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An Investigation of Stannous

Chloride as an Inhibitor of
Lead Corrosion

Subject Area: Infrastructure


An Investigation of Stannous
Chloride as an Inhibitor of
Lead Corrosion

©2010 Water Research Foundation. ALL RIGHTS RESERVED


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An Investigation of Stannous
Chloride as an Inhibitor of
Lead Corrosion

Prepared by:
Raymond M. Hozalski, Ying Tan, and Xiaojun Dai
University of Minnesota, Department of Civil Engineering, Minneapolis, MN 55455-0116

Sponsored by:
Water Research Foundation
6666 West Quincy Avenue, Denver, CO 80235-3098

Published by:

©2010 Water Research Foundation. ALL RIGHTS RESERVED


DISCLAIMER

This study was funded by the Water Research Foundation (Foundation). The Foundation
assumes no responsibility for the content of the research study reported in this publication
or for the opinions or statements of fact expressed in the report. The mention of trade names
for commercial products does not represent or imply the approval or endorsement of the
Foundation. This report is presented solely for informational purposes.

Copyright © 2010
by Water Research Foundation

ALL RIGHTS RESERVED.


No part of this publication may be copied, reproduced
or otherwise utilized without permission.

ISBN 978-1-60573-121-6

Printed in the U.S.A.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


CONTENTS

LIST OF TABLES������������������������������������������������������������������������������������������������������������������������ ix

LIST OF FIGURES���������������������������������������������������������������������������������������������������������������������� xi

FOREWORD����������������������������������������������������������������������������������������������������������������������������� xvii

ACKNOWLEDGMENTS���������������������������������������������������������������������������������������������������������� xix

EXECUTIVE SUMMARY��������������������������������������������������������������������������������������������������������� xxi

CHAPTER 1: INTRODUCTION��������������������������������������������������������������������������������������������������� 1

CHAPTER 2: LITERATURE REVIEW���������������������������������������������������������������������������������������� 3


Lead Corrosion......................................................................................................................3
Effect of Biocides on Corrosion����������������������������������������������������������������������������� 3
Microbially Influenced Corrosion ........................................................................................4
Biofilm�������������������������������������������������������������������������������������������������������������������� 5
Sulfate-Reducing Bacteria�������������������������������������������������������������������������������������� 6
Ammonia-Oxidizing Bacteria��������������������������������������������������������������������������������� 7
Controlling MIC ���������������������������������������������������������������������������������������������������� 7
Corrosion Inhibitors..............................................................................................................8
Orthophosphate������������������������������������������������������������������������������������������������������� 8
Silicate��������������������������������������������������������������������������������������������������������������������� 8
Stannous Chloride��������������������������������������������������������������������������������������������������� 8
Preliminary Stannous Chloride Research in OUR Laboratory.............................................9
Pipe Loop Study������������������������������������������������������������������������������������������������������ 9
Preliminary Bench-Scale Experiments����������������������������������������������������������������� 11

CHAPTER 3: MATERIALS AND METHODS��������������������������������������������������������������������������� 15


Introduction.........................................................................................................................15
Investigation of Tin Chemistry...........................................................................................15
Kinetics of Monochloramine Consumption by Sn2+ �������������������������������������������� 15
Effect of pH on Monochloramine Stability in the Presence of Sn2+ ��������������������� 15
Oxidation of Sn+2 With Monochloramine������������������������������������������������������������� 15
Effect of Tin on Inactivation of Planktonic Bacteria and Biofilms.....................................16
Microorganisms���������������������������������������������������������������������������������������������������� 16
Batch Growth Curves�������������������������������������������������������������������������������������������� 17
Bacterial Activity Assay���������������������������������������������������������������������������������������� 17
Bacterial Inactivation Experiments����������������������������������������������������������������������� 18
Undefined Mixed Cultures������������������������������������������������������������������������������������ 19
Environmental Isolate������������������������������������������������������������������������������������������� 19

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vi | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Continuous-Flow Rotating Disk Reactor�������������������������������������������������������������� 19


Continuous-Flow Glass Capillary Biofilm Reactor���������������������������������������������� 21
Effects of Tin on Lead Corrosion........................................................................................22
Lead Coupon Aging���������������������������������������������������������������������������������������������� 22
Preliminary Corrosion Experiment With New Lead Coupons����������������������������� 23
Lead Corrosion Experiment With Aged and New Lead Coupons������������������������ 24
Analytical Methods.............................................................................................................24
Tin������������������������������������������������������������������������������������������������������������������������� 24
Lead����������������������������������������������������������������������������������������������������������������������� 25
Chlorine����������������������������������������������������������������������������������������������������������������� 25
Lead Surface Mineralogy�������������������������������������������������������������������������������������� 26
Lead Surface Morphology������������������������������������������������������������������������������������ 26

CHAPTER 4: TIN CHEMISTRY������������������������������������������������������������������������������������������������� 27


Introduction.........................................................................................................................27
Results and Discussion.......................................................................................................27
Monochloramine Consumption by Sn2+ ��������������������������������������������������������������� 27
Oxidation of Sn+2 by Monochloramine����������������������������������������������������������������� 27
Kinetics of Monochloramine Consumption���������������������������������������������������������� 31
Summary.............................................................................................................................31

CHAPTER 5: TOXICITY OF TIN TO PLANKTONIC AND BIOFILM BACTERIA��������������� 33


Introduction.........................................................................................................................33
Results and Discussion.......................................................................................................33
Effect of Sn on Bacterial Growth�������������������������������������������������������������������������� 33
Effect of Sn on OUR��������������������������������������������������������������������������������������������� 35
Effect of Sn on Bacterial Inactivation ������������������������������������������������������������������ 35
Effect of Sn on Inactivation of Undefined Mixed Cultures From
the Mississippi River�������������������������������������������������������������������������������������� 41
Effect of Sn on Inactivation of an Isolate From the Mississippi River����������������� 43
Preliminary Biofilm Experiment With Continuous-Flow Rotating
Disk Reactor���������������������������������������������������������������������������������������������������� 44
Continuous-Flow Glass Capillary Biofilm Reactor���������������������������������������������� 46
Summary.............................................................................................................................48

CHAPTER 6: EFFECT OF STANNOUS CHLORIDE ON LEAD CORROSION��������������������� 53


Introduction.........................................................................................................................53
Results and Discussion.......................................................................................................53
Potential Measurements During Coupon Aging��������������������������������������������������� 53
Mineralogy of the Aged Lead Coupons���������������������������������������������������������������� 55
Preliminary Lead Corrosion Experiment With New Lead Coupons�������������������� 59
Lead Corrosion Experiment With Aged and New Lead Coupons������������������������ 61
Summary.............................................................................................................................65

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Contents | vii

CHAPTER 7: CONCLUSIONS AND RECOMMENDATIONS������������������������������������������������ 67


Conclusions.........................................................................................................................67
Recommendations...............................................................................................................68

APPENDIX A: METHOD DEVELOPMENT����������������������������������������������������������������������������� 69

APPENDIX B: PHOTOGRAPHS OF PSEUDOMONAS AERUGINOSA PAO1 BIOFILM


GROWING IN CAPILLARY FLOW CELL REACTORS���������������������������������������������������� 71

APPENDIX C: X-RAY DIFFRACTION SPECTRA OF LEAD COUPONS������������������������������ 79

APPENDIX D: SCANNING ELECTRON MICROGRAPHS OF LEAD COUPONS��������������� 85

REFERENCES��������������������������������������������������������������������������������������������������������������������������� 107

ABBREVIATIONS��������������������������������������������������������������������������������������������������������������������� 111

©2010 Water Research Foundation. ALL RIGHTS RESERVED


©2010 Water Research Foundation. ALL RIGHTS RESERVED
TABLES

3.1 Water matrices used for lead coupon aging....................................................................... 23

3.2 Experimental matrix for lead corrosion experiments......................................................... 24

5.1 Effect of SnCl2 on OUR of four bacterial cultures............................................................ 37

5.2 Viable cell counts of P. aeruginosa PAO1 biofilm developed from continuous
rotating disk reactor experiment with and without SnCl2. ................................................ 47

5.3 Photographs of glass capillary biofilm reactors showing the effect of SnCl2
on the accumulation of P. aeruginosa PAO1 biofilm......................................................... 48

5.4 Photographs of glass capillary biofilm reactors showing the effect of SnCl2
on the removal of an established P. aeruginosa PAO1 biofilm.......................................... 51

6.1 Approximate mineralogical composition on the surface of lead coupons after


70 weeks of aging in lab or tap water as determined by XRD analysis............................. 57

B.1 Control reactor (no Sn added to medium).......................................................................... 72

B.2 0.125 mg/L of Sn added to medium................................................................................... 74

B.3 0.5 mg/L of Sn added to medium....................................................................................... 75

B.4 Removal of established biofilm with 1 mg/L of Sn+2 ........................................................ 78

ix

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©2010 Water Research Foundation. ALL RIGHTS RESERVED
FIGURES

2.1 Diagram showing the process of lead pipe corrosion.........................................................  4

2.2 Schematic diagram illustrating the three stages of biofilm formation................................  5

2.3 Diagram showing the solubility of Sn(II) in equilibrium with Sn(OH)2 (s) at 25°C .........  9

2.4 Comparison of mean lead concentrations for “problem” sampling sites in


the SPRWS system prior to (untreated) and during SnCl2 addition..................................  10

2.5 Sn2+ oxidation by free chlorine (initial concentration of ~10 mg/L as Cl2)
at pH 2.63, Temp. 22°C....................................................................................................  12

2.6 Effect of Sn2+ on the growth of P. aeruginosa PAO1 on 10% LB broth


in batch culture..................................................................................................................  12

3.1 Photograph of the continuous-flow rotating disk reactor system......................................  20

3.2 Photograph of a round glass disk mounted on to the bottom of the spindle of
the modified drill press in the continuous-flow rotating disk reactor system...................  20

3.3 Glass capillary biofilm reactor system..............................................................................  21

3.4 Photograph of the capillary flow cell system for testing the effects of tin on biofilm
development and removal.................................................................................................  22

3.5 Photograph of the flow cell holder (A; BioSurface Technology®) showing
two glass capillary flow cells connected to flexible tubing...............................................  22

3.6 Calibration curve from the analysis of Sn+2 via the TDT colorimetric method................  25

4.1 Oxidation of Sn+2 with monochloramine at pH 2.............................................................  28

4.2 Oxidation of Sn+2 with monochloramine at pH 6.............................................................  28

4.3 Oxidation of Sn+2 with monochloramine at pH 8.............................................................  29

4.4 Oxidation of Sn+2 with monochloramine at pH 9.5..........................................................  29

4.5 Effect of monochloramine dose on the oxidation of Sn+2 at pH 8....................................  30

4.6 Effect of temperature on the reaction rate of monochloramine with Sn+2........................  31

xi

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xii | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

5.1 Effect of SnCl2 on the growth of P. aeruginosa PAO1 (A) and N. europaea (B). P.
aeruginosa PAO1 and N. europaea were cultured in 10% LB broth and ATCC medium
2265, respectively, and initial cell densities were approximately 4.8 × 10–3 mg/L and
0.25 mg/L, respectively.....................................................................................................  34

5.2 Effect of SnCl2 on the OUR of P. aeruginosa PAO1 (24 hr, 80.7 mg biomass/L)
(A), P. aeruginosa PAO1 (72 hr, 36.7 mg biomass/L) (B), E. coli K-12 (24 hr,
102 mg biomass/L) (C), E. coli K-12 (72 hr, 192 mg biomass/L) (D), N. europaea
(14 days, 56.4 mg biomass/L) (E), and an undefined mixed nitrifying culture
(52.7 mg biomass/L) (F)...................................................................................................  36

5.3 Effect of Sn dose on the inactivation of P. aeruginosa PAO1 (72 hour culture age
except graph C, 24 hours) over a range of initial biomass concentrations (in CFU/mL):
3.69 × 104 (A), 5.23 × 104 (B), 1.6 × 105 (C), 1.16 × 107 (D), and 1.05 × 105 (E). All
graphs for experiments performed in lab water except graph E which was performed in
tap water............................................................................................................................  38

5.4 Effect of Sn dose on the inactivation of E. coli K-12 (24 hour: A,B; 72 hour:
C–E) over a range of initial biomass concentrations (in CFU/mL): 9.2 × 104 (A),
1.63 × 107 (B), 6.7 × 104 (C), 3.2 × 107 (D), and 1.26 × 105 (E).......................................  39

5.5 Inactivation of P. aeruginosa PAO1 as a function of Sn dose in µg Sn/CFU...................  40

5.6 Inactivation of E. coli K-12 as a function of Sn dose in µg Sn/CFU................................  40

5.7 Effect of SnCl2 concentration on the inactivation of planktonic Mississippi River


water bacteria....................................................................................................................  42

5.8 Inactivation of planktonic Mississippi River water bacteria as a function of


SnCl2 dose in µg Sn/CFU.................................................................................................  42

5.9 Inactivation of planktonic Mississippi River water bacteria as a function of


SnCl2 or SnCl4 dose in µg Sn/CFU...................................................................................  43

5.10 Effect of stannous chloride concentration on the inactivation of planktonic


Acinetobacter sp. at initial biomass densities of ~103 CFU/mL and ~102 CFU/mL........  44

5.11 Effect of stannic chloride concentration on the inactivation of planktonic


Acinetobacter sp. at initial biomass densities of ~103 CFU/mL and ~102 CFU/mL.........  45

5.12 Inactivation of planktonic Acinetobacter sp. as a function of SnCl2 or SnCl4


dose in µg Sn/CFU............................................................................................................  45

5.13 Photograph of P. aeruginosa PAO1 biofilm grown on rotating glass disks in media
without SnCl2 and with SnCl2. .........................................................................................  46

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Figures | xiii

5.15 A time series of brightness histograms from a capillary flow reactor showing the
development of a P. aeruginosa PAO1 biofilm grown in the absence of
stannous chloride..............................................................................................................  49

5.16 A time series of gray scale histograms derived from the brightness histograms
in Figure 5.15....................................................................................................................  49

5.17 Effect of stannous chloride on the accumulation of P. aeruginosa PAO1


biofilms in capillary flow cells..........................................................................................  50

5.18 Comparison of biofilm accumulation in capillary flow cells when the feed is
switched to media containing stannous chloride (1 mg/L as Sn, n = 3)
after 48 hours versus a control (no Sn, n = 2)...................................................................  51

6.1 Water temperature during lead coupon aging...................................................................  54

6.2 pH as a function of time for the reactor containing lab water amended with
1 mg/L as Cl2 free chlorine and 0.125 mg/L Sn+2.............................................................  54

6.3 Electrical potential vs. SCE for lead coupons incubated in lab water under
the following conditions: Control (A), 0.125 mg/L Sn2+ (B), 1 mg/L as Cl2
free chlorine (C), 1 mg/L as Cl2 free chlorine and 0.125 mg/L Sn2+ (D)..........................  55

6.4 Mean potential measurements from weeks 12 to 16 (n = 5) for lead coupons


aged in lab water (A) and tap water (B)............................................................................  56

6.5 Photos of lead coupons aged for approximately 70 weeks in lab or SPRWS
tap water............................................................................................................................  57

6.6 Examples of spectra from the XRD analysis of a new lead coupon (A) and a
lead coupon aged in lab water with free chlorine (1 mg/L as Cl2) and SnCl2
(0.125 mg/L Sn) (B)..........................................................................................................  58

6.7 SEM images of the surface of a lead coupon aged in lab water with free
chlorine (1 mg/L as Cl2) and SnCl2 (0.125 mg/L Sn).......................................................  59

6.8 SEM image of the surface of a new lead coupon..............................................................  60

6.9 Mean concentration of dissolved Pb (n = 2) versus time in the preliminary


lead corrosion experiment.................................................................................................  61

6.10 Concentration of dissolved lead versus time for the lead corrosion experiment with aged
(A-D) and new (E) lead coupons......................................................................................  62

6.11 Concentration of total lead versus time for the lead corrosion experiment
with aged (A-D) and new (E) lead coupons......................................................................  63

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xiv | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

6.12 SEM image of biofilm on the surface of a lead coupon from Reactor 5
(Sn treatment)....................................................................................................................  64

6.13 SEM image of biofilm on the surface of a lead coupon from Reactor 15 (Control).........  64

A.1 Effect of Sn+2 at concentrations 0.1, 0.5, 5, and 10 mg/L on the kinetics of color
development at a constant toluene-3,4-dithiol concentration of 20 mg/L........................  70

C.1 Spectrum of new lead coupon...........................................................................................  80

C.2 Spectrum of lead coupon aged in lab water only..............................................................  80

C.3 Spectrum of lead coupon aged in tap water only..............................................................  81

C.4 Spectrum of coupon aged in lab water with free chlorine (1 mg/L as Cl2) and
SnCl2 (0.125 mg/L Sn)......................................................................................................  81

C.5 Spectrum of lead coupon aged in lab water with SnCl2 (0.125 mg/L as Sn)....................  82

C.6 Spectrum of lead coupon aged in lab water with free chlorine (1 mg/L as Cl2)...............  82

C.7 Spectrum of lead coupon aged in lab water with monochloramine (1 mg/L as Cl2)........  83

C.8 Spectrum of lead coupon aged in tap water with monochloramine (1 mg/L as Cl2) and
SnCl2 (0.125 mg/L as Sn).................................................................................................  83

D.1 New lead coupon (i.e., reference) ....................................................................................  86

D.2 Coupon A (aged in tap water with Sn+2 with free chlorine) from Reactor 1 (Sn+2 = 0.125
mg/L).................................................................................................................................  87

D.3 Coupon A (aged in tap water with Sn+2) from Reactor 2 (Sn+2 = 0.125 mg/L) ...............  88

D.4 Coupon B (aged in tap water with Sn+2) from Reactor 3 (Sn+2 = 0.125 mg/L) ...............  89

D.5 Coupon B (aged in tap water with monochloramine) from Reactor 4


(Sn+2 = 0.125 mg/L)..........................................................................................................  90

D.6 Coupon C (new coupon) from Reactor 5 (Sn+2 = 0.125 mg/L)........................................  91

D.7 Coupon C (new coupon) from Reactor 6 (Sn+2 = 0.125 mg/L)........................................  92

D.8 Coupon A (aged in tap water with Sn+2 and free chlorine) from Reactor 7 (No Sn) .......  93

D.9 Coupon A (aged in tap water with Sn+2) from Reactor 8 (No Sn) ...................................  94

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Figures | xv

D.10 Coupon B (aged in tap water with Sn+2) from Reactor 9 (No Sn) ...................................  95

D.11 Coupon B (aged in tap water with monochloramine) from Reactor 10 (No Sn)..............  96

D.12 Coupon C (new coupon) from Reactor 11 (No Sn)..........................................................  97

D.13 Coupon C (new coupon) from Reactor 12 (No Sn) .........................................................  98

D.14 Coupon A (aged in tap water with Sn+2 and free chlorine) from Reactor 13 (Control)....  99

D.15 Coupon A (aged with tap water with Sn+2) from Reactor 14 (Control) .........................  100

D.16 Coupon B (aged with tap water with Sn+2) from Reactor 15 (Control)..........................  101

D.17 Coupon B (aged with tap water with monochloramine) from Reactor 16 (Control)......  102

D.18 Coupon C (new coupon) from Reactor 17 (Control)......................................................  103

D.19 Coupon C (new coupon) from Reactor 18 (Control)......................................................  104

D.20 Lead coupon aged in lab water with free chlorine and Sn+2 ...........................................  105

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©2010 Water Research Foundation. ALL RIGHTS RESERVED
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fair method for funding research in the public interest.
A broad spectrum of water supply issues is addressed by the Foundation’s research agenda:
resources, treatment and operations, distribution and storage, water quality and analysis, toxicol-
ogy, economics, and management. The ultimate purpose of the coordinated effort is to assist water
suppliers to provide the highest possible quality of water economically and reliably. The true ben-
efits are realized when the results are implemented at the utility level. The Foundation’s trustees
are pleased to offer this publication as a contribution toward that end.

