BIODEGRADATION OF MONOETHANOLAMINE, ETHYLENE

GLYCOL AND TRIETHYLENE GLYCOL IN LABORATORY

BIOREACTORS
OLE MRKLAS1, , ANGUS CHU2 , STUART LUNN3 and LAURENCE R. BENTLEY1
1
Department of Geology and Geophysics, University of Calgary, 2500 University Drive NW,
Calgary, AB T4N 1N4, Canada; 2 Department of Civil Engineering, University of Calgary, 2500
University Drive NW, Calgary, AB T4N 1N4, Canada; 3 Imperial Oil Resources, 237 Fourth Avenue
SW, Calgary, AB T2P 0H6, Canada
(*author for correspondence, e-mail: omrklas@ucalgary.ca, Tel: 403.237.4289)

(Received 8 April 2003; accepted 9 June 2004)

Abstract. The release of alkanolamines and glycols into the subsurface soils poses a potential hazard
to the environment through impacted soil and groundwater. This study investigated aerobic and
anaerobic biodegradability of monoethanolamine (MEA), ethylene glycol (MEG) and triethylene
glycol (TEG). Significant levels of MEA (31 000 mg/kg), MEG (500 mg/kg) and TEG (2100 mg/kg)
were successfully aerobically biodegraded in bioreactors. The aerobic slurry experiments suggested
initial phosphate (P) limitation, as biodegradation rates increased by one order of magnitude after
phosphate addition. Anaerobic decay of MEA, MEG and TEG was unaffected by P-addition. MEA,
MEG and TEG degradation products such as acetate, ethanol and ammonium at about 75 000 mg/kg,
8100 mg/kg and 8800 mg/kg degraded completely and did not prevent aerobic biodegradation. This
study confirms proposed biodegradation pathways of MEA, MEG, TEG and their breakdown products
in natural soil and groundwater using indigenous microbes. Levels of contamination studied here are
significantly higher than previously reported.
Keywords: biodegradation, contamination, degradation products, ethanolamine, glycol, groundwater,
subsurface

1. Introduction
Alkanolamines and glycols have been commercially available for decades. The
utilization of these substances includes both household and industrial applications.
Glycols and alkanolamines are extensively employed in the sour natural gas industry. Sour natural gas production is a significant portion of the global energy
markets. Alberta produces the majority of Canadian natural gas with a sour natural gas portion of approximately 40% (Sorensen et al., 1999). Alkanolamines are
basic derivatives of ammonium. The optimization and modification of the sweetening process has resulted in a diverse composition of alkanolamine and glycol
mixtures for specific applications (Polasek et al., 1992; Rooney et al., 1998). Unintentional introduction of ethanolamine, ethylene glycol and triethylene glycol into
Water, Air, and Soil Pollution 159: 249263, 2004.
C 2004 Kluwer Academic Publishers. Printed in the Netherlands.


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the subsurface and groundwater is a potential hazard to the environment (Fedorak

et al., 1997; Gallagher et al., 1995; Lintott et al., 1997; Sorensen et al., 1999;
Wrubleski et al., 1997). After their release, alkanolamines and glycols may migrate
into the subsurface and groundwater. The high density and high water solubility of
these species influence their fate and transport in the subsurface and in groundwater (Sorensen et al., 1999). The combination of chemical properties and structures
inherent to these species makes alkanolamines and glycols readily biodegradable.
However, the behavior of alkanolamines in concentrations exceeding 3000 mg/L in
the presence of high levels of aerobic and anaerobic breakdown products (i.e. ammonium, acetate and ethanol) is not well understood, especially in environmental
applications. In fact, studies looking at in-situ fate and transport of alkanolamines,
glycols and degradation products have been complicated by complex geological and
hydrogeological properties (McVicker et al., 1997; Sorensen et al., 1999; StrongGunderson et al., 1995). In addition, it was demonstrated that MEA concentrations
exceeding 1500 mg/kg inhibited in-situ biodegradation (Sorensen et al., 1997).
This study assesses the biodegradability of MEA, MEG and TEG in laboratory bench scale bioreactors. Major breakdown products of MEA, MEG and TEG
biodegradation are ethanol, acetic acid and ammonium (Bradbeer, 1965; BUA,
1994; Jones and Turner, 1973; McVicker et al., 1997). Ethanol and acetic acid may
be completely degraded via the tri carboxylic acid cycle (aerobic) or by methanogenesis (anaerobic) (Gottschalk, 1985). Ammonium degrades by nitrification and
denitrification using shift conditions (aerobic followed by anaerobic conditions) and
carbon source supplementation. The evolution of MEA, MEG and TEG and their
breakdown products ammonium, acetic acid and ethanol is discussed with respect to
the implications for in-situ aerobic and anaerobic degradation at a decommissioned
gas plant site. Indicators such as electrical conductivity and pH measurements may
assist in monitoring degradation patterns. Explaining aerobic and anaerobic degradation pathways in laboratory slurry phase experiments will assist in understanding
in-situ processes and will aid in the design of optimal degradation conditions for
remediation regimes.

