*2.3. Laboratory Microcosm Experiments*

The microcosm was established according to Ting et al. [17] with modifications. Vertical up-flow columns were used to stimulate the release of Hg compounds and examine the efficiency of sorbent cappings on Hg sequestration (Figure 1). The dimensions of the column with an internal height of 15 cm and an internal diameter of 6 cm with glass fiber at the bottom. To investigate how the Hg-contaminated estuary sediment affected by salinity and to understand the stabilizing efficiency of AC, SAC, and FeS cappings, two systems, the freshwater and estuary systems, were set up. Each of the four columns contained 300 g of dried Hg-contaminated sediment from the An-Shun site. The total number of column is 8. Column A was capped with AC (9 g; i.e., 3 wt% AC added); column B was capped with SAC (9 g; i.e., 3 wt% SAC added); column C was capped with FeS (9 g; i.e., 3 wt% FeS added); column D was without capping as the control unit.

**Figure 1.** The diagram of vertical up-flow microcosms construction.

To start up the microcosms, dried Hg-contaminated sediment was firstly added into the column, filled with waters and waited for 24 h to settle. The microcosms were activated to circulate the waters in the column for sediment acclimation and stabilization, and counted as the operation day 1. The capping materials were applied on day 25. The total volume of water used for each experiment was 1.5 L. The water first entering into the bottom of the column was referred as inflow, then vertically moved upwards to fill the column, and then discharged through the outlet. The effluent water were stored in a temporary buffer tank and recirculated into the bottom of column in a closed system by using a peristaltic pump (Lead Fluid BT100S, Hebei, China) to maintain a flow rate of 1 mL/min. The reason that the effluent water was recirculated back to the microcosm column is to establish the mass balance of THg partitioning in various phases (i.e., sediment, capping material, overlying water) in a closed system and evaluate the THg accumulation ability of the various capping materials.

Periodic sampling was in progress while the microcosms were in operation. Each time, 100 mL of water sample was collected from the tube of outflow. After that, the temperature, pH (SunTex SR-2300, New Taipei City, Taiwan), dissolved oxygen (DO; Extech EXStik DO600, Nashua, NH, USA), electrical conductivity (EC; Taina EZDO 6021, Taichung, Taiwan), oxidation reduction potential (ORP; SunTex SR-2300, New Taipei City, Taiwan), trace metals concentration, total Hg (THg), MeHg, DOM, and anions were analyzed. The temporary buffer tank was then refilled with the artificial waters to maintain a constant water amount.

To measure the trace compound concentration, including metals, THg, MeHg, DOM, and anions, the water samples were filtered with 0.45 μm mixed cellulose ester filter (DISMIC-25AS, Toyo Roshi Kaisha, Tokyo, Japan). Trace metals were determined by Inductively couple plasma optical emission spectrometry (Agilent 700, Santa Clara, CA, USA) after the sample was acidified with 0.15% HNO3. Water samples for THg analysis were preserved by adding 0.5% BrCl2 solution and stored in 20 mL glass bottles. THg in sediment and water was analyzed following the USEPA Method 1631 and NIEA W331.50B protocols by using CVAFS. The water samples were preserved by adding 0.2% HCl and stored in 20 mL amber glass bottles in MeHg analysis. MeHg in sediment and water was analyzed following the USEPA Method 1630, NIEA W341.60B and NIEA W540.50B procedures by using gas chromatography/CVAFS (Brooks Rand MERX Integrated Automated MeHg Analyzer, Seattle, WA, USA).

Recovery of Hg in the microcosm system was evaluated based on the Hg concentration in overlying water, cap materials, and sediment. To determine the Hg concentration in cap materials and sediment, digestion was first conducted in a microwave system (Ethos 1600, Milestone, Shelton, CT, USA) with a power setting of 800 W (USEPA method 3051a). After digestion, the THg (μg/mg) in the solid

phase was determined using CVAFS (USEPA method 245.7). QA/QC of data were confirmed based on Hsu et al. [29] and Wang et al. [36]. The recovery values for the QC samples of sediment of THg (NIST 2709a, 0.9 mg/kg) and MeHg (SQC-1238, 10 μg/kg) were 98.6 and 90.0%, respectively. For spiked sediment, the recovery values of THg and MeHg were 94.8 and 95.2%, and were 107.1 and 101.4% for water, respectively. The coefficient of determination (R2) of CVAFS for the aqueous Hg was regularly kept larger than 0.998, the recovery was within 96.2–120%, the precision was within 0.04–5.60%.
