Electronic Supplementary Material

For

Removal of the Sesquiterpene β-Caryophyllene from Air via Biofiltration: Performance Assessment and Microbial Community Structure

William M. Moe*, Weili Hu, Trent A. Key, Kimberly S. Bowman

Department of Civil and Environmental Engineering, 3515B Patrick Taylor Hall, Louisiana State University, Baton Rouge, LA 70803, USA

*Corresponding author

Tel: (225) 578-9174

Email:


Enrichment Culture Development

A sparged-gas bioreactor configuration similar to that reported previously for development of enrichment cultures able to biodegrade a variety of volatile organic compounds (Lee et al., 2002; Atoche and Moe, 2004; Moe and Qi, 2005; Qi and Moe, 2006) was employed to develop an enrichment culture. The 4.0 L glass kettle reactor (Pyrex, Acton, MA) was filled with 2.5 L of nutrient solution containing the following constituents added to tap water: NH4NO3 1.25g/L, KH2PO4 1.0 g/L, MgSO4·7H2O 0.5 g/L, CaCl2·2H2O 0.02 g/L, CuCl2·2H2O 0.17 mg/L, CoCl2·6H2O 0.24 mg/L, ZnSO4·7H2O 0.58 mg/L, MnSO4·H2O 1.01 mg/L, Na2MoO4·2H2O 0.24 mg/L, NiCl2·6H2O 0.10 mg/L and FeSO4·7H2O 1.36 mg/L. The reactor was inoculated with a 0.5 L suspension of commercially available potting soil (Showscape Potting Soil, Phillips Bark, MS, USA) comprised of ground and composted organic forest material, sand, and perlite. Air, at a flow rate of 1.0 L/min, entered the reactor via a gas diffuser stone submerged in the liquid medium. β-caryophyllene was delivered to the influent air supply by manually injecting it into a glass, septum-filled injection port at repeated intervals when it was visually observed that the previous injection had mostly evaporated (approximately 3-day intervals).

On a daily basis, after adding DI water to reach a total volume of 3.0 L in the reactor (to compensate for evaporative losses), 100 mL of the mixed liquid was removed and 100 mL of nutrient solution was added while the reactor remained mixed. This resulted in a hydraulic residence time (HRT) of 30 days. While there was some growth in the aqueous phase (total suspended solids concentration ranging from 19 mg/L to 72 mg/L over the duration of operation), a majority of the biomass was present in the reactor as a fixed film growing attached to the glass sidewalls and inner lid surface above the water level.

At the time of biofilter inoculation (182 days after startup of the sparged gas reactor), biomass growing attached to the sidewall and inner lid surface of the reactor (which had a surface temperature of 34°C) was scraped off and mixed with 500 mL of reactor liquid. The resulting suspension was then homogenized in a laboratory blender before use as inoculum for the laboratory biofilter.

Abiotic Adsorption Test

After the biofilter column was initially assembled but prior to microbial inoculation, β-caryophyllene was supplied to the system to assess the abiotic adsorption capacity of the polyurethane foam packing medium. The experimentally measured breakthrough curve during β-caryophyllene loading to the abiotic column (prior to inoculation) is depicted in Fig. S1. As shown, 5% pollutant breakthrough occurred within one day, and 95% pollutant breakthrough occurred after five days of continuous loading. Mass balance calculations revealed that after complete breakthrough was achieved, the contaminant mass entering and exiting the biofilter column differed by 0.939 g C. Assuming that all of the pollutants measured as C were comprised of β-caryophyllene and using the empirical formula for β-caryophyllene (0.882 g C / per g β-caryophyllene based on the formula C15H24), the pollutant mass accumulating in the biofilter column was calculated to be 1.06 g β-caryophyllene. The corresponding mass of β-caryophyllene adsorbed per unit mass of polyurethane foam was calculated to be 3.63 mg/g.

Fig. S1 Experimentally measured effluent concentration during the abiotic adsorption test conducted prior to biofilter inoculation.

High Purity Versus Low Purity β-Caryophyllene

During the last five days of Period 1 (days 35-40), higher purity β-caryophyllene (>98.5% purity as opposed to >90% purity) was supplied to the biofilter. Fig S2 depicts the influent and effluent concentrations for five days (days 30-35) low purity β-caryophyllene (>90% purity) supplied to the biofilter following five days higher purity β-caryophyllene (>98.5% purity) was applied. As shown in the figure, the mean influent concentration for the higher purity β-caryophyllene duration was 85.89±2.67 ppm C, it did not differ from the low purity β-caryophyllene duration of 83.7±4.0 ppm C. The effluent concentration was quite stable through the ten days at the value of 3.3±0.2 ppm C.