2. Methods and Materials

2.1. CHEMICALS
All chemicals were ACS grade, and purchased from Sigma-Aldrich, Canada or
Fisher Scientific, Canada.
2.2. BIOREACTORS
All bench scale bioreactors utilized the same source of soil and groundwater from
the site at a concentration of 40% w/w (100 g soil + 150 g groundwater). Initial

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251

soil and groundwater sample analyses indicated that the soil samples showed ammonium, but no MEA concentrations and groundwater samples had both ammonium and MEA concentrations. The soil and groundwater samples originated from
the former process area of a decommissioned sour gas plant site from a depth of
about 4 m. This location was chosen, because previous electrical resistivity tomography (ERT) measurements suggested high impact levels. The slurries consisted
of four aerobic (1, 2, 3, 4), two replicate aerobic-abiotic controls, two replicate
anaerobic, and two replicate anaerobic-abiotic controls. The abiotic control slurries were autoclaved three-times on three consecutive days for 1 h at approximately
100 C in autoclave bags. Soil and water samples were analyzed to determine initial concentrations after autoclaving. All Erlenmeyer flasks and glass jars were
autoclaved prior to use. Aseptic sampling procedures were used for the abiotic
controls.
2.2.1. Aerobic Slurries
The aerobic bioreactors were contained in four biotic (labeled 1, 2, 3, 4) and two
abiotic open 500 mL Erlenmeyer flasks. The flasks were wrapped with tin foil
to prevent algae growth. The flasks were mounted onto a shaker table and mixed
gently at constant speed (approximately 300 rpm). The speed was adjusted to ensure
no sedimentation. The geometry of the Erlenmeyer flasks and the fill level (about
3 cm) facilitated optimal mixing and maximal slurry/air interface area for oxygen
transfer. The aerobic bioreactors were weighed during each sampling interval and
sterile distilled deionized water was added to adjust for evaporation. The aerobic
abiotic reactors were fitted with a porous foam stopper, but were not compensated
for evaporation in order to preserve aseptic conditions. Reactors 3 and 4 received
about 40 mg (P)/kg phosphate addition using potassium-di-hydrogen-phosphate
(KH2 PO4 ) on day 11. Slurries 1 and 2 received the same treatment on day 64 of the
experiment. On day 92, slurries 3 and 4 were transferred into an anaerobic glove
box and received 200 mM acetic acid. MEA, MEG, TEG and degradation product
(acetic acid, ethanol, ammonium, nitrite, nitrate) concentrations were monitored.
The aerobic bioreactors were monitored at ambient temperature of 23 C 0.7 C.
2.2.2. Anaerobic Slurries
A dry glove box with continuous nitrogen gas flow under positive pressure created
an oxygen-free environment for two biotic and two abiotic anaerobic slurries. Approximately 250 g slurry material (40% w/w soil) was contained in each 500 mL
glass jar that was wrapped with aluminium foil to prevent algae growth. The lids
were equipped with a pressure release line open to the nitrogen atmosphere. The
slurries were mounted onto magnetic stirrers and intermittently stirred at constant
speed approximately 1 h before each sampling. Anaerobic slurries received 40 mg
(P)/kg phosphate as KH2 PO4 on day 11. The anaerobic slurries were held at constant
temperature of 22 C 1 C.

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O. MRKLAS ET AL.

