-steel U-trap (5) to release CO2 and hold any residual water, and the extracted CO2 is transfered through to the vacuum section for cryogenic purification on a glass U-trap (7). The purified CO2 is then quantified in a calibrated volume (8) using a Baratron gauge. The measured CO2 is then transferred to a pre-baked (500°C for 3 hr) 6-mm OD borosilicate breakseal (9) containing a small amount (5–6 grains) of Silvered Cobaltous/Cobaltic oxide (Elemental Microanalysis, Cat. No. B1120) which has been previously baked at 500°C for 2 hr. The breakseal is sealed with the CO2 sample and baked at 200°C overnight to remove any S-bearing species that may poison the Fe catalyst during graphitization (Palstra and Meijer Reference Palstra and Meijer2014).
CH4
Methane samples are processed on a high temperature oxidation line following the methods of Pack et al. (Reference Pack, Xu, Lupascu, Kessler and Czimczik2015), which is interfaced to the multipurpose vacuum extraction line (not shown in Figure 1) connected to the left of the stainless steel U-trap. In summary, CH4 in headspace is sampled from a septum capped bottle via gas-tight syringe that is pre-flushed with He to eliminate atmospheric CO2 contamination. The sample is injected slowly (<100 mL/min) via septum fitting and needle valve into a flow-through vacuum oxidation line with a carrier of ultra-zero compressed air (UZ air). Maintaining a low injection flow is critical to ensuring the efficiency of the traps and combustion of methane (Pack et al. Reference Pack, Xu, Lupascu, Kessler and Czimczik2015). The carrier is purified with an in-line CO2 trap (EMASorb A, Elemental Microanalysis B1115) and held at a flow rate of 10 mL/min through the line. CO2 and water are preliminarily removed from the sample on a dual chemical trap consisting of EMASorb A and Magnesium Perchlorate mixed with 1/3 Quartz chips (Elemental Microanalysis, B1019 + B1271). This is followed by cryogenically trapping of residual CO2 and water from the sample on a LN2 trap. CO is removed on a subsequent trap consisting of a 290°C oven packed with CuO rods (Fisher Scientific, C474) to oxidize CO to CO2, followed by a second LN2 trap. Methane is combusted to CO2 in a 975°C tube furnace packed with CuO rods (regenerated by the constant flow of UZ air) and Silvered Cobaltous/Cobaltic oxide towards the bottom of the reactor to remove S-bearing species and halides. Any water produced from the combustion is separated from the CO2 on a −80°C trap, and the CO2 from the methane combustion is trapped on a final LN2 trap. Following complete combustion of the sample (∼10 min), the purified CO2 is transferred to the vacuum section of the line (Figure 1B) and sealed in a breakseal.
Graphitization and AMS Analysis
Following CO2 purification, all samples are converted to graphite by hydrogen reduction on custom-build semi-automated graphitization lines (see St-Jean et al. Reference St-Jean, Kieser, Crann and Murseli2017). Graphite samples are pressed into sample targets and subsequently analyzed on a 3MV tandem accelerator mass spectrometer (Kieser et al. Reference Kieser, Zhao, Clark, Cornett, Litherland, Klein, Mous and Alary2015). 14C data are reported as fraction modern carbon (F14C), calculated according to Reimer et al. (Reference Reimer, Brown and Reimer2004) as the ratio of the sample 14C/12C ratio to the standard 14C/12C ratio (Ox-II). The 14C/12C ratios of the samples are background corrected (using the corresponding process blank) and the results of both the samples and standards are corrected for spectrometer and preparation fractionation using the AMS-measured 13C/12C ratio and normalized to the δ13C (PDB) of –25‰. 14C ages are calculated as –8033ln(F14C) and reported in 14C yr BP (BP = AD 1950) as described by Stuiver and Polach (Reference Stuiver and Polach1977). For regular sized samples between 0.7–1.5 mgC, the typical measurement precision reported is < = 3‰ for modern samples. To define limits of detection, if the sample measures within 2σ of the average process blank performance for that material type of the same size (which includes blank contribution from sample processing and measurement), we define this as a “non-finite” or “greater-than” measurement and the results are reported as such. For additional information regarding sample processing at AEL-AMS, see Crann et al. (Reference Crann, Murseli, St-Jean, Zhao, Clark and Kieser2017).
RESULTS AND DISCUSSION
Table 1 summarizes the reference materials used for each material code, including process blank, secondary and modern reference materials. DIC/DOC reference materials are freshly prepared for each batch, where solid materials are dissolved in 20 mL of MilliQ water to yield 1 mgC (ideal), or size-matched to the samples processed alongside. Excellent yields (>95%, calculated from calibrated volume on multipurpose line) are obtained for all sample types. Procedural blank contributions are on the order of 5–7 µg modern carbon for DIC and DOC, and <1 µg modern carbon for CX and CH4. Average performance of each standard is quoted in Table 1.