David E. Rager Robert C. Renner, P.E.


Chair, Board of Trustees Executive Director
Water Research Foundation Water Research Foundation

xvii

©2010 Water Research Foundation. ALL RIGHTS RESERVED


©2010 Water Research Foundation. ALL RIGHTS RESERVED
ACKNOWLEDGMENTS

The authors thank the project advisory committee members (Michael Schock, Mike
Hotaling, and Carol Rego) and project officer (Traci Case) for their advice and support through-
out the project. Finally, the authors thank St. Paul Regional Water Services for providing water
samples and data for use in the project. The authors also than Anne Camper and staff at the Center
for Biofilm Engineering for providing training and advice in the use of capillary flow cell reactors.

xix

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©2010 Water Research Foundation. ALL RIGHTS RESERVED
EXECUTIVE SUMMARY

INTRODUCTION

Due to the concerns regarding high lead exposure via drinking water, the Lead and Copper
Rule (LCR) was established by the US Environmental Protection Agency in 1991. The LCR estab-
lished an action level (AL) of 15 µg/L as total Pb. In response to the LCR, many water treatment
plants across the U.S. have implemented corrosion prevention plans to decrease lead release into
drinking water. The most common technique employed by U.S. water utilities is the addition of
phosphate as orthophosphate, polyphosphate, or a blend of both. Research has demonstrated that
the use of phosphate successfully reduces lead release. The use of phosphate, however, could
stimulate microbial growth because it is an essential nutrient for microorganisms. Furthermore,
phosphate addition by the water utility may create a phosphate removal problem for the local
wastewater treatment facility if it discharges to an environmentally sensitive waterway.
The use of phosphate by St. Paul Regional Water Services (SPRWS) resulted in a dramatic
increase in positive total coliform samples during routine water sampling (Hozalski et al., 2005).
The ability of phosphate to stimulate microbial activity in the SPRWS system was subsequently
confirmed in a pipe loop study (Hozalski et al., 2005). Hence, SPRWS decided to explore alternative
techniques for lead corrosion control.
One alternative corrosion control technique is the use of stannous chloride as a corrosion
inhibitor chemical. Stannous chloride has a wide variety of industrial uses and has been approved
by the US Food and Drug Administration (USFDA) as an antiodixidant and preservative in food
products. Stannous chloride has also been approved for use as a water treatment chemical by NSF
International under Standard 60. Although not commonly used in the water treatment industry,
Hozalski et al. (2005) performed pipe loop experiments and demonstrated that stannous chloride
can decrease lead release and heterotrophic plate counts in comparison to an untreated control.
Unfortunately, at the time of this research effort little was known about tin chemistry in drinking
water and the mechanism by which tin decreases lead corrosion.
Thus, the main goal of this research was to elucidate the mechanism or mechanisms by
which SnCl2 decreases the corrosion of lead and the corresponding release of Pb into the water
supply. The specific objectives of the research were as follows:

1. Improve our understanding of tin chemistry in drinking water


2. Investigate the toxicity of tin to both planktonic and biofilm bacteria
3. Determine the mode of action of SnCl2

The form of tin in water determines how the tin interacts with the lead surface and/or bacte-
ria to reduce corrosion. Also, if tin reacts with chlorine, it will exert a chlorine demand. Tin toxicity
may play a role in corrosion control by limiting microbially influenced corrosion and its addition
to a water supply may enhance biostability. Understanding the mode of action of tin as a corrosion
inhibitor is critical to understand its potential applicability for various distribution systems and
water qualities and to understand its possible application as a corrosion inhibitor.

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xxii | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

RESULTS

Tin Chemistry

The chemistry of tin in drinking water was investigated, including its reactivity with chem-
ical oxidants. Sn+2 was oxidized to Sn+4 by all three of the oxidants investigated: dissolved oxygen,
free chlorine, and monochloramine. In terms of chlorine demand, 0.42 mg of NH2Cl as Cl2 was
consumed per mg of Sn+2 added. Dissolved oxygen oxidizes Sn+2 to Sn+4 at a relatively slow rate
(k = 0.069 hr–1). Both free chlorine and monochloramine oxidize Sn+2 in a matter of seconds (k >
42 hr–1). The oxidation of Sn+2 is typically not complete, resulting in a mixture of Sn+2 and Sn+4 in
the water.

Toxicity of Tin to Planktonic and Biofilm Bacteria

Sn2+ inhibited the growth of Pseudomonas aeruginosa PAO1 and Nitrosomonas europaea
and inhibited the aerobic respiration of P. aeruginosa PAO1, Escherichia coli K-12, N. euro-
paea and a mixed culture of nitrifying bacteria in the presence of excess electron donor. The
Sn2+ minimum inhibitory concentrations for P. aeruginosa PAO1 and N. europaea were 1.5 mg/L
(0.013 mM) and > 4.4 mg/L (0.037 mM), respectively. Sn2+ increased the inactivation of P. aerugi-
nosa PAO1 and E. coli K-12 in the absence of electron donor and the extent of tin inactivation was
a function of the tin to biomass ratio. LD50 (lethal dose at which 50% of of subjects will die) values
for P. aeruginosa PAO1 and E. coli K-12 were 10–6.24 and 10–6.25 mg Sn/CFU, respectively. Thus,
SnCl2 is highly toxic to lab strains of heterotrophic bacteria and is similar to or more toxic than
other heavy metals, with the possible exception of Hg. The observed toxicity of SnCl2 suggests
that the chemical may aid in controlling bacterial regrowth in distribution systems. Although only
limited data with nitrifying bacteria were obtained because of the relatively slow growth of these
organisms, it appears that nitrifiers are more resistant to Sn2+ toxicity than heterotrophic bacteria.
The environmental bacteria appeared to be more resistant to stannous chloride toxicity. There was
little inactivation of an environmental isolate (Acinetobacter sp.). Stannous chloride, however,
inhibited the accumulation of P. aeruginosa biofilm at a concentration of 0.5 mg/L as tin and was
able to remove established P. aeruginosa biofilm at 1 mg/L as tin. All of these inactivation experi-
ments were performed without free chlorine or chloramine present.

Effect of Stannous Chloride on Lead Corrosion

Lead coupons were aged for 70 weeks with lab and tap water in the presence and absence of
chlorine and presence and absence of stannous chloride. Minerals observed on the coupon surfaces
at the end of the aging period did not vary significantly among the different treatments and were
dominated by: lead (II) oxide (PbO), cerussite (PbCO3), and hydrocerussite (Pb3(CO3)2(OH)2­).
Plattnerite (PbO2) was also detected on a few coupons. During the aging period, the electrical
potential values for coupons aged in the presence of stannous chloride were similar to those for
coupons aged without stannous chloride. Some of these aged coupons were placed in semi-batch
reactors containing lab water with 1% R2A medium and inoculated with Mississippi River water
bacteria to investigate the role of microbially-influenced corrosion and the effects of stannous
chloride on lead corrosion. Bacteria and biofilms were prevalent on the lead coupons at the end of
the lead corrosion experiment for all reactors, including those containing tin and those without tin.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Executive Summary | xxiii

Furthermore, stannous chloride did not affect the concentration of dissolved or total lead released
from the coupons.

CONCLUSIONS

Sn+2 was oxidized to Sn+4 by oxidants typically present in treated water: chlorine, mono-
chloramine, and dissolved oxygen. The oxidation of Sn+2 was rapid but typically incomplete,
resulting in a mixture of Sn+2 and Sn+4 in the water even in the presence of residual chlorine. The
measured chlorine consumption (0.42 mg of NH2Cl as Cl2 per mg of Sn+2 added) coupled with the
low recommended dose for water treatment (≤0.2 mg/L as SnCl2) indicates that use of stannous
chloride will result in a minor chlorine demand (<0.053 mg/L as Cl2).
Stannous chloride was highly toxic to laboratory strains of heterotrophic bacteria (P. aeru-
ginosa PAO1 and E. coli K-12) but not as toxic to a laboratory strain of nitrifiers (N. europaea).
Compared to these lab strains, environmental bacteria were significantly more resistant to Sn tox-
icity. For example, an environmental isolate (Acinectobacter sp.) from the Mississippi River was
resistant to inactivation by stannous chloride and stannic chloride. These toxicity and inactivation
experiments were performed with planktonic bacteria in the absence of chlorine to isolate the
effects of the stannous chloride. Additional experiments were performed to evaluate the toxicity of
stannous chloride addition on biofilm bacteria.
Stannous chloride successfully inhibited P. aeruginosa biofilm formation at doses as low
as 0.5 mg/L as Sn+2 and removed established biofilm at a dose of 1 mg/L as Sn+2. Also, because
stannous chloride was only tested on developed biofilm at a concentration of 1 mg/L as Sn in
the absence of chlorine and presence of substantial nutrients, it is unclear how stannous chloride
would impact established biofilm under conditions that better reflect those in water distribution
systems. The observed toxicity of SnCl2 suggests that the chemical may aid in controlling bacterial
regrowth in distribution systems and may inhibit MIC.
Minerals observed on the lead coupon surfaces at the end of the 70 week aging period did
not vary significantly among the different treatments and were dominated by lead oxide (PbO),
cerussite (PbCO3), and hydrocerussite (Pb3(CO3)2(OH)2­). Stannous chloride appeared to have little
effect on the electrical potential during aging or on the lead mineralogy that developed during the
aging process. Aged and new coupons were employed in semi-batch reactors to investigate the
role of MIC and the effects of stannous chloride on lead corrosion (i.e., lead release). Bacteria
and biofilms were prevalent on the lead coupons at the end of the lead corrosion experiment for
all conditions suggesting that microbially-influenced corrosion (MIC) may be important for lead.
Furthermore, stannous chloride did not decrease the accumulation of biofilm on the coupons or
the concentrations of dissolved and total lead released from the coupons. In fact, in many experi-
ments lead levels were greater in the tin-containing bottles in comparison to the other treatments.
In comparison to typical tap waters, these experiments were performed under relatively high nutri-
ent levels and without chlorine. In previous pipe loop experiments with a treated water contain-
ing monochloramine, stannous chloride use reduced lead levels and heterotrophic plate counts in
comparison to an untreated control, but lead levels were still significantly greater than those for a
pipe loop fed orthophosphate. Certainly the experiments performed in this study covered a small
range of possible treated water qualities, so more work is needed to evaluate the effectiveness of
stannous chloride use under a wide range of conditions. Finally, the bacteria on the coupons were
not identified and their role in lead corrosion, if any, was not elucidated.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


xxiv | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

RECOMMENDATIONS

Utilities considering use of stannous chloride for lead corrosion control should proceed
slowly and with caution. In previous pipe loop studies, stannous chloride showed some benefit
with respect to reducing lead and bacterial levels, but the chemical did not decrease lead release or
biofilm accumulation in the batch experiments with lead coupons performed in this work. Coupon
and possibly pipe loop studies are recommended for utilities considering use of stannous chloride
to evaluate whether the chemical might be effective for the given water quality conditions.
More research is needed to identify the bacteria that accumulate on lead surfaces and to elu-
cidate their activity, including a possible role in MIC. Furthermore, additional research is needed
to investigate the effects of stannous chloride on lead corrosion and microbial activity for a wider
range of water qualities including low levels of free chlorine and chloramines. Also, there is a need
for mechanistic studies with more sensitive surface analytical techniques to determine Sn valence
state and surface speciation. In addition, although sensitive techniques are available for measuring
total Sn in water, more sensitive techniques are needed to distinguish between Sn(II) and Sn(IV).
Finally, more information is needed on Sn complexation in water (and related stability constants),
as Sn speciation is likely to have a strong role in bioavailability and toxicity.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


CHAPTER 1
INTRODUCTION

Lead drinking water pipelines were installed throughout the United States (U.S.) in the
past due to its malleable nature and relative resistance to corrosion. When lead was discovered
to be a neurotoxin to humans, however, the installation of lead pipe in water distribution systems
was discontinued. Unfortunately, lead can still enter drinking water from existing lead service
connections, lead solder, and brass devices such as faucets. Due to the concerns regarding high
lead exposure via drinking water, the Lead and Copper Rule (LCR) was established by the US
Environmental Protection Agency in 1991. The LCR established a 90th percentile action level
(AL) of 15 µg/L as total Pb.
In response to the LCR, many water treatment plants across the U.S. have implemented
corrosion prevention plans to decrease lead release into drinking water. The most common tech-
nique employed by U.S. water utilities is the addition of phosphate as orthophosphate, polyphos-
phate, or a blend of both. Research has demonstrated that the use of phosphate successfully reduces
lead release (Hozalski et al., 2005; Edwards and McNeill, 2002; Karalekas, 1983). The use of
phosphate, however, could stimulate microbial growth under some circumstances because it is an
essential nutrient for microorganisms.
The use of phosphate by St. Paul Regional Water Services (SPRWS) resulted in a dramatic
increase in positive total coliform samples during routine water sampling (Hozalski et al., 2005).
During that period, everything in the system was kept constant except for the increased concentra-
tion of phosphate. The ability of phosphate to stimulate microbial activity in the SPRWS system
was subsequently confirmed in a pipe loop study (Hozalski et al., 2005). Hence, SPRWS decided
to explore alternative techniques for lead corrosion control.
Stannous chloride has a wide variety of industrial uses including: a common catalyst in
the pharmaceutical industry; a sensitizing and reducing agent in the production of tin chemicals
and metal manufacturing; and in the manufacturing of common household items such as mirrors,
tanning agents, and oral healthcare products (Mason, 2008). It has been approved by the US Food
and Drug Administration as an antiodixidant and preservative in food products at up to 0.0035%.
Hence, it is presently found in canned goods such as asparagus and fruits (USFDA, 2003). It is
also added to beverages as a color preservative (USFDA, 1982). Stannous chloride has also been
approved for use as a water treatment chemical by NSF International. Although not commonly
used in the water treatment industry, Hozalski et al. (2005) demonstrated that stannous chloride
can decrease lead release and heterotrophic plate counts in comparison to an untreated control.
The overall purpose of the research performed herein was to investigate the mechanism
and effectiveness of stannous chloride for lead corrosion control. For this, three specific objectives
were further considered. The first objective was to investigate the chemical reactions between stan-
nous ion (Sn+2) and common oxidants added to drinking water, namely chlorine and chloramine.
The hypothesis was that stannous chloride will be oxidized by these oxidants to Sn+4. The form
of tin present in water is important to determine its interaction with lead and other constituents in
the water, and therefore, important in understanding lead corrosion control. In addition, it is also
important to know whether stannous chloride would exert a chlorine demand.
There are no known human health effects of ingesting drinking water dosed with SnCl2
but laboratory studies have shown endotoxic effects and reduced cell viability in Escherichia coli
cultures (Dantas et al., 1996) and in mammalian cell cultures (Dantas et al., 2002). For example,

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2 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

SnCl2 concentrations as low as 55.5 mM (10.5 mg/L) induced “low but significant levels of DNA
damage” in human-derived K562 cells (Dantas et al., 2002). Other researchers have reported that
SnCl2 has mutagenic (Singh, 1983) and carcinogenic (Ashby and Tennant, 1991) characteristics. It
is important to note that the low or sub-mg/L concentrations added to drinking water for corrosion
control are about two orders-of-magnitude lower than the lowest levels shown to have deleterious
effects in laboratory cultures. Hence, the second objective was to determine the toxicity of stan-
nous chloride to a wide variety of planktonic and biofilm bacteria. It was hypothesized that stan-
nous chloride would inhibit growth and increase inactivation of bacteria.
The last objective of this work was to investigate the mode of action by which stannous
chloride decreases lead corrosion. One possibility is that a Sn-containing precipitate (e.g., Sn(OH)2,
Sn(OH)4) deposits on the pipe wall providing cathodic or anodic protection. Another possibility is
that stannous chloride decreases the rate of MIC. Microorganisms have the ability to accelerate the
rate of corrosion by direct and indirect means. The addition of stannous chloride might inhibit and
inactivate bacteria, thereby, decreasing MIC and overall lead corrosion.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


CHAPTER 2
LITERATURE REVIEW

LEAD CORROSION

Corrosion may be defined as the deterioration of a metal by either chemical or electro-


chemical effects. The result of corrosion is the oxidation of the metal and release of soluble metal
into the local or surrounding environment (Figure 2.1). The soluble metal ions can combine with
other constituents in the water to form solid precipitates, which can deposit on the metal surface
or be carried away as particulate metal. The corrosion process requires an oxidant to consume the
electrons produced during oxidation of the metal. Oxidants are typically plentiful in water distribu-
tion systems and include chlorine and dissolved oxygen.
Factors contributing to corrosion are complex. For lead corrosion in the water industry, the
complexity of the aqueous matrix continues to challenge researchers. Still, some factors of contri-
bution are widely known. They include but are not limited to: the age of the pipe, the metallurgi-
cal properties of the pipe, the water flow rate, water stagnation times, electrochemical properties
of the pipes, and chemical characteristics of the water. The chemical characteristics of the water
include: temperature, pH, alkalinity or dissolved inorganic carbonate (DIC), corrosion inhibitors
(e.g., phosphate), and biocides. As a result of these complex factors, lead release can vary with
location in a given system and over time. This review discusses some of these aspects and their
contributions to lead corrosion.
The age of pipelines plays an important role regarding corrosion and, thereby, Pb release.
Lead release generally decreases with age because older pipes contain protective layers on their
walls that decrease the release of metallic compounds. For lead pipes in water distribution sys-
tems, these protective films are generally in the form of relatively low solubility lead carbonates
(Schock, 1989). Edwards et al. (2002) confirmed this in a study looking at lead release from lead
pipes of different aging periods while adding different corrosion inhibitors. The study found that
dissolved lead decreased significantly with pipe age, but the decrease of particulate lead was not as
significant (Edwards et al., 2002).
Water alkalinity and pH affect lead solubility significantly. While the dissolved Pb+2 ion
is favored at low pH values, the solid phase lead carbonate (PbCO3) is stable at neutral pH, and
the hydroxycarbonate form (Pb3(CO3)2COH2) is stable at high pH values (Crittenden and MWH,
2005). Hence, a distribution system at higher pH would stabilize the solid phases of lead. Alkalinity
serves to buffer pH and stabilizes the solid phase to enable the formation of protective layers on
the metal surface. The effectiveness at which this protective layer forms depends on pH and DIC.
Schock (1989) constructed lead solubility diagrams based on pH-DIC. The general relationship in
the diagrams suggested a low DIC concentration of (<10 mg C/L) and a high pH (>9) to optimize
the reduction of lead solubility (Crittenden et al., 2005; Schock, 1989).