2.3. ANALYTICAL

PROCEDURES

The slurry sampling procedure involved the removal of about 1 mL of slurry material from the bioreactors. The samples were centrifuged at 10 000 rpm for 5
min. The supernatant was completely removed (water extract). Subsequently, the
remaining soil at a determined moisture content was extracted with 1 M potassium hydroxide (KOH) solution, mixed for 1 h and centrifuged (KOH extract).
The KOH extraction procedure was found to effectively remove sorbed MEA and
ammonium left on the soil after the original soil water extraction. Water and KOH
extracts were analyzed. MEA, ammonium, sodium, magnesium and calcium were
quantified using cation exchange chromatography with suppressed conductivity detection in water mode. Acetic acid, chloride, nitrite, nitrate, phosphate and sulfate
were determined by anion exchange chromatography with chemical suppressed
conductivity detection. Anions were monitored in the water extract. MEG, TEG
and ethanol determinations were carried out on water extract by ion exclusion chromatography with pulsed amperometric detection. Detailed analytical protocols are
described elsewhere (Mrklas et al., 2003; Mrklas, 2002). In addition, during each
sampling interval all slurries were monitored for electrical conductivity (EC), pH
and temperature. The EC values were corrected to a reference temperature of 20 C
using external calibration with EC standards at various temperatures and linear
regression.

3. Results
The analyses of the initial mixture of collected soil and groundwater samples resulted in concentrations of approximately 31 000 mg/kg MEA, 500 mg/kg MEG
and 2100 mg/kg TEG (Table I). The concentrations are comparable to in-situ levels found on-site. Table I outlines the average product and degradation product
concentrations in the soil, groundwater and slurry samples at the beginning of the
aerobic and anaerobic slurry experiments. Additionally, initial (final) phosphate
concentrations, pH and EC were recorded (Table I). Table II summarizes the degradation pathways and Tables III and IV show the biodegradation rates for the various
species.
3.1. A EROBIC

BIOREACTORS

The aerobic reactors were monitored for 98 days. Duplicate reactors showed similar behavior and results of replicates are averaged here. Results from slurries 1
and 2 were averaged (labeled +P64) and monitored without phosphate enhancements for the first 64 days, and with P-enhancements (40 mg P/kg) after day 64
(Figures 1AC). The results of biorectors 3 and 4 were averaged (labeled
+P11). Biodegradation of MEA, MEG and TEG was monitored without P-addition

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BIODEGRADATION OF MEA, MEG AND TEG

TABLE I
Characterization of initial groundwater, soil and slurry concentrations
Concentration
Species

Groundwater(mg/L)

Soil(mg/kg)

Slurry(mg/kg)

MEA
MEG
TEG
Ammonium
Acetic acid
Ethanol
Phosphate, initial (final)
pH
EC(mS/m)

16205 463
NA
NA
3034 75
36410 1137
3938 82
NA
NA
NA

ND
NDa /ND
490a /514
2276 281
ND
ND
NA
7.5a
1.4a

30911 437
474 32
2108 279
8843 64
75139 2347
8127 169
<0.6 (<0.6)
7.0 0.3
15.8 0.9

NA: not analyzed.

ND: not detected.
a
External analyses.
TABLE II
Degradation pathways of MEA, MEG and TEG
Condition

MEA

MEG, TEG

Pathway

Aerobic

Acetic acid, ethanol

ammonium
Acetic acid, ethanol
ammonium
MEA

Acetic acid, ethanol

TCA-cycle
nitrification
Methanogenesis
accumulation
Sorption

Anaerobic
Abiotic

Acetic acid, ethanol

MEG, TEG

until day 11 and with P-addition after day 11. Figure 1A shows aerobic
degradation patterns of acetic acid and ethanol. Figure 1B depicts MEG and
TEG concentrations and Figure 1C depicts MEA averaged measurements +P64
and +P11.
The ethanol concentrations declined from an initial average concentration of
8127 mg/kg to below detection after day 7 and followed similar degradation patterns as indicated by average +P64 and +P11 concentrations. Ethanol degradation
proceeded without a lag period (Figure 1A).
Initial acetic acid concentrations are shown in Table I and first-order biodegradation rates for acetic acid are presented in Table III. The initial acetic acid levels
declined slowly pre P-addition and significant rate increases were observed after
P-addition (Figure 1A). Average aerobic acetic acid biodegradation rates (+P11)
increased approximately 25 times after P-addition. The acetic acid concentrations
declined below detection 10 days after P-addition. The averaged aerobic values

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O. MRKLAS ET AL.

