17β-Estradiol mineralization under field and laboratory incubations
ANNEMIEKE FARENHORST
*
, INOKA AMARAKOON, and LINDSEY ANDRONAK
Department of Soil Science, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada
!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!
*
Address correspondence to Dr. Annemieke Farenhorst, Department of Soil Science, Faculty of
Agricultural and Food Sciences,University of Manitoba, 380 Ellis Building, Winnipeg,
Manitoba, R3T 2N2, Canda; Tel: (204) 474 6858; FAX: (204) 474 7642
E-mail: annemieke.farenhorst@umanitoba.ca
ABSTRACT 1!
2!
3!
Mineralization studies of natural steroid hormones (e.g., 17β-estradiol, E2) are performed in 4!
environmental incubators, usually under a constant temperature such as 20°C. In this paper, we 5!
present a microcosm protocol that quantified the mineralization of E2 in soils under field 6!
temperatures. The nine agricultural soils tested had a wide range of soil organic carbon (1.1 to 7!
5.2%) and clay (9 to 57%) contents. The calculated time over which half of the applied E2 was 8!
mineralized (E2-½) ranged from 299 to 910 d, and total E2 mineralization at 48 d (E2-TOT48) 9!
ranged from 4 to 13%. In subsequent laboratory incubations, the same soils were incubated under 10!
a constant temperature of 20°C, as well as under cyclic temperatures of 14.5°C (14 h) and 11.5°C 11!
(10h), which was within the temperature extremes observed in the field microcosms. E2-½ 12!
ranged from 157 to 686 d at 20°C and from 103 to 608 d at the cyclic temperatures, with the E2-13!
TOT48 ranging from 6 to 21% at 20°C and from 7 to 30% under cyclic temperatures. Despite the 14!
overall 6.75 °C lower mean temperatures under the cyclic versus constant temperatures, E2 15!
mineralization was stimulated by the temperature cycles in three soils. Regardless of the 16!
incubation, the same loamy sand soil always showed larger E2 mineralization than the other 17!
eight soils and this loamy sand soil also had the smallest E2 sorption. Current modeling 18!
approaches do not take into consideration the effects of temperature fluctuations in the field 19!
because the input parameters used to describe degradation are derived from laboratory 20!
incubations at a constant temperature. Across the eight soils, E2-½ was on average 1.7 times 21!
larger and E2-TOT48 was on average 0.8 times smaller under field temperatures than under a 22!
constant 20°C. Hence, we conclude that incubations at 20°C give a reasonable representation of 23!
!
3!
E2 mineralization occurring under field conditions to be expected in a typical Prairie summer 24!
season. 25!
!26!
27!
Keywords: 17β-estradiol, mineralization, temperature, fluctuations, laboratory, field 28!
29!
30!
INTRODUCTION 31!
32!
33!
Vertebrates excrete natural steroid hormones such as 17β-estradiol (E2) and when livestock 34!
manure or sewage sludge is applied to agricultural land, elevated concentrations of steroid 35!
hormone residues are detected in soil.
[1,2]
E2 can be transported from soils into the broader 36!
environment by processes such as surface runoff
[3,4]
and leaching.
[5,6]
The reported half-life of 37!
E2 in agricultural soils is 0.2 to 9.7 days.
[7-9]
A large portion of the initially applied E2 becomes 38!
soil-bound (non-extractable). For example as much as 56% in a silt loam, 70% in a sandy loam 39!
and 91% in a loam soil at only three days after E2 was applied.
[10]
However, the soil-bound 40!
fraction is slowly mineralized.
[10]
The smaller extractable fraction is a mixture of E2 and estrone 41!
(E1), with the ratio E2:E1 decreasing over time. E2 biodegradation to E1 has been observed in 42!
many studies, with E1 being the main or only transformation product of E2 in soils.
[10-12]
E1 can 43!
convert back to E2 but this has been observed only under anaerobic conditions.
[13]
Only a 44!
limited number of bacteria can degrade both E2 and E1,
[14]
and hence E2 and E1 have different 45!
degradation rates in soil.
[10]
46!
47!
!
4!
Mineralization can only be quantified by using radiolabeled [4-
14
C] E2 in microcosm 48!
experiments,
[10,11,15,16]
whereby the recovery of
14
CO
2
indicates that the steroid molecule has 49!
been inactivated because of ring cleavage.
[17]
The time that 50% of the applied E2 is 50!
mineralized (E2-½) can be calculated from these laboratory data. E2-½ has ranged from 294 to 51!
418 d in sewage sludge, from 721 to 869 d in soil and from 2,258 to 14,146 d in biosolids.
[18]
52!
The maximum E2 mineralization (E2-TOT48) is typically less than 20% of the initial E2 applied 53!
to soil microcosms, even for incubations up to 90 days.
[10,19]
54!
55!
Temperature can affect degradation/mineralization through its influence on soil microbial 56!
activity and populations.
[20,21]
Mineralization studies are typically conducted in environmental 57!
chambers at a constant temperature.
[7,16]
Incubation studies using a range of constant 58!
temperatures have shown that E2 persistence in soil decreases with increasing temperatures,
[9,10]
59!
with total E2 mineralization over a 61 day period increasing from 2% at 4
o
C to 14% at 30
o
C.
[10]
60!
Temperature varies under field conditions and, in this paper, we present a microcosm protocol 61!
that can quantify E2 mineralization in the field. The objective of this study was to implement the 62!
in-field microcosm experiment to determine E2 parameters (E2-½, E2-TOT48) in a wide range 63!
of soils and compare these values to E2 parameters derived in the same soils under constant and 64!
cyclic temperatures in a subsequent laboratory incubation. 65!
66!
67!
MATERIALS AND METHODS 68!
69!
70!
!
5!
Chemicals 71!
72!
73!
Analytical-grade 17β-estradiol (98% chemical purity; Sigma Aldrich Chemical Company, St. 74!
Louis, MO) and 4-
14
C labeled 17β-estradiol (99% radiochemical purity; specific activity 45 75!
mCi/mmol; American Radiolabeled Chemicals Incorporated, St. Louis, MO) was used. 76!
77!
Soil Properties 78!
79!
Soil samples (0-10 cm) were collected from the Ap-horizons in a range of agricultural fields in 80!
the Province of Manitoba, Canada. Fields were selected based on soil maps to ensure that the 81!
soils selected had a range of soil textures and organic carbon contents. Soils were sieved (< 2 82!
mm) and frozen at -25 ±2°C. Prior to the microcosm experiments, the moisture content of 83!
thawed soil was determined gravimetrically and distilled water was added to bring the soil to 84!
80% of field capacity minus the liquid volume needed to add the chemical solutions. Field 85!
capacity was determined using laboratory leaching columns (11 cm in height, 2.7 cm radius) and 86!
defined as the amount of gravity retained soil moisture in these columns at 96 hours after 87!
saturation. Portions of soil were also air-dried and analyzed for a range of properties (Table 1). 88!
89!
