assessment of 17beta-estradiol removal from wastewater via abiotic and biotic routes and potential

172 Pages·2017·3.05 MB·English

Clemson University
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5-2013
ASSESSMENT OF 17BETA-ESTRADIOL
REMOVAL FROM WASTEWATER VIA
ABIOTIC AND BIOTIC ROUTES AND
POTENTIAL EFFECTS ON FOOD CHAIN
PATHWAYS
Lance Beecher
Clemson University, [email protected]
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Beecher, Lance, "ASSESSMENT OF 17BETA-ESTRADIOL REMOVAL FROM WASTEWATER VIA ABIOTIC AND BIOTIC
ROUTES AND POTENTIAL EFFECTS ON FOOD CHAIN PATHWAYS" (2013).All Dissertations. 1083.
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ASSESSMENT OF 17β-ESTRADIOL REMOVAL FROM
WASTEWATER VIA ABIOTIC AND BIOTIC ROUTES AND
POTENTIAL EFFECTS ON FOOD CHAIN PATHWAYS

A Dissertation
Presented to
the Graduate School of
Clemson University

In Partial Fulfillment
of the Requirements for the Degree
Doctor of Philosophy
Environmental Toxicology

by
Lance E. Beecher
May 2013

Accepted by:
Dr. Sarah A White, Committee Chair
Dr. Daniel Hitchcock
Dr. Stephen J. Klaine
Dr. Cindy M. Lee

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ABSTRACT

Estrone, 17β-estradiol (E2), and estriol are three of the most relevant natural
hormones and are often found to contaminate the environment. When emanating from
endogenous sources, these three compounds are not toxic or of concern; however, when
estrogens are released into the environment from wastewater effluent they become a
concern. The overall goal of the project was to evaluate abiotic and biotic techniques for
removal of E2 and to quantify survival and reproduction impacts on an algal species and
on an invertebrate species. We accomplished this goal by: 1) quantifying E2 sorption
rates to crushed brick, an industrial mineral aggregate (calcined clay) and sand media by
exposing each to increasing concentrations of E2; 2) determining the removal efficacy of
E2 by the three media from wastewater sources in a pilot scale fixed bed bioreactor; 3)
defining the capacity of algal cells to facilitate E2 removal whether via sorption or
metabolism-aided degradation processes; 4) establishing the impacts of increasing E2
concentrations on the density and growth of algal communities; 5) determining if food-
borne exposure was a critical exposure route to invertebrate species by feeding Daphnia
magna E2 inoculated algae; and 6) assessing the impact of aqueous E2 exposure on the
survival and reproduction of D. magna for two generations.
Sorption onto mineral media, activated sludge, and extended sludge retention time
are key processes influencing EDC degradation in wastewater treatment facilities.
Calcined clay sorbed significantly more E2 from simulated wastewater than either
crushed brick or sand. At concentrations less than 100 ng/L E2, the clay media sorbed
more than 60% of E2 from solution. When the three media were tested in a static bed
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biofilter to examine both sorption and biodegradation by bacterial films established in the
biofilter, nearly all the E2 was removed from solution after 24 hours in the fixed-bed
bioreactor filled with clay. Wastewater collected directly from the activated sludge unit
and the final settling stage were spiked with E2; the concentration E2 in solution was
reduced to below our detection limits (nearly 0 ng/L) after as little as four hours
(activated sludge) to 24 hours (final settling stage), with no media added to provide
surface area for microbial colonization.
Increasing densities of algal (Pseudokirchneriella subcapitata) populations
enhanced removal of E2 from wastewater over a 24-hour period. At algal densities > 24
million cells/mL more than 95% of E2 was removed from solution. When algal
populations of the same density (12 million cells/mL) were exposed to increasing
concentrations of E2 over a 24-hour period, between 85 and 100% of aqueous E2 was
removed at 55, 500 and 1000 ng/L exposure concentrations. Removal of E2 by algae
exposed to higher concentrations (3000 and 10000 ng/L) of E2 averaged ≤ 90%, and both
exposure concentrations were significantly different (P = 0.05) from lower concentrations
(55, 500 and 1000 ng/L). Results suggest that sorption to organic material and
photodegradation are not the main mechanisms driving E2 removal from aqueous
solution, but rather that biological processes (metabolism and degradation) are the
primary removal mechanisms in these lab-scale algal treatment systems.
Finally, invertebrate organisms offer a unique opportunity to evaluate endocrine
disruption activity in lower food level organisms. Daphnia magna were utilized as a
model organism to characterize the effects of E2 on food chain micro-fauna. Endpoints
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evaluated were the survival and reproduction of D. magna after exposure to E2. Survival
and reproduction were recorded for both the first and second generation of D. magna.
During the second generation D. magna experiments, neonates from the first generation
experiments were collected and divided into a group subjected to continued E2 exposure
and a group where E2 was removed and recovery/depuration permitted. Results for food-
borne exposure expressed E2 influence on D. magna populations via aqueous exposure
(bioconcentration) and not by way of food-borne exposure (bioaccumulation). The three
aqueous exposure results emphasized generational effects of E2 exposure to D. magna,
indicating potential generational effects were possible with carryover impacts magnified
in second generation organisms, if exposure to E2 was not removed. When E2 exposure
ceased, D. magna recovered almost immediately and reproduced at levels similar to
control organisms. Results from these experiments will give some insight into treatment
processes to aid in reduction or removal of E2. Reducing E2 levels entering the natural
environment can be beneficial in maintaining a healthy balanced ecosystem.