Effect of Biocides on Corrosion

The biocides most commonly used in the water industries are free chlorine and monochlo-
ramine. Both biocides are strong oxidants and have the ability to oxidize metals. Although free
chlorine is a stronger oxidant and disinfectant, monochloramine is more stable and minimizes the

©2010 Water Research Foundation. ALL RIGHTS RESERVED


4 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

2+ 2- - -
Pb + CO3 ½O2 + H2O + 2e · 2OH

PbCO3 (s)

-
2e

Anode Cathode
Pb0

Figure 2.1 Diagram showing the process of lead pipe corrosion

formation of halogenated disinfection byproducts in the distribution system. For these reasons,
many water utilities have converted or are considering converting from free chlorine to monochlo-
ramine as terminal disinfectant. Unfortunately, this changeover can have dramatic consequences,
such as in the highly publicized case of Washington, D.C. (Renner, 2004; Vasquez et al., 2006).
When the system in Washington D.C. was converted from free chlorine to monochloramine, the
concentrations of lead in some parts of the system increased dramatically. The explanation is a
shift from Pb(IV) minerals to Pb(II) minerals, as Pb(IV) minerals such as PbO2 can form in the
presence of free chlorine while Pb(II) minerals are thought to dominate in systems containing
monochloramine (Edwards and Dudi, 2004). In addition, monochloramine has the ability to attack
brass, so the change may have also increased the leaching of Pb from brass fixtures (Renner, 2004;
Edwards and Dudi, 2004). Another possible explanation considers the role of ammonia oxidizing
bacteria (AOB) (Edwards and Dudi, 2004), as discussed below. Although these common biocides
may increase corrosion in water distribution systems, it is important to remember that biocides
are used is to ensure the delivery of safe pathogen free drinking water to consumers. In addition,
biocides added to water might decrease microbially influenced corrosion by controlling the accu-
mulation of biofilms.

MICROBIALLY INFLUENCED CORROSION

Another potential cause of lead release from the pipe walls is related to the presence of
microorganisms which accelerate corrosion. This process is known as microbially-influenced cor-
rosion (MIC). MIC could potentially accelerate lead pipe degradation by (Sand, 1997):

a. increasing electron transfer from surface


b. destroying the surface protective film layers
c. changing the redox potential of the surface for their metabolic advantage
d. creating highly acidic corrosive substances

Each of these methods could potentially change the surface mineralogy, the metallic structure, and/
or the electro chemical properties of the metal pipe and enhance metal release (Sand, 1997).
The bacteria that may play a role in MIC in water distribution systems include: sulfate
reducers, nitrate reducers, sulfur bacteria and biofilm bacteria (Crittenden et al., 2005). These

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 2: Literature Review | 5

Figure 2.2 Schematic diagram illustrating the three stages of biofilm formation (MSU-CBE©,
P. Dirckx)

organisms are directly associated with the metal surface in biofilms. Biofilm bacteria are difficult
to activate due to their increased resistance to biocides. Biofilm consists of bacteria embedded in
a slime layer of extracellular polymeric substances (EPS). The structure and thickness of these
biofilms are dependent on the species present, nutrient availability, fluid shear, and presents of
biocides.

Biofilm

Biofilm may be produced by many types of bacteria and are believed to be composed of
proteins and polysaccharides. The time length for biofilm formation is dependent on the species
that forms the biofilm and may take from several minutes up to many hours. Nevertheless, it is
believed that there are primarily three stages in biofilm formation (Figure 2.2). The first stage is
the conditioning film, where the formation of proteins and other forms of organic compounds from
the local environment are combined due to electrostatic arrangements. At this stage, the bacteria
are at their planktonic state. The next stage is where the bacteria began to attach to the EPS and is
termed sessile bacteria at this point. The last stage is the actual attachment of the bacteria through
EPS to the film (King, 2007; McKubre and Syrett, 1986).
The EPS in the biofilm is thought to protect the bacteria inside the biofilm from external
potentially harmful substances. The protection effectiveness of these films is dependent on the tox-
icity of the external matter and the bacterial resistance to toxic substances. Additionally, the EPS
films may trap organic or inorganic particles which could serve as food for the bacteria (Taheri et
al., 2005). The EPS could also sorbs heavy metals such as Pb or Cd and reduce their concentration
in the water (Guibaud et al., 2008).
One possible effect of biofilm formation is ennoblement, where the corrosion potential
is displaced to the cathode region, making the metal surface more vulnerable to pitting (Videla,
1996). Other effects of biofilm formation are significant alterations to pH, dissolved oxygen, and
the release of corrosive chemicals to the local water environment. For example, biofilms containing

©2010 Water Research Foundation. ALL RIGHTS RESERVED


6 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

sulfate-reducing bacteria (SRB) produce hydrogen sulfide. The resulting sulfide compound can
alter the surface chemistry and redox potential of the metal.

Sulfate-Reducing Bacteria

To our knowledge, there are no reports providing direct evidence of the MIC of lead sur-
faces. There are, however, many reports concerning MIC of other common water distribution pipe
materials such as copper, iron, and steel. Lytle et al. (2005) found significant concentrations of
iron-sulfide minerals on iron pipe walls which are believed to be the result of SRB activity. SRB
can use the hydrogen produced from the reduction of water as their energy source and sulfate as
the electron acceptor, which releases sulfide in the process. The hydrogen sulfide then reacts with
Fe2+, resulting in the formation of iron-sulfide minerals. The sequence of reactions is shown below.

Fe0 → Fe2+ +2e–  (2.1)

2H+ + 2e– → H2 (2.2)

SO42– + 4H2 → S2– + 4H2O  (2.3)

Fe2+ + HS– → FeS + H+ (2.4)

2FeS + ½H2O + ¾O2 → FeS2 + FeOOH (2.5)

The final products from these reactions on iron pipes may be FeS and FeS2 (Lytle et al., 2005;
McNeil and Odom, 1994). Similarly for copper, McNeil and Odom (1994) proposed the following
reactions, where the final product is CuS:

Cu0 → Cu2+ + 2e– (2.6)

CuO + HS– → CuS +OH– (2.7)

In a similar manner as Fe and Cu, hydrogen sulfide produced by SRB may also react with
Pb on the lead surface to form PbS (McNeil and Odom, 1994; Duncan and Ganiaris, 1987; Stoffyn
and Buckley, 1992, McNeil and Little, 1990)

PbO + HS– → PbS (galena) + OH–  (2.8)

Pb is toxic to microorganisms (Loka Bharathi et al., 1990; Waara, 1991), so the formation of PbS
may be a method of detoxification for SRB (Aiking et al., 1985; Capone et al., 1983). The forma-
tion of PbS in water distribution systems is unlikely, however, because the water typically is oxy-
genated and contains residual chlorine.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 2: Literature Review | 7

Ammonia-Oxidizing Bacteria

MIC may also result from the activity of ammonia oxidizing bacteria. Ammonia formed
from the decay of chloramines can serve as electron donor for AOB, resulting in the formation of
nitrite

NH3 + 3⁄2O2 + → NO2– + H+ + H2O (2.9)

The nitrite that forms may then attack the lead metal to reform ammonia

NO2– + 3Pb0 + 2H2O → NH3 + 3PbO + OH–  (2.10)

or N2

2NO2– + 3Pb0 + 2H2O → N2 + 3PbO + 2OH– (2.11)

as indicated in Edwards and Dudi (2004). In the case of the reformation of ammonia (reaction 10),
the sequence of reactions 9 and 10 becomes collectively autocatalytic as the product from one
reaction serves as the reactant for the other. It is also possible that the nitrite is further oxidized
to nitrate by nitrite oxidizing bacteria with the nitrate directly attacking the lead surface to form
nitrite as reported by Uchida and Okuwaki (1999)

NO3– + Pb0 → NO2– + PbO  (2.12)

Controlling MIC

MIC can be minimized by preventing the formation of biofilm or removing established


biofilm. It is best to prevent biofilm from developing because of the difficulty in removing estab-
lished biofilms. To minimize MIC and corrosion, methods applied in water distribution systems
may be physical, chemical, or electrochemical. Physical treatments involve flushing of the pipes at
high velocity. Chemical treatments involve the addition of biocides and electrochemical methods
refer to cathodic protection. The most common method in the water industry is the application of
biocides due to cost reasons (Javaherdashti, 2008).
Biocides are added in water distribution systems primarily to inactivate pathogens to ensure
safe drinking water for the consumer. The most commonly used biocides are chemical oxidants,
namely free chlorine, chlorine dioxide, ozone, and chloramines. Sometimes multiple chemicals
are used in combination as primary and secondary disinfectants. This is always the case when
chlorine dioxide or ozone is used for primary disinfection as these chemicals do not maintain a
residual in the distribution system. Free chlorine and chloramines help to control microorganisms
in distribution systems but can increase corrosion and the release of metals from pipes (Renner,
2004; Edwards and Dudi, 2004). The addition of free chlorine, and to a lesser extent, chloramines,
also results in the formation of disinfection by-products (e.g., trihalomethanes and haloacetic
acids) from reactions with natural organic matter in the water. While the use of ozone for primary
disinfection reduces the formation of halogenenated disinfection by-products, it might increase

©2010 Water Research Foundation. ALL RIGHTS RESERVED


8 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

assimilable organic carbon and bacterial counts, which in turn could increase pipe corrosion via
MIC (Escobar and Randall, 2001).

CORROSION INHIBITORS

Orthophosphate

Several types of corrosion inhibitors are commonly used in the U.S. Most water utilities
add phosphate to decrease lead corrosion. Generally, phosphate is added as orthophosphate (i.e.,
phosphoric acid or zinc orthophosphate), polyphosphate, or a combination of both. Orthophosphate
works by forming a low solubility hydroxypyromorphite or other Pb(II) orthophosphate mineral
phase on the lead surface. Numerous studies have demonstrated the effectiveness of orthophosphate
for lead corrosion control (Hozalski et al., 2005; Edwards et al., 2002). On the other hand, poly-
phosphate can increase the solubility of metals because it acts as a metal chelator. Polyphosphate is
often considered a corrosion inhibitor, and is typically applied in combination with orthophosphate
for lead corrosion control because it reverts to orthophosphate (Holm and Schock, 1991; Edwards
et al., 2002).

Silicate

Silicate is another corrosion inhibitor that has been investigated for corrosion control. In
pipe loop tests, silicate was effective when applied at concentrations exceeding 20 mg/L SiO2/L
(Schock et al. 2005; Shock, 1989). It was concluded that a long induction period (~8 months) is
necessary for the formation of a protective film (Schock et al. 2005; Shock, 1989).

Stannous Chloride

Stannous chloride (SnCl2) has been used as an antioxidant and preservative in food prod-
ucts and is approved by the U.S. Food and Drug Administration for such purposes (USFDA, 1982;
USFDA, 2003). One regulation specifies a maximum level of 0.0015% as tin in food (USFDA,
1982), while a company was granted a variance to test SnCl2 as a preservative of canned aspara-
gus spears at concentrations up to 35 parts-per-million (ppm) or 0.0035% of SnCl2 in the finished
food (USFDA, 2003). Stannous chloride is a relatively new corrosion inhibitor in that it has only
recently been approved by NSF International for use in potable water distribution systems at con-
centrations in the sub-mg/L to low mg/L range (NSF International, 2008). Little is known about the
effectiveness of SnCl2 for lead corrosion control in potable water distribution systems and to our
knowledge, there was no information available in the peer-reviewed literature prior to the publica-
tion of our manuscript concerning a pipe loop study (Hozalski et al., 2005).
Stannous ion (Sn+2) is sparingly soluble in water (12.6 mM = 1.49 mg/L) at circumneutral
pH and 25°C (Figure 2.3). Thus, a concentrated stock solution must be prepared by dissolving
SnCl2 in strong acid (e.g., 10% HCl). The chemical is typically applied at low doses of ~0.2 mg/L
as SnCl2 (0.13 mg/L as Sn+2). The mode of action of stannous chloride as a corrosion inhibitor
is unclear. When added to water containing dissolved oxygen or chlorine or both, it will likely
be partially or completely oxidized to the stannic or Sn(IV) form (Cotton and Wilkinson, 1980)
because the standard electrode potential for the Sn+4/Sn+2 redox couple is +0.1539 V.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 2: Literature Review | 9

3
2 2+
Sn
1

log concentration (M)


0
-1 Sn(OH)2 (s)
-2
-3 -
Sn(OH)3
-4
-5 0
Sn(OH)+ Sn(OH)2
-6 2+
Sn2(OH)2
-7 2+
Sn3(OH)4
-8
0 2 4 6 8 10 12 14
pH

Figure 2.3 Diagram showing the solubility of Sn(II) in equilibrium with Sn(OH)2 (s) at 25°C

There are no known human health effects of ingesting drinking water dosed with SnCl2
but laboratory studies have shown endotoxic effects and reduced cell viability in Escherichia coli
cultures (Dantas et al., 1996) and in mammalian cell cultures (Dantas et al., 2002). For example,
SnCl2 concentrations as low as 55.5 mM (10.5 mg/L) induced “low but significant levels of DNA
damage” in human-derived K562 cells (Dantas et al., 2002). Other researchers have reported that
SnCl2 has mutagenic (Singh, 1983) and carcinogenic (Ashby and Tennant, 1991) characteristics. It
is important to note that the low or sub-mg/L concentrations added to drinking water for corrosion
control are about two orders-of-magnitude lower than the lowest levels shown to have deleterious
effects in laboratory cultures.
Because of the aforementioned coliform and regrowth issues that occurred during phos-
phate addition, SPRWS was interested in finding an alternative corrosion inhibitor for lead corro-
sion control. Since 2000, SPRWS has been adding stannous chloride to their distribution system
at a concentration of 0.175 mg/L as SnCl2 during the summer season and 0.125 mg/L as SnCl2 the
rest of the year. Results from sampling selected “problem” sites in the full-scale system suggested
that stannous chloride aided in decreasing lead concentrations compared to the preceding period
when no corrosion inhibitor was applied (i.e., “Untreated”) (Figure 2.4). Unlike when phosphate
was applied, no regrowth issues or increases in coliform counts have been observed since SPRWS
began feeding stannous chloride.

PRELIMINARY STANNOUS CHLORIDE RESEARCH IN OUR LABORATORY

Pipe Loop Study

A pipe loop system with five parallel lines comprised of new ductile iron and lead pipes was
operated for 13 months to investigate the effects of corrosion control chemicals on lead release and
bacterial regrowth (Hozalski et al., 2005). Four of the pipe loops were treated with corrosion con-
trol chemicals (orthophosphate, polyphosphate, orthophosphate/polyphosphate blend, and SnCl2)

©2010 Water Research Foundation. ALL RIGHTS RESERVED


10 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

30
Untreated
25
SnCl2

20

Total Pb, ug/L


15

10

0
A B C D E
Sampling site

Figure 2.4 Comparison of mean lead concentrations for “problem” sampling sites in the
SPRWS system prior to (untreated) and during SnCl2 addition. The lead AL is indicated by
the horizontal line.

and one served as an untreated control. To our knowledge, this was the first direct comparison of
SnCl2 and phosphate-based corrosion inhibitors for lead corrosion control over an extended period
of time in a flow-through pipe loop system. The pipe loop system received chloraminated and
filtered surface water from a full-scale lime softening plant and was monitored for total Pb, dis-
solved Pb, heterotrophic plate count (HPC), and coliform concentrations. Total Pb concentrations
in each of the treated loops were significantly lower (95% confidence level) than in the untreated
control, and orthophosphate (1 mg/L as P) consistently yielded the lowest Pb concentrations of all
of the chemicals tested including SnCl2 (0.125 mg/L). Nevertheless, none of the chemicals was
able to consistently maintain total Pb concentrations below the 15 mg/L action level for the 8-hour
stagnation time sample in the new lead pipes used for this study. In addition, HPC concentrations
in all of the loops amended with phosphate-containing chemicals exceeded those in the untreated
control loop during the warm summer months. Conversely, HPC levels in the SnCl2 loop were
significantly lower than in the control during the same period.
Corrosion control chemicals typically limit corrosion by combining with the metal ions
released at the anode to form a low solubility solid scale, such as the hydroxypyromorphite formed
when orthophosphate combines with Pb2+. Given the lack of information available in the peer-
reviewed literature on SnCl2 for corrosion control, it is unclear how the chemical slows release of
Pb. In order to address this issue, X-ray diffraction (XRD) analyses were performed on samples
of pipe removed from the SnCl2 loop. The interior of the pipe surface contained a whitish-yellow
solid which consisted primarily of hydrocerrusite (Pb3(CO3)2(OH)2) and no tin minerals were
detected. Possible explanations for the failure to detect tin minerals include the presence of a thin
tin mineral layer with insufficient mass to be detectable by the X-ray diffractometer, existence of
tin in an amorphous phase, loss by dissolution while the pipe loop was shut down prior to sample
collection, or loss during removal and subsequent handling of the pipe section. Thus, more work
was needed to study the mode of action of SnCl2 for control of Pb corrosion.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 2: Literature Review | 11

Utilities like SPRWS are faced with a dilemma; use phosphate to provide the best lead
corrosion control but risk enhancing regrowth, or use SnCl2, which is poorly understood and may
not adequately control lead release (based on the results of the pipe loop study). SPRWS opted to
use SnCl2 before the pipe loop study was completed because the utility had no other viable alter-
native. The results of the pipe loop study and full-scale system monitoring at SPRWS suggested
that SnCl2 can decrease lead release (in comparison to an untreated control) without negatively
affecting microbiological water quality. Nevertheless, our lack of understanding of tin chemistry in
treated drinking water and of the mode of action of the chemical in lead corrosion control severely
limited our ability to predict its applicability and effectiveness for different water systems. Thus,
the research described herein was performed to investigate tin chemistry and the mechanisms by
which tin controls lead corrosion.