Performance of all process blanks for typical sized samples (0.7–1.5 mgC) are illustrated in Figure 2. During this 2-yr period, 34 DIC (NaHCO3) and 53 DOC (kHP) standards were analyzed. The slightly elevated DIC blank samples (>1 pMC; 03-2016 and 03-2017) are likely attributable to atmospheric adsorption of CO2 on the stock of NaHCO3, despite being stored in a desiccant cabinet. Heating the powder in a 60ºC drying oven for 1 hr prior to use helps to reduce the blank. For DOC samples, a clear reduction of the blank is observed in early 2017 when a new stock of sodium persulfate was opened (same catalogue number, different lot number). Blanks are consistently reduced thereafter; demonstrating the importance of verifying the purity of each chemical lot. With the new stock of persulfate, process blanks are now consistently <0.7 pMC (>40,000 14C yr BP) for water samples, averaging around 0.55 and 0.44 pMC for DIC and DOC, respectively. This value is comparable to other DOC extraction methods such as UV oxidation (∼0.7 pMC blank; Druffel et al. Reference Druffel, Griffin, Walker, Coppola and Glynn2013), and somewhat improved compared to other previously cited references including wet oxidation and evaporation (∼1.0 pMC blank: Burr et al. Reference Burr, Thomas, Reins, Jeffrey, Courtney, Jull and Lange2001; Leonard et al. Reference Leonard, Castle, Burr, Lange and Thomas2013). We attribute the variability of the blank for both DIC and DOC to be related to modern carbon contamination during sample processing, from the bottles/septa, reagents, and He-carrier flow through line. Tests with varying amounts of MilliQ (10–100 mL) and AgNO3 (1–10 mL) showed no significant change to the DOC blank. As expected, the relative contribution of the blank is elevated for smaller sized samples, illustrated in Figure 3. At ∼0.2 mgC, the blank is ∼3.6 pMC for DOC and ∼1.7 pMC for DIC; this indicates that the majority of the carbon contamination is contributed from the persulfate oxidation.
Figure 2 Process blank performance of water and gas samples at AEL by material code for standard sample sizes (0.7–1.5 mgC).
Figure 3 Blank contribution relative to sample size for waters (DIC and DOC).
For secondary references, an average value of 41.06 ± 0.25 pMC was obtained from 30 analyses of IAEA-C2 (DIC), which agrees well with the consensus value. 4 SWRNOM and 6 OXII (DOC) secondary standards were run with average values of 106.12 ± 0.32 and 132.61 ± 0.45 pMC, respectively. The slightly depleted OXII value may be attributable to the fact that oxalic acid oxidizes very easily (Brett Walker, personal communication), and may have been pre-oxidized during the DIC sparge.
A total of 16 CX and 17 CH4 blank standards (14C-free gas mixtures) were analyzed, all returning exceptionally low blanks, with values typically <0.1 pMC (>55,000 14C yr BP) and varying less than 0.2 pMC. The CX and CH4 standards are more reproducible than the DIC and DOC materials as less sample preparation and handling is involved. Secondary reference materials for gas samples are not available at this time.
Highlights of the sample preparation method for waters include relative ease of preparation and high throughput. For DIC, typically 3–4 samples and 2 standards can be prepared and extracted in one day. DOC samples are slightly more labor intensive requiring a 2-day sample prep process since it is necessary to remove the DIC fraction prior to oxidation. Tests were completed leaving the reacted bottles 1, 3, and 7 days following heating, with no change observed with respect to sample yield or blank contribution, owing to the integrity of the butyl septa. Therefore, many samples (up to 20) can be prepared at once and left for up to 1 week until extracted, facilitating sample preparation. For gas samples (CX/CH4) where there is minimal preparation required, typical throughput is ∼8 samples per day. The wet chemical oxidation technique for DOC described in this paper has some disadvantages, namely that it can only be applied for non-saline waters as high concentrations of dissolved chloride will interfere with the persulfate oxidation. For 13C stable isotope IRMS analysis, using the wet oxidation technique, the present routine limit for salt content is 1.5% (brackish water), which is considerably less than seawater (3.5%). So far, this limit has not been tested for the 14C sample preparation. Small concentration samples (<1 ppmC) also prove challenging due to the large volume of water needed and limited reaction bottle size (max. 700 mL sample volume in 1000 mL bottle); however, multiple aliquots can be combined for a larger sample yield.
CONCLUSION AND FUTURE WORK
New techniques for preparation of water (DIC/DOC) and gas (CO2/CH4) samples for 14C analysis at AEL-AMS highlight wet oxidation for dissolved organic carbon in freshwaters. Samples are prepared with relative ease and reasonably high throughput for a manual technique (5–8 per day) as many samples can be prepared at once prior to extraction. Measurement of reference materials demonstrate our ability to process water and gas samples with low procedural blanks: <0.7 pMC (>40,000 14C yr) for DIC/DOC, and <0.1 pMC (>55,000 14C yr) for CH4/CX. Work underway to improve sample processing of waters and gases at AEL-AMS includes a number of manual and automated method development projects:
Modification of the gas extraction line to accommodate larger volumes for ppm concentration gas mixtures (including atmospheric CO2 and CH4).
Interface a robotic auto-sampler with a semi-automated gas cleanup line (St. Jean et al. Reference St-Jean, Kieser, Crann and Murseli2017) for rapid analysis of carbonates, CO2 in gas mixtures and waters (DIC and DOC).
Adapt the wet oxidation method for DOC for brackish waters.
Design and build a semi-automated, multi-capacity low-background UV-oxidation system for saline and low DOC concentration water samples.
ACKNOWLEDGMENTS
This work was funded by the Canadian Foundation for Innovation and the Ontario Research Fund. We gratefully acknowledge the expertise and support from the G. G. Hatch Stable Isotope Laboratory for helping to get this method off the ground and running. We also thank Xiaomei Xu and Brett Walker for their helpful discussions. Thanks to Natalia Baranova, Mike Grinter, and Anthony Lapp for their pioneering work on sample preparation, and to Emily Barber and Karolina Krym for countless hours extracting samples. We thank the reviewers for their comments in helping to improve the manuscript.