TABLE III
Average degradation rates (/S.D.) of carbon species in aerobic bioreactor sets +P64
and +P11
+P11

+P64a

Compound

Aerobic (day1 )

Aerobic P-enhanced (day1 )

Aerobic (day1 )

Ethanol
MEG
Acetic acid
MEA
TEG

0.56 0.04
0.067 0.001
0.018 0.006
0.011 0.001
0.0115 0.0003

0.47030 0.00001
0.10 0.03
0.145 0.001

0.56 0.04
0.02
0.014 0.005
0.0078 0.0001
0.005 0.002

Bioreactor +P64 degradation rates P-enhanced not shown due to insufficient sampling frequency.

TABLE IV
Average anaerobic biodegradation rates(/S.D.)
Compound

Anaerobic (day1 )

Degradation kinetics

Acetic acid
Ethanol
MEA
TEG

0.0054 0.0001
0.0040 0.0007
0.0022 0.0002
0.0023 0.0003

first order
first order
zero order
zero order

+P64 showed steady acetic acid decay until day 64 and significant rate increases
were observed after day 64. (Figure 1A). The abiotic controls showed steady acetic
acid concentrations.
The MEG concentrations decreased initially at higher rates in both averaged
values +P64 and +P11. Average MEG concentrations (+P64) decreased by 36%
until day 7 and then did not significantly change until phosphate addition on day
64. MEG concentrations (+P11) declined to below detection after day 15 and
after P-addition on day 11 (Figure 1B). The abiotic controls showed steady MEG
concentrations.
MEA biodegradation had zero-order biodegradation rates (Figure 1C). Prior
to P-addition the average MEA biodegradation rates +P11 and +P64 were similar (Table III). However, these rates increased by one order of magnitude after
P-addition in both reactor sets (Figure 1C). The abiotic controls showed steady
MEA concentrations. Biodegradation rates and standard deviations are presented in
Table III and followed zero-order degradation kinetics.
MEA and ammonium partitioned into soil and water fractions (Figures 1C).
The KOH extraction method resulted in nearly complete desorption of MEA
and ammonium as indicated of recovery rates of 109 and 97% (n = 3),

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BIODEGRADATION OF MEA, MEG AND TEG

A
P-addition day 11

P-addition day 64

100000
C [mg/kg]

80000
60000
40000
20000
0
0

20

40

60

80

100

Time [d]
Ac +P64

Ac +P11

Ac abiotic

EtOH +P64

EtOH +P11

B
P-addition day 11

P-addition day 64

2500
C [mg/kg]

2000
1500
1000
500

0
0

20

MEG +P11

40
MEG +P64

60
Time [d]
TEG +P64

80

100

TEG +P11

C
P-addition day 11

P-addition day 64

C [mg/kg]F

40000
30000
20000
10000
0
0
MEA T+P11

10

20

30

MEA W+P11

40

50
Time [d]

MEA S+P11

60

70
MEA abiotic

80

90

100

MEA T+P64

Figure 1. Average (a) acetate and ethanol concentrations, (b) MEG and TEG concentrations and (c)
MEA concentrations in soil (S), water (W) and total (T) in [mg/kg] during aerobic biodegradation
versus time in [days]. Phosphate addition occurred on day 11 (+P11) and day 64 (+P64) as indicated
by the arrows. Error bars represent one standard deviation.

256

O. MRKLAS ET AL.

respectively. The recovery rates were calculated as detected MEA and ammonium concentrations. The total concentrations [mg/kg] were calculated as the sum
of water extract concentration in [mg/kg] and the amount in the KOH extract in
[mg/kg].
TEG biodegradation rates +P11 increased by one order of magnitude after day
21 and reached below detection levels after 32 days post P-addition (Figure 1b).
Average TEG biodegradation rates +P64 were slower until the phosphate addition
on day 64. The abiotic controls showed steady TEG concentrations.
The carbon utilization in all aerobic bioreactors occurred in the order ethanol,
MEG, acetic acid, MEA and TEG. The EC values followed similar trends as
acetic acid and MEA concentrations in the biotic and abiotic reactors, until day 60
where elevated nitrite and nitrate concentrations caused an increase in EC values
(Figures 1AC, 2 and 3).
3.2. NITROGEN

DISTRIBUTION

The aerobic slurries were monitored for nitrogen containing species MEA, ammonium, nitrite and nitrate. Figure 3 shows averaged nitrogen concentrations +P11.
The concentration changes (+P11) of nitrogen containing species observed were
divided into four phases labeled A, B, C, and D. Phase A represents pre-phosphate
enhanced conditions and spanned the first 11 days of the experiment. Increased
carbon utilization due to P-enhancement occurred in phase B and aerobic nitrogen transformation from ammonium to nitrite and nitrate is seen in phase C.
Phase D started on day 92 after transferring the aerobic slurries 3 and 4 into
an anaerobic environment and consequently anaerobic utilization of nitrate by
denitrification.
P-addition day 11