Preliminary Bench-Scale Experiments

Preliminary experiments were performed by the principal investigator while on sabbatical


at the Center for Biofilm Engineering at Montana State University. The goals of the experiments
were to: (1) develop methods for analyzing Sn2+ and Sn4+ in aqueous samples, (2) evaluate the rate
and extent of Sn2+ oxidation to Sn4+ in the presence of free chlorine and dissolved oxygen, and
(3) evaluate the toxicity of Sn2+ to bacteria including Pseudomonas aeruginosa PAO1.
Reports in the literature (Cheng et al., 1992; Neil et al., 1995) of a colorimetric method for
analyzing Sn2+ served as the starting point for method development. Also, total tin can be read-
ily analyzed by standard methods for analysis of metals in water (Standard Methods) including
atomic absorption spectrophotometry (AAS) and inductively coupled plasma–optical emission
spectrometry (ICP-OES) or ICP-mass spectrometry (ICP-MS). Because only Sn2+ and Sn4+ will
be present in water, the approach taken was to adapt the method for analysis of Sn2+ for use in
our experiments, measure total tin using ICP-OES or AAS, and determine Sn4+ by difference. The
colorimetric method involves reaction of Sn2+ with toluene-3,4-dithiol to form Sn(II)-dithiolate
producing a red color (Cheng et al., 1992; Neil et al., 1995).
Results from a batch Sn2+ oxidation experiment are shown in Figure 2.5. All experiments
to date have been performed at low pH (less than 3) so that high Sn2+ concentrations could be
used to provide clear results from the colorimetric method. Several observations from this data
(and subsequent experiments, data not shown) can be made: (1) Sn2+ is stable when no oxidant is
present, (2) the chlorine concentration is stable over the short time of the experiment when no Sn2+
is added, (3) Sn2+ is oxidized by free chlorine and exerts a chlorine demand, (4) the kinetics of
the oxidation reaction are so rapid that the reaction is over in less than 1 minute, and (5) the reac-
tion does not go to completion (i.e., there is still Sn2+ remaining) despite the presence of residual
chlorine. From subsequent experiments, the oxidation rate in the presence of free chlorine was
estimated (k > 42 hr–1). The effect of chlorine dose on the conversion of Sn2+ was investigated
and as expected, the fraction of the initial Sn2+ oxidized to Sn4+ increased with increasing chlorine
dose and was complete (100% oxidation) at very high chlorine doses (100 mg/L). The oxidation
of Sn2+ by dissolved oxygen (k = 0.069 hr–1) was much slower than by free chlorine and was also
incomplete (data not shown). The incomplete oxidation of Sn2+ in the presence of free chorine
(see Figure 2.5) can be explained by the Nernst equation:

[Sn 2 +] [OH -] [Cl -]


 (2.13)
( RT )
=
HOCl] exp – ∆G
[Sn ] [
4+ 0

©2010 Water Research Foundation. ALL RIGHTS RESERVED


12 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

12

10

Conc., mg/L
8 Chlorine consumed

0
0 1 2 3 4 5
Time, min.

Sn2+ Chlorine
Sn2+ (No Chlorine Control) Chlorine (No Sn2+ Control)

Figure 2.5 Sn2+ oxidation by free chlorine (initial concentration of ~10 mg/L as Cl2) at pH 2.63,
Temp. 22°C

0.45
0.4 No Sn2+ Control
0.35 0.25 mg Sn2+/L
0.3 0.5 mg Sn2+/L
0.25 0.75 mg Sn2+/L
OD 600

0.2 1.0 mg Sn2+/L


0.15 1.5 mg Sn2+/L
0.1 Control ave
0.05 1.0 mg/L ave
0
1.5 mg/L ave
-0.05
0 10 20 30 40 50
Time, hr

Figure 2.6 Effect of Sn2+ on the growth of P. aeruginosa PAO1 on 10% LB broth in batch
culture (OD600 = optical density at 600 nm)

which indicates that the ratio of Sn2+ to Sn4+ at equilibrium is inversely proportional to the chlorine
concentration, which is consistent with the data collected to date. The equation also indicates that
the ratio of Sn2+ to Sn4+ is proportional to the concentration of OH–, suggesting that the extent of oxi-
dation of Sn2+ will decrease with increasing pH. More data are needed to evaluate whether the equa-
tion shown here adequately describes the experimental observations, especially the effect of pH.
Preliminary experiments were also performed to evaluate the toxicity of tin to bacteria
(Figure 2.6). Results to date suggest that concentrations less than 1 mg Sn2+/L (added as SnCl2)

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 2: Literature Review | 13

do not have a significant effect on the growth of P. aeruginosa PAO1. P. aeruginosa PAO1 is fre-
quently used in laboratory investigations and was selected for this research because it is known to
form biofilms. At a tin concentration of 1 mg/L, growth was significantly delayed in comparison
to the control, but the final cell densities were the same. At a concentration of 1.5 mg/L, however,
growth was completely inhibited. Dantas et al. (1996) reported the rapid inactivation of E. coli
dosed with 37 mM SnCl2 (4.4 mg/L as Sn2+) and attributed this to the production of reactive oxy-
gen species generated in a Fenton-like reaction. Thus, it is clear that SnCl2 is toxic/inhibitory to
bacteria. This represents a positive benefit in that its use may aid in controlling bacterial regrowth
in distribution systems. Conversely, it raises some concerns regarding potential toxicity to higher
organisms, especially drinking water consumers. More work was needed to investigate the effects
of SnCl2 on bacteria, especially biofilm bacteria that predominate in water distribution systems.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


©2010 Water Research Foundation. ALL RIGHTS RESERVED
CHAPTER 3
MATERIALS AND METHODS

INTRODUCTION

Laboratory experiments were performed to investigate whether stannous chloride is an


effective lead corrosion inhibitor and its mode of action. Batch experiments were performed to
investigate the reactivity of stannous ion with drinking water oxidants followed by determining the
toxicity of stannous and stannic chloride to planktonic and biofilm bacteria. Finally, batch experi-
ments with lead coupons were performed to investigate the effects of tin on lead corrosion.

INVESTIGATION OF TIN CHEMISTRY

Kinetics of Monochloramine Consumption by Sn2+ 

A monochloramine stock solution (~40 mg/L) was prepared by adding ammonium chloride
slowly (drop wise at 10 µL) to a solution of sodium hypochlorite in 4 mM bicarbonate buffer at pH
9.6 (Kipper et al., 2006; Piyachaturawat, 2005). Then, 500 mL of diluted monochloramine solution
(~9 mg/L as Cl2) was placed into a beaker and chilled to the desired testing temperature (1, 5 or
10 °C). After measuring the initial monochloramine concentration and pH, SnCl2 was added to a
final concentration of 9 mg/L as Sn. Ten milliliter aliquots of the solution were removed periodi-
cally for analysis of chloramine concentration at approximately the following reaction times: 0,
10, 30, 60, 120, and 300 seconds. The concentrations of chloramine were measured using the N, N
diethyl-p-phenylenediamine-ferrous ammonium sulfate (DPDFAS) method (Standard Methods).

Effect of pH on Monochloramine Stability in the Presence of Sn2+ 

A 25 mM bicarbonate solution was prepared in ultra-pure water and the pH was adjusted
with concentrated HCl to the desired value (2, 6, 8, or 10). Monochloramine was added to a final
concentration of approximately 1 mg/L as Cl2 and the solution was transferred into a series of 7
to 8 flasks. After testing the pH of each flask, Sn+2 was added at concentrations ranging from 0 to
5 mg/L. After shaking the flasks thoroughly, the solutions were tested for monochloramine residual
using the DPDFAS.

Oxidation of Sn+2 With Monochloramine

The oxidation of Sn+2 to Sn+4 by monochloramine was evaluated by measuring the con-
centration of Sn+2 using a colorimetric method (described below) in addition to measuring the
chloramine residual using the DPDFAS method. A Sn+2 solution (~2.5 mg/L as Sn) was prepared in
24 mM bicarbonate buffer solution at approximately pH 8.5. The Sn+2 solution was added to eight
100 mL Erlenmeyer flasks which were dosed with monochloramine at concentrations ranging
from 0 to 1 mg/L as Cl2. After shaking each flask, 10 mL aliquots were removed for measurement
of Sn+2 while the remainder was used for the monochloramine analysis.

15

©2010 Water Research Foundation. ALL RIGHTS RESERVED


16 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

EFFECT OF TIN ON INACTIVATION OF PLANKTONIC BACTERIA AND BIOFILMS

Microorganisms

Three laboratory stains of bacteria were selected as model organisms for use in the toxic-
ity experiments: two heterotrophs (Pseudomonas aeruginosa PAO1 and Escherichia coli K-12)
and one autotroph (Nitrosomonas europaea). In addition, a few experiments were performed with
an undefined mixed population of nitrifying bacteria, with undefined mixed cultures from the
Mississippi River, and with an environmental isolate from a Mississippi River culture. Although
not likely to be the dominant organisms, P. aeruginosa and E. coli were selected because they
have been detected in water distribution systems (Tokajian et al., 2005; Emde et al., 1992) and for
experimental convenience as described below. N europaea has been used as a model nitrifier in
other microbial inactivation studies (Oldenburg et al., 2002).
P. aeruginosa PAO1 was acquired from the Center for Biofilm Engineering (CBE) at
Montana State University. P. aeruginosa PAO1 is frequently used in laboratory investigations and
was selected for this research because it is known to form biofilms. This bacterium has a genetic
insert that causes the cells to express green fluorescent protein (GFP) and that provides for resis-
tance to the antibiotic carbenicillin. P. aeruginosa PAO1 was stored on Luria-Bertani (LB) agar
plates and cultured in LB media containing 250 mg/L carbenicillin. E. coli K-12 was also acquired
from the CBE at Montana State University. Similar to the P. aeruginosa, this bacterium has a
genetic insert that causes the cells to express a fluorescent marker (DS-Red) and the organism is
also carbenicillin resistant. E. coli K-12 was stored on LB agar plates and cultured in LB broth
containing 250 mg/L carbenicillin. Organisms with fluorescent markers were selected for possible
use in glass capillary biofilm reactor experiments to facilitate real-time imaging of biofilms using
scanning-laser confocal microscopy.
N. europaea was purchased from the American Type Culture Collection (ATCC 19718).
Upon arrival, the bacterial suspension (1 mL) was transferred to a 125 mL erlenmeyer flask con-
taining 10 mL of ATCC growth medium 2265. After 14 days of incubation on a shaker table in the
dark at room temperature, the suspension (10 mL) was transferred to a 500 mL erlenmeyer flask
containing 100 mL of ATCC growth medium 2265. This step was repeated to generate sufficient
biomass for the toxicity experiments.
The undefined mixed culture of nitrifying bacteria was obtained from a membrane-
aerated bioreactor originally inoculated with a mixed culture of nitrifying bacteria that had been
enriched from wastewater collected from the Metropolitan Wastewater Treatment Plant (St. Paul,
MN, USA). The reactor was fed a carbon-free synthetic wastewater enriched with ammonium
(40–50 mg/L as N) at pH 8–8.4 and operated at room temperature (23 ± 2°C). More details on the
bioreactor experiments can be found elsewhere (Motlagh, 2008).
Undefined mixed cultures of bacteria were obtained from the Mississippi River in
Minneapolis, MN to represent environmental bacteria. River water was collected on four occa-
sions during the spring and summer of 2008 (April 19, 2008, June 17, 2008, June 23, 2008, and
June 26, 2008). An environmental isolate was obtained by selecting a colony from an R­2A agar
plate inoculated with Mississippi River water collected on July 20, 2008.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 3: Materials and Methods | 17

Batch Growth Curves

The effect of tin on the growth of P. aeruginosa PAO1 and N. europaea was investigated
by performing batch experiments over a range of tin doses. The bacteria were cultured as described
above. For the P. aeruginosa PAO1, a suspension (OD600 = 1.20, ~480 mg/L) was diluted 100 times
and then 40 mL aliquots of the diluted culture were added to 50 mL conical bottom plastic tubes
each containing 40 mL of 10% LB broth spiked with SnCl2 at concentrations (as Sn) ranging from
0 mg/L (control) to 1.5 mg/L (0 to 310 mg Sn/mg biomass). The tubes were prepared in duplicate.
One mL aliquots of a N. europaea suspension (OD600 = 0.062, ~25 mg/L) were transferred to six
250 mL Erlenmeyer flasks containing 100 mL of ATCC growth media 2265 spiked with SnCl2 at
concentrations (as Sn) ranging from 0 mg/L (control) to 4.4 mg/L (0 to 18 mg Sn/mg biomass). The
flasks were incubated on a shaker table in the dark at room temperature. Periodically, aqueous sam-
ples were withdrawn to measure optical density at 600 nm (OD600) and nitrate and nitrite (N. euro-
paea experiment only). OD600 was determined using a spectrophotometer (Shimadzu UV-1601PC).
Nitrate and nitrite were analyzed by ion chromatography (Metrohm Peak 761 Compact IC).

Bacterial Activity Assay

Oxygen uptake rate (OUR) testing was used to investigate the effects of relatively low
doses of SnCl2 (10.4–83.5 mg Sn/mg biomass) on bacterial activity. OUR was evaluated for the
single species and mixed cultures under three different tin treatments: no tin (control), tin in solu-
tion (Sn), and pretreatment of the bacteria with tin for 1 hour prior to transferring into Sn-free solu-
tion for the OUR experiment (Sn pretreated). P. aeruginosa PAO1 and E. coli K-12 were grown in
LB with 250 mg/L carbenicillin and harvested after 24 hours (exponential phase) or 72 hours (late
stationary/early decay phase) of incubation. N. europaea was grown in ATCC growth medium
2265 and harvested after approximately 14 days. The bacteria were collected by centrifuging at
5000  rpm for 25 min, washing with phosphate buffer solution and re-suspending in phosphate
buffer. A 2 mL aliquot of the suspension was transferred to a 300 mL biochemical oxygen demand
(BOD) bottle filled with sterile minimal medium amended with a high substrate concentration
(500 mg acetate/L for heterotrophs and 700 mg ammonia-N/L for autotrophs). The biomass con-
centration (dry weight) in the bottles ranged from 37 to 192 mg/L. The BOD bottle was placed
on a stir plate with a stir bar to provide mixing. A dissolved oxygen (DO) probe (YSI 200-BOD
probe) was used to monitor the DO concentration in the BOD bottle over a 1-2 hour period. The
OUR was obtained from the slope of the DO versus time plot and then normalized by the biomass
concentration.
Prior to the OUR experiments, an oxygen leakage experiment was conducted. A BOD
bottle was filled with deoxygenated (via stripping with nitrogen gas) distilled water and the DO
concentration was monitored over a 24 hour period. The oxygen leakage into the bottle from the
atmosphere caused a DO increase of 0.01 mg/L/hr, which was 2 to 3 orders of magnitude less than
the observed OUR in the active bottles. Thus, oxygen leakage was considered negligible.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


18 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Bacterial Inactivation Experiments

Lab Strains

First, all glassware for the biological experiments was cleaned with detergent and tap water,
followed by acid washing (overnight soak) and then rinsing at least three times with distilled water.
Finally, the glassware was autoclaved.
A single colony of either P. aeruginosa PAO1 or E. coli K-12 from a LB streak plate
was inoculated into LB media containing 250 mg/L carbenicillin and the bacteria were harvested
after 24 hours (exponential phase) or 72 hours (late stationary/early decay phase). The bacterial
suspension was centrifuged (10,000 rpm for 5 min) and the supernatant was discarded. The cells
were washed once in phosphate buffered saline (PBS), centrifuged and resuspended in PBS to the
desired final stock concentration (105–109 per mL). Then, 100 mL of the bacterial suspension was
transferred to a 1 mL vial containing 900 mL of test water containing different concentrations of
SnCl2 (0, 0.125, or 1 mg/L as Sn). Samples were withdrawn from the vial periodically (0, 5, 10,
20, 40, and 60 min) for enumeration.
Two different test waters were used in the bacterial inactivation experiments: a water pre-
pared in the laboratory by adding 10 mM NaHCO3 to ultrapure water (MilliQ system) and adjust-
ing the pH to 8.0 with HCl (lab water) and a treated water sample collected from the St. Paul
Regional Water Services (St. Paul, MN) filtration plant prior to any corrosion inhibitor addition
(tap water). The lab water is similar to the NSF 61 Section 9 test water (NSF International 2001)
except that chlorine was omitted to avoid rapid inactivation of the bacteria. The tap water sample
was dechlorinated and stored in a polystyrene bottle at 4°C. The tap water was brought to room
temperature prior to each set of experiments.
The experiments with N. europaea were performed similarly except that the bacteria were
grown in ATCC medium 2265 for approximately 2 weeks. The nitrifier suspension was centrifuged
(10,000 rpm, 5 min) and the supernatant was discarded. The pellet was washed with and resus-
pended in PBS to a stock concentration of 2 × 107 per mL. Then, 100 mL of this suspension was
transferred to 900 mL of lab water containing different SnCl2 concentrations (0.125 or 1 mg/L as
Sn). After 60 minutes of incubation, 200 mL of the suspension was transferred to a separate 1 mL
vial for enumeration of viable and non-viable bacteria using the Live/Dead assay.
P. aeruginosa and E. coli were enumerated using the spread plate method with LB agar.
The samples were serially diluted as necessary with PBS solution for plating to achieve a target of
300-3000 bacteria per mL. Then, a 100 mL aliquot of diluted sample was spread onto a LB plate.
After 3 days of incubation at room temperature, the colonies were counted.
N. europaea was enumerated using the Live/Dead assay (Sigma-Aldrich). The two dyes
(carboxyfluorescein diacetate and propidium iodide) were applied to the bacterial suspension and
the suspension was then incubated at 30°C for 15 minutes. After the incubation period, the sus-
pension was filtered through a 25 mm diameter 0.2 mm pore size polycarbonate membrane filter
(Millipore). Approximately 10 mL of distilled water was then filtered through the membrane to
remove excess dye. The filter was then mounted on a glass microscope slide with a drop of immer-
sion oil and a cover slip. The bacteria were counted by epifluorescence microscopy at a total mag-
nification of 1000× using a 100× oil immersion objective.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 3: Materials and Methods | 19

Undefined Mixed Cultures

Samples of water were collected from the Mississippi River in Minneapolis, MN on four
occasions (April 19, 2008, June 17, 2008, June 23, 2008, and June 26, 2008) to obtain bacteria
for batch inactivation experiments. Samples of river water and 10 times diluted river water were
spiked with selected concentrations of SnCl2 or SnCl4 (control: 0 mg Sn/L, 0.125 mg Sn/L, 1 mg
Sn/L, or 5 mg Sn/L) to provide a wide range of tin exposures (i.e., mg Sn/CFU). To maintain the
same water composition as the undiluted samples, the river water was diluted with filter-sterilized
Mississippi River water. After 1 hr of contact time between the bacteria and the stannous chloride
or stannic chloride, 100 μL aliquots were withdrawn and spread on R2A agar plates. Triplicate
plates were prepared per test condition. The plates were incubated at 30°C and counted after
approximately 24 hr.