Fig. S2 Influent and effluent concentrations from high purity β-caryophyllene experiment at the end of Period 1.

On day 120, higher purity β-caryophyllene (>98.5% versus >90%) was supplied to the biofilter for a duration of 4.85 days. Fig S3 depicts influent and effluent pollutant concentrations during this short period of time. As it shown, the influent concentration was 90.9±1.2 ppm C, essentially the same as the previous influent concentration measurement, and the effluent concentration was 3.34±0.37 ppm C which is quite similar to the effluent mean of 3.3±0.2 ppm C in Period 3B.

Fig. S3 Influent and effluent concentrations measured during high purity β-caryophyllene test in Period 3B.

Scanning Electron Microscopy

A representative scanning electron microscopy (SEM) image of packing medium samples collected from the biofilter on day 145 are shown below in Fig. S4. A large majority of the cells were rod-shaped (0.2 to 0.5 µm in width × 1 to 7 µm in length) or cocci (0.2 to 1 µm diameter), consistent with the morphology of bacteria. Some images revealed the presence of what appeared to be filamentous fungi and higher organisms (e.g., nematodes); however, their relative abundance was generally low.

Fig. S4 SEM image of biofilter packing medium sampled from the inlet section of the biofilter on day 145. Bar represents 1.0 µm.


Physicochemical Properties of b-caryophyllene

Because experimental data are not available regarding several physicochemical properties of b-caryophyllene, estimation methods were applied using models freely available in the EPI SuiteTM version 4.10 software package (US EPA, 2011, http://www.epa.gov/oppt/exposure/pubs/episuitedl.htm). This approach has been utilized previously to estimate properties of other sesquiterpenes (Jenner et al., 2011). Tabulated results are provided in Table S1.

Table S1: Physicochemical properties of b-caryophyllene

Parameter / Value
Molecular Formula / C15H24
CAS # / 87-44-5
Molecular Weight (g/mole) / 204.36
Density at 20ºC (g/mL) / 0.902
Boiling point (°C) / 262-264
Log KOW (dimensionless) / 6.30a
Water Solubility at 25°C (mg/L) / 0.05 b -0.54 c
Henry’s Law Constant at 25°C (atm·m3/mol)
(unitless) / 0.69d
28.2d
Vapor Pressure (Pa at 25 °C) / 4.16e

a KOW=octanol-water partition coefficient, estimated using WSKOWWin version 1.67, atom/fragment contribution method.

b Water solubility estimated using WSKOWWin version 1.41 regression equation.

c Water solubility estimated using WATERNT version 1.01.

d Henry’s Law constant estimated using the Bond contribution method in HenryWin version 3.20.

e Vapor pressure estimated as the mean of the Antoine and Modified Grain Method in MPBPWin version 1.43.

The estimated vapor pressure for b-caryophyllene shown in Table S1 is somewhat higher than the value of 1.1 Pa estimated using the alternative approach of Hoskovec et al. (2005).


References Cited

Atoche JC, Moe WM (2004) Treatment of MEK and toluene mixtures in biofilters: Effect of operating strategy on performance during transient loading. Biotechnol Bioeng 86:468-481. doi: 10.1002/bit.20064

Hoskovec M, Grygarova D, Cvačka J, Streinz L, Zima J, Verevkin SP, Koutek B (2005) Determining the vapour pressures of plant volatiles from gas chromatographic retention data. J Chromatography A 1083:161-172. doi: 10.1016/j.chroma.2005.06.006

Jenner KJ, Kreutzer G, Racine P (2011) Persistency assessment and aerobic biodegradation of selected cyclic sesquiterpenes present in essential oils. Environ Toxicol Chem 30:1096-1108. doi: 10.1002/etc.492

Lee S, Moe WM, Valsaraj KT, Pardue JH (2002) Effect of sorption and desorption-resistance on aerobic trichloroethylene biodegradation in soils. Environ Toxicol Chem 21(8):1609–1617. doi: 10.1897/1551-5028(2002)021<1609:EOSADR>2.0.CO;2

Moe WM, Qi B (2005) Biofilter treatment of volatile organic compound emissions from reformulated paint: Complex mixtures, intermittent operation, and startup. J Air Waste Manag Assoc 55:950-960. doi: 10.1080/10473289.2005.10464687

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