20

P-addition day 64

EC [mS/cm]

16
12
8
4
0
0

20

EC +P11

40

Time [d] 60

EC +P64

80

100

EC abiotic

Figure 2. Electrical conductivity (T = 20) in [mS/cm] in aerobic slurries +P64, +P11 (biotic) and C1
(abiotic) versus time [days]. Phosphate addition occurred on day 11 (+P11) and on day 64 (+P64).
Error bars represent one standard deviation.

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BIODEGRADATION OF MEA, MEG AND TEG

C(N) [mmol/kg]

500

NH3

400

+P
NH4+

300

NO2-

200

NO3-

100

N2

0
0
NH4 T-N
NO2

20

40 Time [d]
NH4 S-N
NO3

60
MEA T-N

80

100
NH4 W-N

Figure 3. Nitrogen species concentrations: Total ammonium-nitrogen (NH4 Total) and ammonium
nitrogen in KOH extract (NH4 Soil), total MEA nitrogen (MEA Total), total nitrite nitrogen (NO2)
and total nitrate nitrogen (NO3) concentrations in [mmol-N/kg] averaged +P11 versus time [days].
Phases A, B, C and D indicate stages of biodegradation.

Overall ammonium concentrations decreased during the entire period of the experiment. Figure 3 presents ammonium concentrations sorbed onto the soil fraction
and total concentrations in +P11. Total ammonium levels rapidly decreased during
phase A and until day 15. A slower decrease was observed between day 15 and
day 58, as indicated in phase B. A sharp decline was observed during phase C
after day 58. The sorbed ammonium concentrations were constant until day 13 and
increased from 79 mmol-N/kg to 175 mmol-N/kg until day 23 during phase B of
the experiment. The MEA degraded in phase B from 102mmol-N/kg starting on
day 13 to below detection. A sharp decrease of ammonium levels was observed
in both soil and total concentrations after day 58. The total ammonium concentration in the two abiotic controls averaged 420 58 mmol-N/kg during the entire
study.
Average nitrite concentrations (+P11) increased on day 58 and reached an average maximum of 54 mmol-N/kg in phase C on day 72. Nitrate production occurred
on day 64 and reached an average maximum level of 29 mmol-N/kg on day 92. In
phase D and after day 92, slurries 3 and 4 were placed in a dry glove box under
anaerobic conditions. The addition of 200mmol/kg acetic acid supplied sufficient
available organic carbon to initiate anaerobic denitrification. The nitrate levels declined to below detection levels on day 98 in phase D of the experiment. Finally,
bioreactors 3 and 4 (+P11) indicated that MEA, MEG, TEG, acetic acid, ethanol,
ammonium, nitrate and nitrite concentrations were below detection on day 98 at
the end of the experiment. The pH values increased with declining acetic acid
and increasing ammonium. As ammonium concentrations declined, pH declined
as well (Figure 4). The abiotic controls indicated no significant pH changes during
the entire experiment.

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O. MRKLAS ET AL.

10.00

+P

+P

pH [-]

9.00
8.00
7.00
6.00
0

20

40

pH +P11

Time [d]

60

pH +P64

80

100

pH abiotic

Figure 4. Average aerobic pH values +P64, +P11 (biotic) and control (abiotic) versus time [days].
Error bars represent one standard deviation. Phases A, B, C and D indicate stages of biodegradation.
P-addition day 11

80000

C [mg/kg]

60000
40000
20000
0
0

MEA total

20

EtOH

40

Time [d]

MEA abiotic

60

NH4 total

80

100

TEG

Ac

Figure 5. Average anaerobic biotic concentrations and control (abiotic) of acetate, ethanol, MEG,
TEG and MEA [mg/kg] during biodegradation versus time [days]. Error bars represent one standard
deviation.