Environmental Isolate

A sample of water was collected from the Mississippi River in Minneapolis, MN on July 20,
2008. An aliquot of 100 μL from the sample was withdrawn and plated onto R­2A agar. This plate
was then incubated at 30°C for three days. On the last day of incubation, a single bacterial colony
of a dominant morphology was removed with a flame-sterilized loop and inoculated into 10 mL of
R­2A broth (Difco® recipe without the agar). This solution was placed onto a shaker table at room
temperature overnight. After measuring the OD600 of this solution, a 1 mL aliquot of the solution
was washed and resuspended in lab water (Ultra pure water with 10 mM of NaHCO3). The cell
suspension was then diluted with lab water to target concentrations of 103 and 104 CFU. The diluted
samples were then spiked into solutions with selected concentrations of SnCl2 or SnCl4 (control:
0 mg Sn/L, 0.125 mg Sn/L, 1 mg Sn/L, or 5 mg Sn/L). After 1 h of contact time between the
bacteria and the stannous chloride or stannic chloride solution, 100 μL aliquots were withdrawn
and spread on R2A agar plates. Triplicate plates were prepared per test condition. The plates were
incubated at 30°C and counted after approximately 24 h.
The isolate was identified by amplifying and sequencing the 16S rRNA gene. The gene was
amplified by colony polymerase chain reaction (PCR). The PCR was performed in a final volume
of 50 uL containing H2O (37.35 µL), of PCR buffer (Promega) (10 µL), dNTPs (0.4µL), DNA
primers 27F and 1522R (2 µL), Taq polymerase (Promega) (0.25 µL) and a small patch of cells
from a colony grown on an agar plate. The PCR program was set for 25 cycles. The PCR product
was purified with the GeneClean II kit and was sent for sequencing to the Biomedical Genome
Center. The two sequencing primers used were 338F and 907R.

Continuous-Flow Rotating Disk Reactor

The rotating disk reactor was a simple system for growing biofilm and involves rotating
a disk (typically made of glass) seeded with bacteria in a liquid growth media for a period of
time sufficient to allow a biofilm to develop. A photograph of the setup is included as Figure 3.1.
The entire system was operated in a biosafety cabinet to minimize the risk of contamination. A
round glass disk (2.2 cm diameter) was mounted to the bottom of the spindle of a modified drill
press using stopcock grease (Figure 3.2). For our experiments, the disk was rotated at 104 rpm
approximately 2 mm below the surface of the media. To begin an experiment, the glass disk was
seeded by adding 100 mL of media containing approximately 109 colony forming units (CFU)/mL

©2010 Water Research Foundation. ALL RIGHTS RESERVED


20 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Peristaltic
Pump Press
Drill

Reactor
Waste
Feeding Container
Media

Figure 3.1 Photograph of the continuous-flow rotating disk reactor system

Spindle

Glass Disk

Figure 3.2 Photograph of a round glass disk mounted on to the bottom of the spindle of the
modified drill press in the continuous-flow rotating disk reactor system

of P. aeruginosa PAO1 and then rotating the disk in the bacterial suspension for 1 hr. After 1 hr,
the reactor was replaced with a clean reactor containing 80 mL of 10% LB media and a peristaltic
pump was used to supply 10% LB into the reactor continuously at 18 mL/hour for approximately
72 h. Media was being pumped out at the same rate as the fresh media input so as to maintain a
constant media volume in the reactor. Experiments were performed with media containing 1 mg/L
(as Sn) of SnCl2 and without SnCl2 (control). After 72 hr, the resulting biofilm was scraped off
into a 20 mL PBS solution and homogenized with a tissue homogenizer. Five milliter of the bio-
mass solution was used to enumerate viable bacteria via epifluorescence microscopy magnification
following sample preparation using the Live/Dead Assay (Molecular Probes, Inc., Eugene, OR).
Fluorescent images of viable cells were taken at a total magnification of 1000× using a Nikon E600

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 3: Materials and Methods | 21

Figure 3.3 Glass capillary biofilm reactor system. Biofilms are grown under continuous-flow
conditions. The apparatus consists of a vented medium feed carboy, a flow break, a filtered
air entry, a peristaltic pump, the glass capillary and flow cell holder, and a waste carboy.
These components are connected by flexible silicone tubing (MSU-CBE©, J. Meyer).

microscope. The number of viable cells was counted via these fluorescent images to calculate the
concentration of viable cells grown on the disk.

Continuous-Flow Glass Capillary Biofilm Reactor

A schematic diagram of the continuous-flow glass capillary biofilm reactor system is shown
in Figure 3.3 and photographs of the actual setup are shown in Figures 3.4 and 3.5. Sterile 1% LB
media was prepared without SnCl2 (control) or with SnCl2 (0.125 mg/L, 0.5 mg/L, or 1 mg/L as
Sn+2) adjusted to pH 7.0 with HCl and NaOH in ten liter glass containers. The sterile media was
pumped into autoclaved borosilicate glass capillaries (1 mm × 1 mm × ~10 cm) at 1 mL/min using
a peristaltic pump to condition the capillary and remove air bubbles; which disturbs and prevents
biofilm accumulation. Then, the flow was paused and the capillary was inoculated by injecting
500 mL of a suspension containing ~109 CFU/mL of P. aeruginosa PAO1 using a syringe. The
suspension was allowed to remain in the reactor for 1 hour before the flow was restored. The reac-
tor system was run continuously for 48 h. Images of biofilm development were taken at 0 h, 18 h,
24 h, and 48 h via light microscopy at 40× total magnification using a Nikon E600 microscope.
Two capillaries were run per experiment and each experiment was repeated two times for a total of
4 replicates per condition (control, 0.125 mg/L, 0.5 mg/L, or 1 mg/L as Sn+2).
Additional experiments were performed to investigate the ability of stannous chloride to
remove an established biofilm. For these experiments, the system was run for 48 h without stan-
nous chloride to create a well developed biofilm. Then, the feed was switched to a carboy contain-
ing medium with 1 mg/L stannous chloride as Sn+2. Images were collected during the initial biofilm
development (0 h, 18 h, 24 h, 48 h) and after switching the feed to the media containing stannous

©2010 Water Research Foundation. ALL RIGHTS RESERVED


22 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Microscope

Medium

Flow Cell
Reactor
Peristaltic
pump

Figure 3.4 Photograph of the capillary flow cell system for testing the effects of tin on biofilm
development and removal

(A)   (B)
Figure 3.5 Photograph of the flow cell holder (A; BioSurface Technology®) showing two glass
capillary flow cells connected to flexible tubing. The glass capillaries (B) have a square cross
section (1 mm × 1 mm) allowing direct noninvasive observation of biofilm growing on the
inside. The flow cell holder can hold up to four flow cells as shown.

chloride (72h and 96 h). Control experiments were also performed by operating the system with
stannous chloride-free medium for the entire 96 hours.

EFFECTS OF TIN ON LEAD CORROSION

Lead Coupon Aging

A sheet of lead (6 feet × 6 feet × 1⁄16 inch) was purchased from SPS Companies, Inc. The
sheet was then cut into (3 inch × 0.5 inch × 1⁄16 inch) rectangular coupons. A round hole (diameter
= 1⁄8 inch) was cut near one end to facilitate coupon removal and/or hanging as needed. Lead cou-
pons were cleaned prior to use to remove any surface deposits by the following process: soaking

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 3: Materials and Methods | 23

Table 3.1
Water matrices used for lead coupon aging
Beaker Water type Sn, mg/L Chlorine concentration, mg/L as Cl2
 1 Lab water 0.125 1.0 (free chlorine)
 2 Lab water 0 1.0 (free chlorine)
 3 Lab water 0.125 0
 4 Lab water 0 0
 5 Tap water 0.125 1.0 (free chlorine)
 6 Tap water 0 1.0 (free chlorine)
 7 Tap water 0.125 0
 8 Tap water 0 0
 9 Lab water 0.125 1.0 (monochloramine)
10 Lab water 0 1.0 (monochloramine)
11 Tap water 0.125 1.0 (monochloramine)
12 Tap water 0 1.0 (monochloramine)

in 0.1 M NaOH for ~2 minutes, soaking in 0.1 M HCl for ~2 minutes, rinsing with Milli-Q water
using a squirt bottle, soaking in 95% ethanol, then air drying for a few minutes.
Four lead coupons were placed in a 250 mL glass beaker containing 200 mL of test water.
Twelve beakers were prepared containing different test waters (Table 3.1). Two base waters (“lab
water” and “tap water”) were used and modified to test the effects of chlorine and stannous chlo-
ride on the corrosion rate and surface mineralogy. Lab water was prepared with ultrapure water
(MilliQ system) with 10 mM of NaHCO3 adjusted with HCl and NaOH to pH 8. Tap water was
water taken from SPRWS before the addition of chlorine, chloramine, or stannous chloride; the pH
of this water was approximately 8.3.
The beakers were incubated in the dark at room temperature without mixing. The test
waters in the beakers were prepared and changed weekly for the first 16 weeks and then changed
non-systematically for the next 54 weeks. Temperature and pH of the solutions and the electrical
potential of one coupon from each beaker were monitored before and after each water change.
Electrical potential was measured relative to a standard calomel electrode (SCE) with a Keithley
electrometer using a Accumet probe. At the end of the 70-week aging period, one representative
coupon from each treatment was sacrificed for analysis of surface mineralogy via X-ray diffraction
(XRD, Bruker AXS- D-5005).

Preliminary Corrosion Experiment With New Lead Coupons

A series of nine semi-batch reactors were prepared by placing a new lead coupon into a
250 mL glass bottle containing 200 mL of lab water (10 mM of Na2CO3 at pH 8) amended with 1%
R2A media. The bottles were divided into three sets of triplicate bottles representing different treat-
ments: (1) inoculated with bacteria (No Sn), (2) inoculated with bacteria and spiked with stannous
chloride at 0.125 mg/L as Sn+2 (Sn), and (3) no bacteria or tin (Control). For the inoculated reac-
tors, an undefined mixed culture of Mississippi River bacteria was added at the beginning of the
experiment at a dose of approximately 103 CFU/mL. A plytetrafluoroethylene (PTFE)-coated stir
bar was added to each bottle and the bottles were incubated on stir plates at room temperature for
approximately 9 weeks. The R2A-amended lab water was changed every ten days. During water
changes, approximately 12 mL samples were withdrawn via syringe and filtered through 0.2 µm
syringe-mounted filters for dissolved Pb and dissolved tin analysis, and additional 12 mL samples

©2010 Water Research Foundation. ALL RIGHTS RESERVED


24 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Table 3.2
Experimental matrix for lead corrosion experiments
Sn
Coupon (0.125 mg/L as Sn+2) No Sn Control
label Coupon type Reactor number
A Aged with tap water, Sn, 1  7 13
and free chlorine
B Aged with tap water and Sn 2  8 14
C Aged with tap water 3  9 15
and free chlorine
D Aged with tap water 4 10 16
and monochloramine
E New 5, 6 11, 12 17, 18

were withdrawn for total Pb and total tin analysis. The samples were spiked with acid (5% concen-
trated nitric acid for Pb samples or 10% concentrated HCl for Sn samples) and were analyzed for
tin and Pb within 2 weeks of collection.

Lead Corrosion Experiment With Aged and New Lead Coupons

Eighteen semi-batch reactors were prepared by placing either a new or aged lead coupon
into a 250 mL glass bottle containing 200 mL of lab water (10 mM of NaHCO3 at pH 8) amended
with 1% R2A medium (Table 3.2). The bottles were divided into three sets representing different
treatments: (1) inoculated with bacteria (No Sn), (2) inoculated with bacteria and spiked with stan-
nous chloride at 0.125 mg/L as Sn+2 (Sn), and (3) no bacteria or tin (Control). For the inoculated
reactors, an undefined mixed culture of Mississippi River bacteria was added at the beginning of
the experiment at a dose of approximately 103 CFU/mL. A PTFE-coated stir bar was added to
each bottle and the bottles were incubated on stir plates at room temperature for approximately
8 weeks. The R2A-amended lab water was changed weekly. The control reactors (reactors 13 to
18, Table 3.2) were placed under an ultraviolet (UV) lamp in the biosafety cabinet 2-3 times per
week for more than one hour in an attempt to prevent significant bacterial activity. During water
changes, approximately 12 mL samples were withdrawn via syringe and filtered through 0.2 µm
syringe-mounted filters for dissolved Pb and dissolved tin analysis, and additional 12 mL samples
were withdrawn for total Pb and total tin analysis. The samples were spiked with acid (5% concen-
trated nitric acid for Pb samples or 10% concentrated HCl for tin samples) and were analyzed for
tin and Pb within 8 weeks of collection.

Analytical Methods

Tin

The colorimetric method for analysis of Sn+2 in aqueous samples involves the addition of
toluene dithiolate (TDT) to form a Sn(II)-TDT complex that produces a red color which can be
measured using a ultraviolet-visible light spectrophotometer (Hach) at a wavelength (λ) of 550 nm
(Clark, 1936). To the 10 mL sample, 10 µL of concentrated acid (37%) and 100 µL of TDT solu-
tion (2 g/L) were added. After incubating for 30 minutes at room temperature to allow the color

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 3: Materials and Methods | 25

12

10 y = 28.6x
R2 = 0.998
8
Sn (mg/L)

6
+2

0
0 0.1 0.2 0.3 0.4
Absorbance at 550 nm

Figure 3.6 Calibration curve from the analysis of Sn+2 via the TDT colorimetric method

to develop, the solution was placed in a 1 cm pathlength disposable plastic cuvette and the absor-
bance at 550 nm was measured. A calibration curve was determined for each experiment (e.g.,
Figure 3.6). More details on the method optimization and testing are provided in Appendix A.
Total tin in aqueous samples was analyzed via ICP-AES (Perkin Elmer Optima 3000) at
the University of Minnesota Research Analytical Laboratory or via graphite furnace AAS (Perkin
Elmer 500) in the Department of Civil Engineering. Sn4+ was determined by subtracting the Sn2+
concentration from the total tin concentration.

Lead

Lead concentrations in acidified aqueous samples were analyzed via ICP-AES (Perkin
Elmer Optima 3000) at the University of Minnesota Research Analytical Laboratory or via graph-
ite furnace AAS (Perkin Elmer 500) in the Department of Civil Engineering. For AAS, calibration
standards were prepared by diluting a AAS grade lead stock solution (Perkin Elmer) with 2% nitric
acid. Qualitative assurance/quality control (QA/QC) samples included blanks and standards and
represented 10% of the total number of samples analyzed. All samples were reanalyzed if the QA/
QC samples were not within 5% of the expected values.

Chlorine

Using 10 mL of the monochloramine stock solution to confirm that no free chlorine was
present with the DPD method (Hach), monochloramine in this solution was analyzed using the
N, DPDFAS method (Standard Methods). To determine the concentration of monochloramine,
10 mL of monochloramine solution was first added to 10 mL solutions including 5 mL of N, N
diethyl-p-phylenediamine, 5 mL of phosphorous buffer, and a few drops of potassium iodide.
The resulting color of this solution was dark pink. This solution was then titrated with a ferrous
ammonium sulfate solution until the color of the solution dissipated.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


26 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Lead Surface Mineralogy

The surface mineralogy of the aged lead coupons was analyzed with a wide angle XRD
(Bruker-AXS-D-5005). The diffractometer works at 2.2 kW using Cu as its emission source. A
piece of the aged lead coupons was cut with a razor blade (approximately 1 cm2) and placed into
the diffractometer chamber for reading. After adjusting the generator voltage to 45 kV and current
to 40 mA, the sample was scanned with the following parameters: 2-θ angles from 10° to 90°, 2-θ
step interval of 0.04°, and a dwelling time of 0.8 seconds. The spectra were compared with spectra
from the reference database to identity the minerals present and to quantify the relative % abun-
dance. The matching was restricted to Pb and tin minerals. Analysis of XRD data was done using
Jade software version 8.0.

Lead Surface Morphology

The surface morphology of lead coupons used in the corrosion experiments was investi-
gated using scanning electron microscopy (SEM). For sample preparation, a small piece (approxi-
mately 1 cm × 0.3 cm) was cut from each coupon with a razor blade and then fixed by immersing in
a phosphate buffer saline (PBS) solution containing 2.5% glutaraldehyde and 2.0% formaldehyde
for 2.5 hours. The coupons were then removed and immersed in 25% ethanol and then 50% etha-
nol for 5 minutes each. Then, the coupons were incubated in a solution containing 70% ethanol
overnight followed by 5 minute immersions in 95% ethanol and then 100% ethanol. Afterwards,
the samples were dehydrated via critical point drying with CO2. The samples were then mounted
onto conductive carbon tabs and coated with an approximately 5 nm thick layer of gold using an
argon ion beam coater. Imaging of the coupons was done with a Hitachi S-4700 cold field emission
scanning electron microscope at 3 kV with a working distance of approximately at 5 to 7 mm at the
University of Minnesota Characterization Facility.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


CHAPTER 4
TIN CHEMISTRY

INTRODUCTION

Dissolved tin occurs in the +II oxidation state as stannous ion (Sn+2) and in the +IV oxida-
tion state as stannic ion (Sn+4). The oxidation state of tin is important for understanding the role
of tin in water chemistry and its interactions with the lead surface. For example, in preliminary
experiments we observed that Sn+2 was oxidized to Sn+4 by two oxidants often present in drink-
ing water: dissolved oxygen and free chlorine. Hence, Sn+2 exerted a minor chlorine demand.
Additional research was performed to investigate the reaction of Sn+2 with monochloramine as
described below.