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ACKNOWLEDGEMENTS

I would like to thank my major advisor Sarah White and my other committee
members Cindy Lee, Steve Klaine and Dan Hitchcock for their guidance and support
during my graduate experience. I would like to thank Basma Demiri who provided field
and lab assistance, Billy Bridges for his recommendations for statistical analysis and
Melissa Riley and Sandra Gray who provided exceptional support and guidance
throughout the many pathways of chemical analysis undergone throughout the project.
I would like to thank my mom, Barbara Goff, for her love and support throughout
my career. Most importantly, I would like to thank my wife, Paula Beecher, and my two
daughters, Kayleigh and Caroline Beecher, for their limitless patience and love they have
provided me during this all-encompassing process. They have cheered me throughout
hard times and given me the encouragement to follow my dream. I dedicate this degree
to my two daughters and hope this gives them the belief that if you work hard your
dreams will come true.

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TABLE OF CONTENTS
Page
TITLE PAGE ...................................................................................................................... i
ABSTRACT ........................................................................................................................ ii
ACKNOWLEGEMNTS ......................................................................................................v
LIST OF TABLES ........................................................................................................... viii
LIST OF FIGURES .............................................................................................................x
PREFACE ...................................................................................................................... xviii
CHAPTER 1: LITERATURE REVIEW .............................................................................1
Endocrine disrupting compounds and health effects ................................................................... 1
Structure and properties of natural estrogenic compounds ......................................................... 2
Commercial Use of Estrogens ..................................................................................................... 4
Sources and Fate in the Environment .......................................................................................... 5
Wastewater Treatment ............................................................................................................... 12
Alternative Treatments .............................................................................................................. 15
Radioimmunoassay Analysis..................................................................................................... 19
References ................................................................................................................................. 20
Chapter 2: MEDIA CHOICE, SORPTION AND BIODEGRADATION
OF 17β-ESTRADIOL IN STATIC BED BIOFILTERS ...........................................29
Abstract ..................................................................................................................................... 29
Introduction ............................................................................................................................... 30
Materials and Methods .............................................................................................................. 35
Data Analysis ............................................................................................................................ 48
Results and Discussion .............................................................................................................. 50
Conclusions ............................................................................................................................... 64
Acknowledgements ................................................................................................................... 64
References ................................................................................................................................. 66
Appendix A ............................................................................................................................... 72
Appendix B................................................................................................................................ 74
Chapter 3: ALGAE-AIDED (Pseudokirchneriella subcapitata)
REMOVAL OF 17β-ESTRADIOL FROM WASTEWATER ..................................75
Abstract ..................................................................................................................................... 75
Introduction ............................................................................................................................... 76
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Table of Contents (Continued)
Page
Materials and Methods .............................................................................................................. 80
Results and Discussion .............................................................................................................. 88
Conclusions ............................................................................................................................. 104
Acknowledgements ................................................................................................................. 105
References ............................................................................................................................... 106
Chapter 4: EFFECTS OF A HUMAN DERIVED HORMONE ON
MULTIGENERATIONAL SURVIVAL AND FECUNDITY OF
WATER FLEA, Daphnia magna .............................................................................110
Abstract ................................................................................................................................... 110
Introduction ............................................................................................................................. 111
Material and Methods: ............................................................................................................. 114
Results and Discussion ............................................................................................................ 119
Conclusions ............................................................................................................................. 141
Acknowledgements ................................................................................................................. 143
References ............................................................................................................................... 144
Chapter 5: OVERALL CONCLUSIONS AND FUTURE
RECOMMENDATIONS .........................................................................................148
Conclusions ............................................................................................................................. 148
Future Recommendation ......................................................................................................... 149

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LIST OF TABLES
Table Page
1-1. Physical-chemical properties of estrone, 17β-estradiol, estriol. ..................................4
1-2. Typical concentration of estrogenic compounds1 produced by human
females. .............................................................................................................5
1-3. Estimated yearly steroid hormone excretion (tons/yr)1 by farm
animals in the United States. ............................................................................6
1-4. Estrogens in milk and milk products in pg/ml or pg/g1. ..............................................8
2-1. Constituents1 used in the makeup of synthetic wastewater used as
substrates in biodegradation experiments comparing calcined
clay, crushed brick and sand. ..........................................................................42
2-2. Quality assurance/quality control evaluation of radioimmunoassay kit
analyzing a serum based analysis with water samples. Serum
based standards were diluted with DI water and processed.
Mean values and percent coefficient of variation are
presented. ........................................................................................................49
2-3. Freundlich and Langmuir coefficients derived from sorption
isotherms for live algae under light and dark conditions along
with live and dry algae. E2ad, average 17β-estradiol (E2)
adsorbed to substrate; b, slope; K, Freundlich sorption
coefficient; Smax, sorption maxima; k, binding strength. .............................52
3-1. Chemical makeup of nutrient water used to culture
Pseudokirchneriella subcapitata. Nutrient water was prepared
by mixing 3 mL of each solution to 1 liter of DI water and
autoclaved for sterilization. ............................................................................82
3-2. Freundlich and Langmuir coefficients derived from sorption
isotherms for live algae under light and dark conditions along
with live and dead algae. E2 , average E2 adsorbed to
ad
substrate (standard deviation); b, slope; K, Freundlich sorption
coefficient; S , sorption maxima; k, binding strength. ................................98
max
4-1. Moderately hard water components used to produce water for
culturing and experimental medium for testing survival and
reproduction of Daphnia magna. .................................................................116
viii
List of Tables (Continued)
Table Page
4-2. Chronic LC 1and LOEC2 (21 day) values for Run 1, 2 and 3
50
(Generation 1 and 2) and combined runs of D. magna
experiments. The standard deviation is provided for LC in
50
the parenthesis. .............................................................................................127

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