3.3. ANAEROBIC

BIOREACTORS

The four anaerobic slurries were monitored for 98 days (Figure 5). The anaerobic
bioreactors A1 and A2 showed similar degradation trends and results were averaged.
MEA, MEG and TEG degradation was monitored without phosphate addition until
day 11 and with P-addition after day 11. Table 1 presents the initial concentrations
of chemical species in the soil, groundwater and slurries.
The alkanolamine, glycols and degradation products were not completely degraded in the anaerobic bioreactors. The biotic bioreactors showed MEA and TEG
concentrations decreasing by 23 and 25% during the study. Ammonium concentrations were constant in both abiotic and biotic anaerobic slurries. MEG concentrations in the biotic anaerobic slurries were stable during the entire period of the

BIODEGRADATION OF MEA, MEG AND TEG

259

study and similar to MEG concentrations in the abiotic reactors. The biodegradation rates for the compounds were generally one magnitude lower than the aerobic
biodegradation rates (Tables 3 and 4).

4. Discussion
The presence of MEA, MEG, TEG and their breakdown products ethanol, acetic
acid and ammonium in the collected soil and groundwater samples agrees with
previous proposed degradation pathways of alkanolamines and glycols (Bradbeer,
1965; BUA, 1994; Jones and Turner, 1973; McVicker et al., 1997). In general,
MEA, MEG and TEG biodegradation results in ethanol, acetic acid and ammonium
evolution through deamination (MEA) and dehydrogenase (glycols) (Gottschalk,
1985).
The species present (MEA, ammonium) supplied adequate nitrogen concentrations and it was observed that the aerobic biodegradation increased by one order
of magnitude and was successfully completed after the addition of phosphate. The
aerobic bioreactors showed complete biodegradation of ethanol, acetic acid, MEG,
MEA and TEG within 21 days after phosphate addition. The 3000 mg/L MEA
concentrations found in this study degraded completely and did not inhibit aerobic biodegradation. These biodegradation rates agree with recent literature (BUA,
1994; Philip, 1991; Sorensen et al., 1997). Therefore, it was concluded that the
slurries were initially operated under phosphate-limited conditions during the first
phase of the experiment. These results suggest that aerobic field conditions with
adequate phosphate supplies are optimal for MEA, MEG and TEG biodegradation.
Significant changes in anaerobic biodegradation rates were not observed with the
addition of phosphate. The anaerobic biodegradation rates of ethanol, acetic acid,
MEA and TEG were constant even after phosphate addition during the entire period
of the experiment. Acetic acid, however, seemed to be the preferential substrate in
the anaerobic reactors, as the degradation rate constants were the highest. MEG
concentrations did not change during the experiment. Therefore, anaerobic degradation of ethanol, MEA, MEG and TEG may be inhibited by present acetic acid and
ammonium concentrations. In fact, it was previously shown that acetic acid concentrations of approximately 2000 mg/L significantly inhibited methane production in
anaerobic environments (Mawson et al., 1991). Furthermore, it was reported that
anaerobic methane production was inhibited due to high ammonium/ammonia concentrations (Gallert et al., 1998). Thus, anaerobic decay of ethanol, MEA and TEG
was much slower than aerobic degradation, possibly attributed to inhibition and
was not complete at the end of the 98 day experiment.
MEA has been described as an ammonium derivate with comparable properties (Bollmeier, 1991; Dow, 1980). Ammonium can be sorbed onto clay minerals
(Lumbanraja and Evangelou, 1990; Olsen et al., 1999). MEA distribution between
water and soil played a major role in recent laboratory and field investigations

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O. MRKLAS ET AL.