RESULTS AND DISCUSSION

Monochloramine Consumption by Sn2+ 

The consumption of monochloramine by Sn+2 at different pH values is shown in Figures 4.1


to 4.4. A linear regression was performed for each data set and the equation for the line and the
corresponding R2 value are given in each plot. The method detection (MDL) for chloramine using
the DPDFAS method is 0.05 mg/L as Cl2; hence, chloramine concentrations within about a factor
of 2 of the MDL were not included in the linear regressions.
At pH 2, the concentration of monochloramine added was approximately 1 mg/L as Cl2.
The measured monochloramine concentration at a Sn+2 dose of 0 mg/L, however, was approxi-
mately 0.47 mg/L as Cl2. For pH 6, 8, and 9.5, the measured monochloramine concentration at a
Sn+2 dose of 0 mg/L was approximately 1 mg/L as Cl2. This is not surprising as monochloramine is
relatively unstable at low pH. Nevertheless, the slopes of the plots were similar and the mean value
was 0.42 mg of NH2Cl as Cl2 per mg of Sn+2 added. The expected value was 0.59 mg monochlo-
ramine as Cl2/mg Sn2+ according to the following proposed reaction stoichiometry:

Sn2+ + NH2Cl + 2H+ → Sn4+ + NH4+ + Cl– (4.1)

The observed difference suggests that not all of the Sn+2 added was oxidized to Sn4+ during the
reaction. Preliminary experiments performed with free chlorine also resulted in incomplete Sn+2
oxidation except at extremely high chlorine doses, such as 100 mg/L as Cl2.

Oxidation of Sn+2 by Monochloramine

Adding monochloramine to Sn+2 at pH 8 and measured for Sn+2 residuals with TDT sug-
gests an average of 1.30 mg of Sn2+ per mg of NH2Cl (as Cl2) as shown in Figure 4.5. This ratio is
lower than the expected value at 1.70 mg of Sn2+ per mg of NH2Cl based on the assumed reaction
stoichiometry (Equation 4.1).

27

©2010 Water Research Foundation. ALL RIGHTS RESERVED


28 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

1.2

1
NH2Cl residual (mg/L as Cl 2)

0.8
y = -0.415x + 0.44
R2 = 0.914
0.6

0.4

0.2

0
0 0.5 1 1.5 2 2.5 3 3.5 4 4.5
+2
Sn (mg/L)

Figure 4.1 Oxidation of Sn+2 with monochloramine at pH 2. The open symbols were excluded
from the linear regression.

1.2

1
NH2Cl residual (mg/L as Cl 2)

0.8
y = -0.414x + 0.93
R2 = 0.996
0.6

0.4

0.2

0
0 0.5 1 1.5 2 2.5 3 3.5 4 4.5
+2
Sn (mg/L)

Figure 4.2 Oxidation of Sn+2 with monochloramine at pH 6. The open symbols were excluded
from the linear regression.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 4: Tin Chemistry | 29

1.2

1.0
NH2Cl residuals (mg/L as Cl 2)

0.8
y = -0.419x + 0.9808
2
R = 0.976
0.6

0.4

0.2

0.0
0 0.5 1 1.5 2 2.5 3 3.5 4 4.5
+2
Sn (mg/L)

Figure 4.3 Oxidation of Sn+2 with monochloramine at pH 8. The open symbols were excluded
from the linear regression.

1.2

1
NH2Cl residuals (mg/L as Cl 2)

0.8
y = -0.431x + 0.989
R2 = 0.946
0.6

0.4

0.2

0
0 0.5 1 1.5 2 2.5 3 3.5 4 4.5
Sn+2 (mg/L)

Figure 4.4 Oxidation of Sn+2 with monochloramine at pH 9.5. The open symbols were excluded
from the linear regression.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


30 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

A 0.45

0.4

Sn+2 residual (mg/L)


0.35

0.3

y = -1.1467x + 0.3553
0.25 R2 = 0.9825

0.2
0 0.02 0.04 0.06 0.08 0.1 0.12

NH2Cl dose (mg/L)

B 0.45

0.4
Sn+2 residual (mg/L)

0.35

y = -1.42x + 0.308
0.3 R2 = 0.967

0.25

0.2
0 0.02 0.04 0.06 0.08 0.1 0.12

NH2Cl dose (mg/L)

C 0.45

0.4
Sn+2 residual (mg/L)

0.35

0.3 y = -1.34x + 0.415


R2 = 0.9825
0.25

0.2
0 0.02 0.04 0.06 0.08 0.1 0.12

NH2Cl dose (mg/L)

Figure 4.5 Effect of monochloramine dose on the oxidation of Sn+2 at pH 8. Three replicate
experiments shown as plots A, B, C.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 4: Tin Chemistry | 31

10
9 0.70-3.3 °C
8 5.0 -7.89 °C
10.3-12.1 °C
NH2Cl (mg/L as Cl 2)

7
6
5
4
3
2
1
0
0 50 100 150 200 250 300 350

Time (s)

Figure 4.6 Effect of temperature on the reaction rate of monochloramine with Sn+2. The Sn+2
dose and monochloramine (mg/L as Cl2) dose were both approximately 9 mg/L.

Kinetics of Monochloramine Consumption

The consumption of monochloramine was rapid and was complete in less than 10 seconds
for all temperatures tested (Figure 4.6), so the estimated pseudo-first order rate constant (k) is
greater than 227 hr–1. The oxidation of Sn+2 with free chlorine was similarly fast (k > 42 hr–1) while
the oxidation of Sn+2 with dissolved oxygen was much slower (k = 0.069 hr–1). This suggests that:
(1) the chlorine demand of Sn+2 will be virtually instantaneous and (2) kinetic experiments with
chlorine and monochloramine can be performed in room air rather than in an anaerobic glove-bag
without significant interference from reactions with dissolved oxygen.

SUMMARY

The chemistry of tin in drinking water was investigated, including its reactivity with chem-
ical oxidants. Sn+2 was oxidized to Sn+4 by all three of the oxidants investigated: dissolved oxygen,
free chlorine, and monochloramine. Dissolved oxygen oxidizes Sn+2 to Sn+4 at a relatively slow
rate (k = 0.069 hr–1). Both free chlorine and monochloramine oxidize Sn+2 in a matter of seconds
(k > 42 hr–1). The oxidation of Sn+2 is typically not complete, resulting in a mixture of Sn+2 and
Sn+4 in the water. The measured chlorine consumption (0.42 mg of NH2Cl as Cl2 per mg of Sn+2
added) coupled with the low recommended dose for water treatment (≤0.2 mg/L as SnCl2) indi-
cates that use of stannous chloride will result in a minor chlorine demand (<0.053 mg/L as Cl2).

©2010 Water Research Foundation. ALL RIGHTS RESERVED


©2010 Water Research Foundation. ALL RIGHTS RESERVED
CHAPTER 5
TOXICITY OF TIN TO PLANKTONIC AND BIOFILM BACTERIA

INTRODUCTION

Stannous chloride has been shown to be genotoxic to two facultative anaerobic bacteria
(Escherichia coli and Salmonella typhimurium) and the fungus Saccharomyces cerevisiae at con-
centrations in the range of 25 to 75 mM (Dantas et al., 2002; Pungartnik et al., 2005). Additional
research was needed, however, to determine the toxicity of stannous chloride to a wider variety of
bacteria including known biofilm forming organisms (e.g., Pseudomonas aeruginosa) and ammo-
nia oxidizing bacteria (e.g., Nitrosomonas europaea). To our knowledge, all toxicity testing to
date has involved laboratory strains grown planktonically. Research was needed to test the toxic-
ity of stannous chloride to environmental bacteria. Thus, undefined consortia and an isolate from
the Mississippi River were included in our toxicity testing. We also tested the ability of stannous
chloride to decrease the development of biofilms and to remove established biofilms. Finally,
additional experiments were performed to test the toxicity of the stannic ion (Sn+4) because Sn+2 is
oxidized to Sn+4 by chlorine or dissolved oxygen.

RESULTS AND DISCUSSION

Effect of Sn on Bacterial Growth

SnCl2 concentrations up to 0.75 mg/L as Sn (~160 mg Sn/mg biomass) did not have a
significant effect on the growth of P. aeruginosa PAO1 (Figure 5.1A). At a Sn concentration of
1 mg/L (~210 mg Sn/mg biomass), growth was significantly inhibited in comparison to the control,
but the final cell densities were similar. At a concentration of 1.5 mg/L (~310 mg Sn/mg biomass),
however, growth was completely inhibited. Thus, the MIC for P. aeruginosa PAO1 is 1.5 mg/L
(0.013 mM). Interestingly, viable bacteria were recovered from the 1.5 mg/L Sn dose on LB agar
plates throughout the experiment at a nearly constant density of approximately 50% that of the
initial density (1.8 × 104 CFU/mL). Thus, it appears that about half of the inoculated bacteria were
inactivated and the remaining bacteria were unable to grow until they were removed from the Sn
containing solution, diluted, and spread on plates.
The initial N. europaea growth rate was slow (apparent lag phase) for the first 7 days
and then increased rapidly (Figure 5.1B). For Sn concentrations up to 1.0 mg/L (~4 mg Sn/mg
biomass), there were no significant differences in growth rate. When the Sn concentration was
≥2 mg/L (~8 mg Sn/mg biomass), however, the “lag phase” was extended a few more days before
the N. europaea growth rate increased to a similar rate as the other treatments. Also, at the end of
the experiment (20 days), there were no significant differences in bacterial densities as indicated by
OD600 values (Figure 5.1B). Thus, the MIC for N. europaea is greater than 4.4 mg/L (0.037 mM).
Higher SnCl2 concentrations were not investigated because the maximum solubility at circumneu-
tral pH is 4.4 mg/L as Sn. The nitrite results (data not shown) largely mimicked the growth curve
results, except that only the 4.4 mg/L Sn dose (~18 mg Sn/mg biomass) had a significant impact
on nitrite production. As expected, no nitrate was detected in any of the samples, indicating that

33

©2010 Water Research Foundation. ALL RIGHTS RESERVED


34 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

0.4
A
0.3 control
0.25 mg/L Sn
0.5 mg/L Sn
0.75 mg/L Sn
0.2 1.0 mg/L Sn
OD600

1.5 mg/L Sn

0.1

0.0

-0.1
0 10 20 30 40 50

Time, hours

0.07
B
0.06

0.05

0.04
OD600

0.03 control
0.25 mg/L Sn
0.02 0.5 mg/L Sn
1.0 mg/L Sn
0.01 2.0 mg/L Sn
4.4 mg/L Sn
0.00

0 5 10 15 20 25

Time, days

Figure 5.1 Effect of SnCl2 on the growth of P. aeruginosa PAO1 (A) and N. europaea (B).
P. aeruginosa PAO1 and N. europaea were cultured in 10% LB broth and ATCC medium
2265, respectively, and initial cell densities were approximately 4.8 × 10–3 mg/L and
0.25 mg/L, respectively.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 5: Toxicity of Tin to Planktonic and Biofilm Bacteria | 35

the ammonia oxidizing bacterium N. europaea solely converted ammonia to nitrite and no further
nitrification from nitrite to nitrate occurred.

Effect of Sn on OUR

For all treatments and both culture ages (24 and 72 hours) for the P. aeruginosa PAO1
experiments, DO concentrations decreased significantly, indicating that viable bacteria were pres-
ent under all test conditions (Figures 5.2A and 5.2B). The OUR values for the controls were the
greatest of the three treatments and decreased for the Sn and Sn pretreated samples (Table 5.1). The
OUR decreased by 29% (24 hour) and 49% (72 hotur) for the Sn test compared with the respective
Sn-free controls. OUR values for the Sn and Sn pretreated tests for each culture age were similar.
Similar to the P. aeruginosa PAO1 experiments, DO concentrations decreased significantly
for all of the E. coli K-12 experiments, indicating that viable bacteria were present under all test
conditions (Figures 5.2C and 5.2D). In the presence of Sn, the OUR decreased by 86–90% com-
pared with the controls. The OURs for the Sn pretreatment tests, however, were similar to their
respective controls.
Unlike the heterotrophic cultures, the OUR values for the three treatments for the N. euro-
paea culture were similar (Figure 5.2E) despite a relatively high Sn dose of 35.5 mg Sn/mg biomass.
An undefined mixed nitrifying culture was also tested and the results are shown in Figure 5.2F. The
OUR for the undefined mixed nitrifying culture (3.03 mg O2/mg biomass/min) in the absence of Sn
was approximately an order of magnitude greater than that for the N. Europaea culture (0.29 mg
O2/mg biomass/min). The OUR for the mixed culture, however, decreased by 29–36% for the Sn
tests in comparison to the control.
These results suggest that Sn at doses ranging from 10.4–27.3 mg Sn/mg biomass inhibited
the biological activity of P. aeruginosa PAO1 and E. coli K-12 but did not inactivate the bacteria.
The E. coli K-12 were inhibited to a much greater extent than the P. aeruginosa PAO1 at com-
parable Sn doses. Nevertheless, when the Sn was removed in the pretreatment experiments, the
E. coli K-12 quickly recovered in the presence of Sn-free media containing substrate (acetate). The
P. aeruginosa PAO1, however, were unable to recover from the 1 hour Sn exposure. Thus, both
heterotrophs were sensitive to Sn toxicity but the response pattern was different. For the nitrifiers,
N. europaea was relatively resistant to Sn toxicity compared to the heterotrophs and the mixed
nitrifying culture was only moderately affected by the highest biomass normalized Sn doses used
in the experiments (38.0–83.5 mg Sn/mg biomass).

Effect of Sn on Bacterial Inactivation

In the absence of Sn, there was little change in P. aeruginosa PAO1 viable cell density
expressed as colony forming units/mL (CFU/mL) over the 60 minute incubation period for all
experiments (Figure 5.3). The addition of SnCl2 clearly increased the rate of inactivation of P. aeru-
ginosa PAO1, especially for initial cell densities less than 106 CFU/mL. For the same Sn concen-
tration, the rate and extent of inactivation increased with decreasing initial cell density. Extent of
inactivation is defined as follows:

Inactivation = (CFU of control – CFU of test case)/CFU of control

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36 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

7.2 9

7.0 A D
8
6.8

6.6
7
6.4
DO, mg/L

DO, mg/L
6.2 6

6.0
5
5.8
control control
5.6
1mg/L Sn 4 2 mg/L Sn
5.4 1mg/L Sn Pretreated 2 mg/L Sn Pretreated
5.2 3
0 10 20 30 40 50 60 70 0 10 20 30 40 50 60 70

Time, min Time, min


7.0 8.5

6.5
B E
8.0

6.0
7.5
DO, mg/L

DO, mg/L
5.5
7.0
5.0

6.5
4.5
control
1 mg/L Sn control
4.0 6.0
1 mg/L Sn Pretreated 2 mg/L Sn
3.5
2 mg/L Sn Pretreated
5.5
0 10 20 30 40 50 60 70 0 20 40 60 80 100 120 140
Time, min Time, min
5.4 9

5.2
C F
8

5.0 7

4.8 6
DO, mg/L

DO, mg/L

4.6 5

4.4 4

4.2
control 3 control,
4.0 2 mg/L Sn 2 mg/L Sn
2
2 mg/L Sn Pretreated 4.4 mg/L Sn
3.8
1
0 10 20 30 40 50 60 70
0 10 20 30 40 50
Time, min Time, min

Figure 5.2 Effect of SnCl2 on the OUR of P. aeruginosa PAO1 (24 hr, 80.7 mg biomass/L) (A),
P. aeruginosa PAO1 (72 hr, 36.7 mg biomass/L) (B), E. coli K-12 (24 hr, 102 mg biomass/L)
(C), E. coli K-12 (72 hr, 192 mg biomass/L) (D), N. europaea (14 days, 56.4 mg biomass/L) (E),
and an undefined mixed nitrifying culture (52.7 mg biomass/L) (F)

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 5: Toxicity of Tin to Planktonic and Biofilm Bacteria | 37

Table 5.1
Effect of SnCl2 on OUR* of four bacterial cultures
Culture
Mixture
nitrifying
P. aeruginosa PAO1 E. coli K-12 N. europaea culture
Treatment 24 hr 72 hr 24 hr 72 hr 2 weeks
Control 0.52 (0) 0.75 (0) 0.18 (0) 0.36 (0) 0.29 (0) 3.05 (0)
Sn 2 0.37 (12.4) 0.38 (27.3) 0.02 (19.6) 0.03 (10.4) 0.28 (35.5) 1.95 (38.0)
2.17 (83.5)
Sn pretreatment† 0.36 (12.4) 0.30 (27.3) 0.16 (19.6) 0.34 (10.4) 0.29 (35.5) —
*OUR expressed in units of µg O2/mg biomass/min.

Sn dose in mg Sn/mg biomass given in parentheses.

For the lowest cell density (3.69 × 104 CFU/mL) and a SnCl2 concentration of 1 mg/L as Sn, 88%
of the bacteria were inactivated within 5 minutes and 99% were inactivated within 40 minutes
(Figure 5.3A). At the highest initial cell density (1.16 × 107 CFU/mL) and 1 mg/L as Sn, only 17%
were inactivated after 60 minutes (Figure 5.3D). The test water the organisms were suspended in
(i.e., lab water versus tap water) had no effect on the rate of inactivation by SnCl2. The tap water
pH (7.87-8.05) was similar to that for the lab water (8.0), with the most relevant difference being
that the tap water contained natural organic matter (~4 mg/L as C). It was expected that exponential
phase organisms may be more susceptible to inactivation as they are rapidly respiring and pumping
in nutrients. Nevertheless, the results suggest that growth phase did not have a significant effect on
the susceptibility of P. aeruginosa PAO1 to inactivation by Sn. Similar results were obtained for
E. coli K-12 (Figure 5.4).
For N. europaea in lab water, dead bacteria accounted for only 2% and 4% of the total for
Sn doses of 0.125 mg/L and 1 mg/L, respectively after 60 minutes of incubation (data not shown).
In these experiments the initial cell density was 1.8 × 106 /mL, so the biomass-normalized Sn doses
were 6.9 × 10–8 mg/bacterium and 5.6 × 10–7 mg/bacterium, respectively. For comparison purposes,
biomass-normalized Sn doses comparable to the latter dose inactivated approximately 50% of the
P. aeruginosa and E. coli.
From Figures 5.3 and 5.4, it is obvious that the Sn/CFU ratio, not simply the Sn concen-
tration, is the critical parameter for predicting the extent of inactivation. The lower the initial
bacterial concentration for a fixed Sn dose, the more efficiently Sn inactivated the bacteria. Thus,
the amount of Sn taken up by the bacteria is critical for inactivation. At lower initial bacteria con-
centrations, the mass ratio of Sn/bacteria was high, therefore bacteria could take up sufficient Sn
to be inactivated. At high initial bacteria concentrations, the mass ratio of Sn/bacteria was lower,
therefore not all bacteria could take up sufficient Sn to be inactivated.
Because the ratio of Sn to biomass (CFU/mL) dictated the extent of inactivation, log-linear
toxicity plots of fraction of bacteria inactivated versus SnCl2 dose (in mg Sn/CFU) were created
(Figures 5.5 and 5.6). The data for all experiments were included as there was no apparent effect
of culture age or test water on extent of inactivation. The data points represent the average of the
inactivation values at 40 and 60 minutes incubation time as there was little change after 40 min-
utes (Figures 5.3 and 5.4). Both log-linear toxicity plots resembled “S” shaped curves similar to
typical toxicity plots and the data were fit with a five parameter logistic model (Baud, 1993). The
fitting was done by a trial and error method using Microsoft Excel software to minimize the sum

©2010 Water Research Foundation. ALL RIGHTS RESERVED


38 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

A D
1.2 1.2

1.0
1.0

0.8
0.8
0 mg/L Sn
0.6 0.125 mg/L Sn
C/C0

C/C0
1 mg/L Sn 0.6
0 mg/L Sn
0.4
0.125 mg/L Sn
0.4
0.2 1 mg/L Sn

0.2
0.0

0.0
0 10 20 30 40 50 60 70 0 10 20 30 40 50 60 70

Time, min Time, min

B E
1.2 1.2

1.0 1.0

0.8 0.8
0 mg/L Sn 0 mg/L Sn
0.6 0.125 mg/L Sn 0.6 0.125 mg/L Sn
C/C0

C/C0

1 mg/L sn 1 mg/L Sn
0.4 0.4

0.2 0.2

0.0 0.0

0 10 20 30 40 50 60 70 0 10 20 30 40 50 60 70

Time, min Time, min

C
1.2

1.0

0.8 0 mg/L Sn
0.125 mg/L Sn
0.6 1 mg/L Sn
C/C0

0.4

0.2

0.0

0 10 20 30 40 50 60 70

Time, min

Figure 5.3 Effect of Sn dose on the inactivation of P. aeruginosa PAO1 (72 hour culture
age except graph C, 24 hours) over a range of initial biomass concentrations (in CFU/mL):
3.69 × 104 (A), 5.23 × 104 (B), 1.6 × 105 (C), 1.16 × 107 (D), and 1.05 × 105 (E). All graphs for
experiments performed in lab water except graph E which was performed in tap water.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 5: Toxicity of Tin to Planktonic and Biofilm Bacteria | 39

A C
1.2 1.2

1.0 1.0
0 mg/L Sn
0.8
0.125 mg/L Sn 0.8 0 mg/L Sn
1 mg/L sn 0.125 mg/L Sn
0.6 0.6 1 mg/L Sn
C/C0

C/C0
0.4 0.4

0.2 0.2

0.0 0.0

0 10 20 30 40 50 60 70 0 10 20 30 40 50 60 70

Time, min Time, min


B D
1.2 1.2

1.0 1.0

0.8 0.8
C/C0

C/C0

0.6 0.6
0 mg/L Sn
0 mg/L Sn
0.125 mg/L Sn
0.4 0.125 mg/L Sn
1 mg/L Sn 0.4
1 mg/L Sn
0.2 0.2

0.0 0.0
0 10 20 30 40 50 60 70 0 10 20 30 40 50 60 70
Time, min Time, min

E
1.2

1.0

0.8 0 mg/L Sn
0.125 mg/L Sn
0.6 1 mg/L Sn
C/C0

0.4

0.2

0.0

0 10 20 30 40 50 60 70

Time, min

Figure 5.4 Effect of Sn dose on the inactivation of E. coli K-12 (24 hour: A,B; 72 hour: C–E)
over a range of initial biomass concentrations (in CFU/mL): 9.2 × 104 (A), 1.63 × 107 (B),
6.7 × 104 (C), 3.2 × 107 (D), and 1.26 × 105 (E). All graphs for experiments performed in lab
water except graph E which was performed in tap water.

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40 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

100%

90%

80%

70%

60%
Inactivation
50%

40%

30%

20%

10%

0%
-9 -8 -7 -6 -5 -4 -3
log (dose)

Figure 5.5 Inactivation of P. aeruginosa PAO1 as a function of Sn dose in µg Sn/CFU. Data


points (filled circles) were fit with a five parameter logistic model (solid line).

100%

90%

80%

70%

60%
Inactivation

50%

40%

30%

20%

10%

0%
-9 -8 -7 -6 -5 -4 -3
log (dose)

Figure 5.6 Inactivation of E. coli K-12 as a function of Sn dose in µg Sn/CFU. Data points
(filled circles) were fit with a five parameter logistic model (solid line).

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 5: Toxicity of Tin to Planktonic and Biofilm Bacteria | 41

of squared errors. LD50 values were calculated by entering an inactivation of 0.5 into the best fit
equations and solving for the Sn dose. The LD50 for P. aeruginosa PAO1 (10–6.24 mg Sn/CFU) was
similar to that for E. coli K-12 (10–6.25 mg Sn/CFU).
Stannous chloride increased the inactivation of P. aeruginosa PAO1 and E. coli K-12 in
the absence of electron donor and the extent of Sn inactivation was a function of the Sn to bacte-
ria ratio. The observed dependence of inactivation on the biomass normalized dose is significant
because most bacterial toxicity studies in the literature simply report toxicity values in concentra-
tion units (e.g., mM). In order to compare toxicity of metals or other chemicals reported in the
literature, we recommend normalizing the dose by the initial biomass concentration.
Dantas et al. (1996) reported the rapid inactivation (~90% in 40 minutes) of E. coli K12
AB1157 (~108 cells/mL) in 0.9% NaCl containing 37 mM SnCl2 (4.4 mg/L as Sn2+). The estimated
dose of 4.4 × 10–8 mg/CFU resulted in a much greater inactivation than would be predicted from
Figures 5.5 and 5.6 (~10%), suggesting that the strain used in the Dantas et al. (1996) study was
more sensitive to Sn toxicity. The toxicity of SnCl2 was attributed to oxidative stress and DNA
damage resulting from the production of reactive oxygen species inside the cell in a Fenton-like
reaction (Dantas et al., 1996; Dantas et al., 1999; Dantas et al., 2002; Pungartnik et al., 2005). It
is surprising, therefore, that the E. coli K12 AB1157 would be sensitive to Sn toxicity because the
strain is known to be DNA repair proficient.
The toxicity of Sn is similar to or greater than other heavy metals, with the possible excep-
tion of Hg. For example, the Zn, Pb, and Cu MBCs for planktonic P. aeruginosa PAO1 (Teitzel
and Parsek, 2003) expressed on a biomass normalized basis are 10–3.9, 10–4.3, and 10–6.2 mg/CFU.
The LD50 values were not reported but are estimated to be approximately an order of magnitude
lower. Certainly caution must be exercised in comparing metal toxicity results between studies
as differences in solution composition (e.g., pH) can alter metal speciation and have a significant
impact on toxicity. Nevertheless, by comparison to reported literature values, it appears that SnCl2
is highly toxic to some lab strains of heterotrophic bacteria. This suggests a potential beneficial use
of this chemical for controlling bacterial regrowth in distribution systems. Given that viable bacte-
rial numbers in water distribution systems are typically very low (<100 CFU/mL) and that organ-
isms are often stressed by the presence of residual chlorine, it is likely that the addition of SnCl2 to
water at concentrations in the range of 0.1 to 0.2 mg/L as Sn has a significant effect on the viability
of heterotrophic bacteria. In fact, we previously reported that the addition of SnCl2 significantly
reduced heterotrophic plate count levels in comparison to a no Sn control in chloraminated pipe
loop experiments (Hozalski et al., 2005). Although only limited data with nitrifying bacteria were
obtained to date because of the relatively slow growth of these organisms, it appears that nitrifiers
are more resistant to Sn2+ toxicity than heterotrophic bacteria.

Effect of Sn on Inactivation of Undefined Mixed Cultures From the Mississippi River

The ��������������������������������������������������������������������������������������
concentration�������������������������������������������������������������������������
of viable planktonic Mississippi River water heterotrophic bacteria col-
lected on April 19, 2008 decreased with increasing concentration of SnCl2 as shown in Figure 5.7.
The extent of inactivation is plotted versus the log of normalized SnCl2 dose expressed as mg Sn/
CFU in Figure 5.8. The batch inactivation data were fitted with a straight line to yield a LD50 value
of 10–2.47 mg Sn/CFU. This LD50 value is almost four orders of magnitude greater than the values
obtained for lab strains of P. aeruginosa and Escherichia coli (10–6.24 and 10–6.25, respectively),
suggesting that the river water bacteria are much more resistant to Sn toxicity than the lab strains.
If the data from all four river sampling dates are combined, however, no correlation is observed

©2010 Water Research Foundation. ALL RIGHTS RESERVED


42 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

1600

1400 Undiluted
10X diluted
1200

1000
CFU/mL

800

600

400

200

0
0 1 2 3 4 5 6
Sn+2 (mg/L)

Figure 5.7 Effect of SnCl2 concentration on the inactivation of planktonic Mississippi River
water bacteria. Data points represent the average of triplicate plates. Water sample collected
on April 19, 2008.

80%

70%

60%

50%
Inactivation

40% y = 0.2299x + 1.0692


R2 = 0.8407
30%

20%

10%

0%
-4.5 -4 -3.5 -3 -2.5 -2 -1.5 -1 -0.5 0
Log Dose (µg/CFU)

Figure 5.8 Inactivation of planktonic Mississippi River water bacteria as a function of SnCl2
dose in µg Sn/CFU. Water sample collected on April 19, 2008.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 5: Toxicity of Tin to Planktonic and Biofilm Bacteria | 43

100%

80% Sn+2
Sn+4
60%
Linear (Sn+2)
40% Linear (Sn+4)

20%
Inactivation

y (Sn+2) = 0.0152x + 0.268


0% R2 = 0.0012

-20%
y (Sn+4) = 0.0748x + 0.523
-40% R2 = 0.0407

-60%

-80%
-5 -4 -3 -2 -1 0
Log Sn Dose (μg/CFU)

Figure 5.9 Inactivation of planktonic Mississippi River water bacteria as a function of


SnCl2 or SnCl4 dose in µg Sn/CFU. Compilation of data from experiments on water samples
collected on April 19, 2008, June 17, 2008, June 23, 2008, and June 26, 2008.

(Figure 5.9). One possible explanation for the lack of correlation for the lumped data from four
sampling events is that the microbial community composition (and Sn resistance) changes signifi-
cantly with each sampling. The variability could also be attributed to changes in water chemistry
that affected the complexation and speciation of the Sn. Also, the significant amount of scatter in
the results is attributed to the inherent complexity of mixed microbial communities obtained from
the environment. Despite the significant scatter, most samples exhibited a positive inactivation.
These results indicated that stannous chloride is toxic to some bacterial species in the river water.
Additionally, the data in Figure 5.9 suggest that the toxicity of Sn+4 and Sn+2 are similar.

Effect of Sn on Inactivation of an Isolate From the Mississippi River

The toxicity of Sn+2 and Sn+4 to an environmental isolate obtained from the Mississippi
River in Minneapolis (Acinetobacter sp.) is shown in Figures 5.10 and 5.11. The plots showed
that tin had very little effects on Acinetobacter sp. viability. Significant scatter was observed in
the inactivation versus exposure data (Figure 5.12). Relative to the reference, approximately 50%
of the samples exhibited some inactivation while approximately 50% exhibited no inactivation or
even negative inactivation (i.e., growth). This suggested that Acinetobacter sp. isolated was resis-
tant to Sn+2 and Sn+4. According to the literature, Acinetobacter sp. is a common soil bacterium
that is resistant to many common antibiotics such as penicillin and chloramphenicol (Rahal, 2006;
Gerischer�����������������������������������������������������������������������������������������
,����������������������������������������������������������������������������������������
2008). In addition, some bacteria belonging to the Acinetobacter genus are also consid-
ered resistant to heavy metals (Prasad, 2001).

©2010 Water Research Foundation. ALL RIGHTS RESERVED


44 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

2500 10^3 CFU/mL


10^2 CFU/mL

2000

1500
CFU/mL

1000

500

0
0 1 2 3 4 5 6
Sn+2 (mg/L)

Figure 5.10 Effect of stannous chloride concentration on the inactivation of planktonic


Acinetobacter sp. at initial biomass densities of ~103 CFU/mL and ~102 CFU/mL. Experiment
performed on July 25, 2008.

Preliminary Biofilm Experiment With Continuous-Flow Rotating Disk Reactor

P. aeruginosa PAO1 biofilm developed as white colonies primarily on the periphery of the
disks (Figure 5.13). The consistent color and morphology of the colonies suggested that contami-
nation of the system was not a problem. The color and morphology of the biofilms for the tests with
and without SnCl2 were similar, however, the colony size for the biofilms grown without SnCl2
appeared to be larger (Figure 5.13).
The biomass was scraped from the surface of the disks, collected on filters, stained with
the LIVE/DEAD reagents, and counted via epifluorescence microscopy. The images contained
only rod-shaped cells characteristic of P. aeruginosa (Figure 5.14), again suggesting that little or
no contamination occurred during the experiment. The viable cells were counted to determine the
viable cell counts per area of disk as shown below:

Number of cells = Average cell # Total Sample Volume area of filter (cm 2)
cm 2 area of field(cm 2)
× Aliquot Sample Volume × area of glass disk (cm 2)
The limited data suggested that there was no difference between the viable cell counts obtained
for biofilm grown in the presence of Sn (1 mg/L as Sn+2) versus the control without Sn (Table 5.2).
The lack of effect of stannous chloride on biofilm development was likely due to the high
nutrient load applied to the system (10% LB). The rate of biofilm accumulation is a function of
biofilm growth, biofilm inactivation, and physical removal (shear loss and sloughing). At high sub-
strate concentrations, the growth rate should be at or near a maximum value such that an increase

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 5: Toxicity of Tin to Planktonic and Biofilm Bacteria | 45

2500 10^3 CFU/mL


10^2 CFU/mL
2000

1500
CFU/mL

1000

500

0
0 1 2 3 4 5 6
Stannic chloride (mg/L)

Figure 5.11 Effect of stannic chloride concentration on the inactivation of planktonic


Acinetobacter sp. at initial biomass densities of ~103 CFU/mL and ~102 CFU/mL. Experiment
performed on July 25, 2008.

40%
Sn+2
20% Sn+4
Linear (Sn+2)
0% Linear (Sn+4)
Inactivation

-20%

y(Sn+2) = -0.0785x - 0.277


-40%
R2 = 0.0617

-60% y(Sn+4) = 0.0333x + 0.178


R2 = 0.0321
-80%
-4.5 -4 -3.5 -3 -2.5 -2 -1.5 -1 -0.5 0
Log Dose (μg/CFU)

Figure 5.12 Inactivation of planktonic Acinetobacter sp. as a function of SnCl2 or SnCl4 dose
in µg Sn/CFU. Compilation of data from experiments performed on July 22, 2008, July 25,
2008, and August 5, 2008.

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46 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

No Sn 1 mg/L as Sn+2

Figure 5.13 Photograph of P. aeruginosa PAO1 biofilm grown on rotating glass disks in media
without SnCl2 (left) and with SnCl2 (right)

Figure 5.14 Image of viable P. aeruginosa PAO1 cells from the rotating disk reactor experiment
taken via epifluorescence microscopy

in inactivation rate could be masked. Ultimately there is a tradeoff between using higher substrate
concentrations to minimize the time required to obtain quantifiable biofilm levels and operating
under the oligotrophic conditions that are representative of distribution systems. Thus, the media
concentration was decreased to 1% for the glass capillary reactor experiments.

Continuous-Flow Glass Capillary Biofilm Reactor

Effect of Stannous Chloride on Biofilm Development

Representative images from the glass capillary biofilm reactors (Table 5.3) clearly illus-
trate the inhibition of accumulation of P. aeruginosa PAO1 biofilm with increasing concentration
of stannous chloride (See Appendix B for complete set of images). Stannous chloride slowed
biofilm development in some experiments at doses as low as 0.125 mg/L (Table 5.3). When the

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 5: Toxicity of Tin to Planktonic and Biofilm Bacteria | 47

Table 5.2
Viable cell counts of P. aeruginosa PAO1 biofilm developed from continuous
rotating disk reactor experiment with and without SnCl2
Sn+2 (mg/L) Viable cell counts (cells/cm2)
0 5.02 × 105 (n =1)
1 6.11 × 105 ± 5.28 × 105 (n = 3)
n = number of samples.

concentration was increased to 0.5 mg/L, biofilm formation was severely inhibited and completely
prevented in most experiments (Table 5.3 and Appendix B). Differences in biofilm morphology
were also observed, as biofilms grown in the absence of stannous chloride tended to form larger
colonies than when grown in the presence of stannous chloride (Table 5.3). This observation is in
agreement with the aforementioned results from the rotating disk reactor experiments.
To quantify the accumulation of P. aeruginosa PAO1 biofilm in the capillaries, bright-
ness histograms were obtained using ImageJ software. One histogram was created for each of
the biofilm images taken at specific times (Figure 5.15). The brightness values were converted to
gray scale values by subtracting each brightness value from the maximum brightness value of 255
(Figure 5.16). The mean gray scale value was then calculated for each image and the mean initial
or time zero value for the time series was subtracted from each mean value in the series to obtain
a differential gray scale value (Dgray scale value). Thus, the Dgray scale value would directly
indicate the extent of biofilm development, being zero at time zero and increasing as biofilm accu-
mulates in the capillary.
The mean Dgray scale values from all experiments were plotted versus time (Figure 5.17).
Stannous chloride had little effect on biofilm development at a concentration of 0.125 mg/L as
Sn in these experiments (1% LB media). At time = 48 hours, the mean Dgray scale values for the
control and 0.125 mg/L experiments were not significantly different at the 95% confidence level.
At a concentration of 0.5 mg/L as Sn, stannous chloride severely inhibited biofilm development
(Figure 5.17). In fact, biofilm development was completely inhibited for most runs with 0.5 mg/L
as Sn in the media and the mean Dgray scale values were not significantly different from zero at all
time points. At time = 48 hours, the mean Dgray scale values for the control and 0.5 mg/L experi-
ments were significantly different at the 95% confidence level.