(Mrklas et al., 2001; Mrklas 2002; Sorensen et al.,1999). This study presents for
the first time the detailed distribution of both ammonium and MEA in water and
soil fractions in slurry reactors during enhanced aerobic and anaerobic biodegradation. Earlier studies determined MEA and ammonium distribution coefficients of
about 1 L/kg in the same soil samples employed here (Mrklas, 2002). These distribution coefficients suggested that sorption may have limited biodegradation. The
results presented here show that MEA biodegraded faster in the water phase than
MEA sorbed on the soil phase. Thus available or free MEA concentrations in the
water phase were limited by sorption during enhanced biodegradation. Bioavailability of substrate plays a major role in biodegradation of contaminants (Alexander,
1999). Cation exchange and/or sorption fix the substrate and may inhibit microbial
utilization of the sorbed substrate (Nielsen et al., 1996). Therefore, the distribution of substrate in a soil-water mixture determined the availability of substrate
in the water phase and, thus influences the bulk biodegradation rates. Such limitation occurred during the enhanced aerobic degradation of MEA, as MEA concentrations in the water phase (available) declined faster than in the soil phase
(sorbed).
In addition, it was demonstrated that a decrease of total MEA concentration due
to biodegradation resulted in increased ammonium sorbed onto the soil fraction
(Figure 3). The biodegradation of alkanolamines occurred partly under ammonium
evolution (Bradbeer, 1965; Jones and Turner, 1973). As MEA molecules desorbed
from the soil phase and partitioned into the water phase the produced ammonium
sorbed onto the available cation exchange sites. The overall ammonium concentration, however, declined as the pH increased to a maximum of 8.9 and ammonia
volatilization became a major release pathway. The pH increase was a direct result
of acetic acid biodegradation (Figures 1a and 4). Ammonium (NH+
4 ) and ammonia (NH3 ) partitioning is mainly influenced by pH changes (Canter, 1997; Larsen
et al., 2001). The ammonium equilibrium in the slurries consisted of (1) ammonium
ions in the water phase and (2) soil phase, (3) ammonia gas dissolved in the water
phase and (4) ammonia gas in the vapor phase. As the pH increased the portion of
dissolved ammonia gas (NH3 ) increased in relation to the dissolved ammonium and
consequently resulted in an increase of ammonia gas in the vapor phase escaping
the open bioreactors. At a pH of 8.9 about 30% of the ammonium concentration
is gaseous ammonia and it partitions into the atmosphere following Henrys law
(Larsen et al., 2001). A maximum pH of 8.9 was observed during the P-enhanced
phase of the experiment in reactors 3 and 4 and therefore, ammonia volatilization
appeared to be the major mechanism for ammonium loss. Ammonia release will
not be as large a factor in highly buffered field sites, but may play a major role
in areas with high pH values. In general, results demonstrated that it is necessary
to understand the roles of sorption of both MEA and ammonium during MEA
biodegradation.
Nitrification and denitrification were demonstrated using sequential aerobic and
anaerobic conditions in +P11 (Figure 3). The production of nitrite (NO
2 ) before

BIODEGRADATION OF MEA, MEG AND TEG

261

nitrate (NO
3 ) during the aerobic phase suggested the inhibition of Nitrobacter
species that are capable of nitrate production utilizing nitrite. Anthonisen et al.
(1976) demonstrated that dissolved ammonia and nitric acid concentrations inhibited and delayed nitrate evolution in slurry experiments. The formation of the
intermediate hydroxylamine occurs during nitrification and before the evolution
of nitrite (Brock and Madigan, 1991; Gottschalk, 1985). Consequently, we interpret the observed patterns of nitrite and nitrate evolution as being due to inhibition caused by dissolved ammonia and some loss of nitrogen to hydroxylamine
formation.
Both aerobic and anaerobic abiotic controls demonstrated that chemical transformation of MEA, MEG and TEG was insignificant.
5. Conclusions
This biodegradation study of MEA, MEG, TEG and their degradation products
employed natural soil, groundwater and indigenous microbes. Phosphate limitations
played a significant role in aerobic biodegradation at the concentrations observed
here. This suggests that impacted areas in field scenarios may show elevated levels
for long periods until aerobic conditions and phosphate addition occur.
The KOH method enabled complete MEA and ammonium extraction from the
soil fraction due to increased pH (MEA desorption) and excess potassium concentration (ammonium cation exchange). This suggests that MEA sorption may
not entirely be contributed to cation exchange. The implications for the field are
that groundwater sample analyses only determine a portion of the total ammonium
and MEA concentrations in the subsurface. The degradation patterns indicated that
carbon species degraded aerobically in the order ethanol, MEG, acetic acid, MEA
followed by TEG. Therefore, TEG will degrade after the other species in this series
of contaminants and may persist in the environment the longest. The significant
acetic acid concentrations seen here at neutral pH values were buffered by the
system. At sites with low buffering capacity increasing acetic acid concentrations
caused by biodegradation may require carbonate/bicarbonate addition in order to
create neutral pH levels and optimize biodegradation.
Nitrification and denitrification were observed by sequentially changing aerobic
to anaerobic conditions, supplementing with acetate and utilizing indigenous microorganisms. Shifting aerobic and anaerobic conditions in the field, i.e. seasonal
changes or site heterogeneity (saturated/unsaturated zones) may allow for natural
in-situ nitrification and denitrification.
Acknowledgement
This study was supported by the Natural Sciences and Engineering Research Council of Canada (NSERC) and Imperial Oil Resources.

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References
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