Removal of Established Biofilms With Stannous Chloride

Stannous chloride was shown to inhibit P. aeruginosa PAO1 biofilm accumulation in


the capillary flow cell experiments discussed above. Additional experiments were performed to
determine if stannous chloride could remove established biofilms (Table 5.4). The capillary flow
cell reactors that were run with feeding media continuously without stannous chloride showed
continuous increase of biofilm accumulation over 96 hours (Table 5.4, Figure 5.18). When the feed
was changed to a media containing 1 mg/L of stannous chloride at time = 48 hours, the biofilm
decreased with time (Figure 5.18).
The first stage in biofilm formation is the attachment of planktonic or sessile bacteria to
the surface. The ability of stannous chloride to inhibit the development of P. aeruginosa PAO1
biofilm in the continuous-flow glass capillary reactor experiments is consistent with the observed
toxicity of stannous chloride to planktonic or sessile P. aeruginosa PAO1 discussed above. The
second stage of formation is colonization, where the attached bacteria begin to grow and produce

©2010 Water Research Foundation. ALL RIGHTS RESERVED


48 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Table 5.3
Photographs of glass capillary biofilm reactors showing the effect of SnCl2 on the
accumulation of P. aeruginosa PAO1 biofilm
Sn+2
(mg/L) 0h 24 h 48 h

0.125

0.5

EPS. Guibaud et al. (2008) suggested that EPS has a sorption affinity for heavy metals. Hence, the
EPS may provide some protection to the cells in the biofilm, allowing the biofilm to accumulate if
the concentration of Sn+2 is low. When the concentration of Sn+2 is sufficiently high (≥0.5 mg/L as
Sn in these experiments), then biofilm development can be severely inhibited and established bio-
films can be removed. The actual concentration required to inhibit biofilm development in water
distribution systems will depend on a number of factors including the organisms present and their
resistance to Sn toxicity, the amount of growth substrate available, the water temperature, and the
concentration and type of disinfectant. A stannous chloride dose of 0.175 mg/L as SnCl2 was suf-
ficient to significantly reduce heterotrophic plate count levels relative to an untreated control in
a pipe loop system fed treated water containing chloramines at ~2.5 mg/L as Cl2 (Hozalski et al.
2005).

SUMMARY

Sn2+ inhibited the growth of Pseudomonas aeruginosa PAO1 and Nitrosomonas europaea
and inhibited the aerobic respiration of P. aeruginosa PAO1, Escherichia coli K-12, N. europaea
and a mixed culture of nitrifying bacteria in the presence of excess electron donor. The Sn2+ MICs

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 5: Toxicity of Tin to Planktonic and Biofilm Bacteria | 49

25,000
0h
18 h
20,000 24 h
48 h

15,000

10,000

5,000

0
0 50 100 150 200 250

Figure 5.15 A time series of brightness histograms from a capillary flow reactor showing the
development of a P. aeruginosa PAO1 biofilm grown in the absence of stannous chloride

25,000
0h
18 h
20,000 24 h
48 h

15,000
Counts

10,000

5,000

0
0 50 100 150 200 250

Figure 5.16 A time series of gray scale histograms derived from the brightness histograms in
Figure 5.15

©2010 Water Research Foundation. ALL RIGHTS RESERVED


50 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

40

35
Control
0.125 mg/L Sn
30
∆ Gray Scale Value 0.5 mg/L Sn
25

20

15

10

0
0 18 24-26 48

Tim e (h)

40

35
Control
0.125 mg/L Sn
30
∆ Gray Scale Value

0.5 mg/L Sn
25

20

15

10

0
0 18 24-26 48
Time (h)

Figure 5.17 Effect of stannous chloride on the accumulation of P. aeruginosa PAO1 biofilms
in capillary flow cells. (A) Data for all experiments included (number of experiments = n = 7
for Control, n = 11 for 0.125 mg/L Sn, and n = 13 for 0.5 mg/L Sn). (B) Same as A except data
for two outlier experiments on 6/16/08 for 0.5 mg/L Sn were eliminated. Error bars represent
one standard deviation from the mean value.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 5: Toxicity of Tin to Planktonic and Biofilm Bacteria | 51

Table 5.4
Photographs of glass capillary biofilm reactors showing the effect of SnCl2
on the removal of an established P. aeruginosa PAO1 biofilm
Sn+2
(mg/L) 48 h 73 h 96 h

80

70 Control
0 to 1mg/L Sn at 48 hours
60
∆ Gray Scale Value

50

40

30

20

10

0
0 23-24 48 56 71-73 96
Time (h)

Figure 5.18 Comparison of biofilm accumulation in capillary flow cells when the feed is
switched to media containing stannous chloride (1 mg/L as Sn, n = 3) after 48 hours versus
a control (no Sn, n = 2). Error bars represent one standard deviation from the mean value.

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52 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

for P. aeruginosa PAO1 and N. europaea were 1.5 mg/L (0.013 mM) and > 4.4 mg/L (0.037 mM),
respectively. Sn2+ increased the inactivation of P. aeruginosa PAO1 and E. coli K-12 in the absence
of electron donor and the extent of Sn inactivation was a function of the Sn to biomass ratio. LD50
values for P. aeruginosa PAO1 and E. coli K-12 were 10–6.24 and 10–6.25 mg Sn/CFU, respectively.
Thus, SnCl2 is highly toxic to lab strains of heterotrophic bacteria and is similar to or more toxic
than other heavy metals, with the possible exception of Hg. The observed toxicity of SnCl2 sug-
gests that the chemical may aid in controlling bacterial regrowth in distribution systems. Although
only limited data with nitrifying bacteria were obtained because of the relatively slow growth of
these organisms, it appears that nitrifiers are more resistant to Sn2+ toxicity than heterotrophic
bacteria. The environmental bacteria appeared to be more resistant to stannous chloride toxicity.
There was no significant increase in inactivation of the environmental isolate Acinetobacter sp.
Stannous chloride, however, inhibited the accumulation of P. aeruginosa biofilm at a concentra-
tion of 0.5 mg/L as Sn and was able to remove established P. aeruginosa biofilm at 1 mg/L as Sn.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


CHAPTER 6
EFFECT OF STANNOUS CHLORIDE ON LEAD CORROSION

INTRODUCTION

In Chapter 5, the toxicity of tin to planktonic and biofilm bacteria was demonstrated.
Stannous chloride was highly toxic to laboratory strains of heterotrophic bacteria and considerably
less toxic to ammonia oxidizers and environmental bacteria. The stannic ion, formed from the oxi-
dation of stannous ion as discussed in Chapter 4, also exhibited some bacterial toxicity in limited
testing. These results suggested that stannous chloride could decrease lead corrosion by inhibiting
bacteria activity and MIC. Thus, experiments were performed using new and aged lead coupons
in semi-batch reactors to investigate whether MIC of lead occurs under these test conditions and if
stannous chloride can mitigate MIC.

RESULTS AND DISCUSSION

Potential Measurements During Coupon Aging

The lead coupons were aged for 70 weeks. During the first 16 weeks, the solutions in the
reactors were changed regularly (every 7 to 10 days) and the reactors were monitored for pH, tem-
perature, and lead coupon electrical potential. After week 16, the reactor solutions were changed
less frequently and monitored less frequently.
Temperature remained relatively constant at approximately 23–24 °C during the aging
period, except a modest temperature spike during weeks 2 to 5 (Figure 6.1). This temperature spike
was due to air conditioning issues in the building. The pH of the solutions amended with SnCl2
were less than the target pH of 8 during the first 4 weeks because the acidic SnCl2 stock solution
caused the pH to decrease (Figure 6.2). Additional base was added from week 5 onward to correct
this problem (Figure 6.2). The pH typically increased between water changes from 8 to 8.5 for the
lab water reactors (e.g., Figure 6.2) and from 8 to 8.3 for the tap water reactors.
The electrical potential of the lead coupons increased in the first few weeks, especially
for the lab water conditions, suggesting that the surface was aging and a passive layer was form-
ing resulting in a reduced corrosion rate (Figure 6.3). After 8 weeks of incubation, the potentials
appear to have stabilized at between –0.2 and –0.3 mV for the lab water conditions and –0.3 to
–0.4 mV for the tap water conditions (Figure 6.4). The lead coupons in lab water with free chlorine
addition appear to have reached a stable potential value more rapidly, suggesting that free chlorine
helped to oxidize the lead surface to more quickly form a passivation layer. Free chlorine had less
of an impact in the tap water reactors, likely because the normal organic matter (NOM) in the tap
water consumed some of the chlorine. Sn addition did not have a significant effect on the potential
of the lead surface, regardless of water quality. Finally, the potentials in tap water were compara-
tively lower than the potentials in lab water, suggesting that the tap water is more corrosive than
the lab water.

53

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54 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

26.5

26.0

Temperature, degrees C 25.5

25.0

24.5

24.0

23.5

23.0

22.5
0 2 4 6 8 10 12 14 16 18
Time, weeks

Figure 6.1 Water temperature during lead coupon aging

8.8

8.6

8.4

8.2

8.0
pH

7.8

7.6 before change


after change
7.4

7.2
0 2 4 6 8 10 12 14
Time, weeks

Figure 6.2 pH as a function of time for the reactor containing lab water amended with 1 mg/L
as Cl2 free chlorine and 0.125 mg/L Sn+2. Before change refers to measurements made just prior
to replacing the test water and after change refers to immediately after the test water was replaced.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 6: Effect of Stannous Chloride on Lead Corrosion | 55

A B
0 0

-0.1 -0.1

-0.2 -0.2

Potential, mv
Potential, mv

-0.3 -0.3

-0.4 -0.4

before change
-0.5 -0.5 before change
after change
after change
-0.6 -0.6
0 2 4 6 8 10 12 14 16 18 0 2 4 6 8 10 12 14 16 18

Time, weeks Time, weeks

C D
0 0

-0.1 -0.1
Potential, mv

-0.2
-0.2
Potential, mv
-0.3
-0.3

-0.4
-0.4

-0.5 before change


-0.5 before change
after change
after change
-0.6
0 2 4 6 8 10 12 14 16 18 -0.6
0 2 4 6 8 10 12 14 16 18
Time, weeks
Time, weeks

Figure 6.3 Electrical potential vs. SCE for lead coupons incubated in lab water under the
following conditions: Control (A), 0.125 mg/L Sn2+ (B), 1 mg/L as Cl2 free chlorine (C), 1 mg/L
as Cl2 free chlorine and 0.125 mg/L Sn2+ (D). Before change refers to measurements made just
prior to replacing the test water and after change refers to immediately after the test water was
replaced.

Mineralogy of the Aged Lead Coupons

In comparison to the uniform gray color of the new lead coupons, the aged coupons exhib-
ited significant coloration attributed to accumulation of surface deposits. Lead coupons aged in
lab water were generally reddish purple in color while the lead coupons aged in tap water were
generally greenish and gray in color (Figure 6.5). The color of lead (II) oxide can be red (litharge)
or yellow (massicot). White deposits were also common on the surface of many of these coupons.
Cerussite is colorless or white in color, sometimes with a grey or greenish tint and hydrocerussite
is colorless, gray, or white. Plattnerite is brown-black or black in color. The presence of chlorine or
tin during aging seemed to have little effect on the visual appearance of the aged coupons.
Lead minerals on the coupons and their respective percentage compositions determined
by XRD analysis are given in Table 6.1. An example XRD spectrum is shown in Figure 6.6 (See
Appendix C for the complete set of XRD spectra of the aged coupons). The minerals observed
included: lead (II) oxide, cerussite, and/or hydrocerussite. Plattnerite was detected at low levels in

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56 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

A
before after before after before after before after
0
1 2 3 4 5 6 7 8
-0.05

-0.1
Mean potential (mV)

-0.15

-0.2

-0.25

-0.3

-0.35

-0.4
Sn + Cl2 Cl2 only Sn only Control
-0.45
Sample

B
before after before after before after before after
0
-0.05 1 2 3 4 5 6 7 8

-0.1
Mean potential (mV)

-0.15
-0.2
-0.25
-0.3
-0.35
-0.4
-0.45
Sn + Cl2 Cl2 only Sn only Control
-0.5
Sample

Figure 6.4 Mean potential measurements from weeks 12 to 16 (n = 5) for lead coupons aged
in lab water (A) and tap water (B). Error bars represent one standard deviation from the mean
values. Before refers to measurements made just prior to replacing the test water and after refers to
immediately after the test water was replaced.

a few samples where free chlorine was added (Table 6.1). The reported value of 37% plattnerite
for the lab water only sample is suspect, as it is not possible for this mineral to form in the absence
of a strong oxidant such as free chlorine. Unfortunately, the data analysis was not completed until
several weeks after the samples were analyzed and we were unable to rerun the sample. No tin-
containing minerals were found on the surface of any of the coupons, which is consistent with
findings from the pipe loop study (Hozalski et al. 2005). Similarly, SPWRS did not detect any tin-
containing minerals on the interior surfaces of lead service connections removed from the water
distribution system after more than five years of stannous chloride usage (data not shown).

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 6: Effect of Stannous Chloride on Lead Corrosion | 57


Figure 6.5 Photos of lead coupons aged for approximately 70 weeks in lab (left) or SPRWS
tap water (right)

Table 6.1
Approximate mineralogical composition on the surface of lead coupons after 70 weeks of
aging in lab or tap water as determined by XRD analysis
Cerussite Plattnerite Hydrocerussite
Coupon Lead (Pb0) PbO (PbCO3) (PbO2) (Pb3(CO3)2(OH)2­)
Reference 100% — — — —
Cl2-LW — 75% 20% 5.4% —
Cl2-TW — 78% 3.2% — 19%
LW only — 63% — 37% —
TW only 17% 54% 29% — —
NH2Cl-LW 14% 58% 25% — 4.4%
NH2Cl-TW 14% 66% 0.2% — 20%
Sn-LW — 44% 56% — —
Sn-TW 19% 59% 2.7% — 20%
Sn+Cl2 -LW — 83% 2.0% — 15%
Sn+Cl2- TW 20% 51% 27% 0.2% 1.7%
Sn+NH2Cl-LW 1.6% 68% 21% — 9.2%
Sn+NH2Cl-TW 29% 45% 20% — 5.3%
LW = Lab Water, TW = Tap Water

Finally, the surface morphology of one of the coupons aged in lab water was examined
using SEM (Figure 6.7). A new lead coupon was also examined as a reference (Figure 6.8). As
expected, the surface of the new coupon was much smoother than the surface of the aged cou-
pon. Surprisingly, bacterial communities and individual bacteria were observed on the surface of
the aged lead coupon (Figures 6.7). No nutrients or organic carbon were added to the lab water
used to age the coupons, so in theory there was little available to sustain bacteria growth. Also,
lead is known to be toxic to bacteria. Perhaps the nutrient deficiency explains the lack of EPS
characteristic of biofilms growing in nutrient-rich environments. It is interesting to note that

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58 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure 6.6 Examples of spectra from the XRD analysis of a new lead coupon (A) and a lead
coupon aged in lab water with free chlorine (1 mg/L as Cl2) and SnCl2 (0.125 mg/L Sn) (B).
Example reference spectra are shown at the bottom of each figure.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Chapter 6: Effect of Stannous Chloride on Lead Corrosion | 59

(continued)
Figure 6.7 SEM images of the surface of a lead coupon aged in lab water with free chlorine
(1 mg/L as Cl2) and SnCl2 (0.125 mg/L Sn)

commonly-observed rod-shaped and spherical (i.e., cocci) bacteria were observed along with the
less commonly observed spirochetes (Figure 6.7C). Unfortunately, the roles of these bacteria in
lead corrosion, if any, cannot be discerned from a SEM image.

Preliminary Lead Corrosion Experiment With New Lead Coupons

As expected, the concentrations of dissolved Pb release from new lead coupons decreased
with time (Figure 6.9). Also, the results were similar regardless of the treatment (i.e., control, Sn,

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60 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure 6.7 (Continued)

Figure 6.8 SEM image of the surface of a new lead coupon

or no Sn). The Sn reactor, however, does appear to be trending upward over the last 3 samples,
suggesting that the Sn might be interfering with passivation of the lead surface. To evaluate the
role of MIC, the Sn and No Sn reactors were inoculated with Mississippi River water bacteria
while the Control was not inoculated. The Control reactors, however, appeared to become con-
taminated with bacteria (i.e., cloudy) even though the reactor water was changed inside a biosafety
cabinet. Thus, it is not surprising that the Control and No Sn dissolved lead results were similar.
The dissolved lead levels were typically between 100 and 400 mg/L after day 20. Such lead levels

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Chapter 6: Effect of Stannous Chloride on Lead Corrosion | 61

1.2

1 Sn No Sn Control
Dissolved Pb (mg/L)

0.8

0.6

0.4

0.2

0
0 10 20 30 40 50 60 70
Time (Day)

Figure 6.9 Mean concentration of dissolved Pb (n = 2) versus time in the preliminary lead
corrosion experiment

are approximately consistent with lead solubility in the presence of lead carbonate minerals and
exceed the lead AL by a factor of 10 or more.

Lead Corrosion Experiment With Aged and New Lead Coupons

Additional lead corrosion experiments were performed with the aged lead coupons. New
lead coupons were also included to repeat the preliminary experiment. Similar to the prelimi-
nary experiment, dissolved lead levels decreased initially and then stabilized after approximately
15  days (Figure 6.10). It was expected that the aged coupons would not exhibit the significant
initial decrease observed for the new lead coupons. One possible explanation for this behavior is
some disturbance to the corrosion products on the coupons during transfer to the new reactors. The
lead levels were between ~200 and 500 mg/L after day 15, which are in approximate agreement
with lead carbonate solubility. Again, there was no improvement in dissolved lead levels with the
Sn treatment. In fact, lead levels for the reactors containing Sn were typically greater than those for
the No Sn and Control reactors. This is contradictory to the results from the pipe loop experiments
(Hozalski et al., 2005), where lead levels decreased in comparison to an untreated control. The
difference in results from these two studies are likely due to differences in the chemical composi-
tion of the waters, as the pilot-scale experiments used treated water from the full-scale plant which
contained natural organic matter and a chloramine residual.
The total lead levels were much more erratic than the dissolved lead levels and did not
exhibit a consistent trend (Figure 6.11). Total lead concentrations generally varied between 0.4
and 2.0 mg/L. Again, the results for the three treatments (Sn, No Sn, and Control) were similar for
the new and aged coupons. The variability in total lead levels may be due to coupon disturbance

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62 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

A B

1.2 1.2
Sn Sn
1 No Sn 1 No Sn

Dissolved Pb (mg/L)
Control Control
Dissolved Pb (mg/L)

0.8 0.8

0.6 0.6

0.4 0.4

0.2 0.2

0 0
0 5 10 15 20 25 30 35 40 0 5 10 15 20 25 30 35 40

Time (day) Time (day)

C D

1.2 1.2
Sn Sn
1 No Sn 1 No Sn
Control

Dissolved Pb (mg/L)
Dissolved Pb (mg/L)

Control
0.8 0.8

0.6 0.6

0.4 0.4

0.2 0.2

0 0
0 5 10 15 20 25 30 35 40 0 5 10 15 20 25 30 35 40
Time (day)
Time (day)

1.2
Average, Sn
1 Average, no Sn
Average, Control
Dissolved Pb (mg/L)

Sn
0.8 No Sn
Control
0.6

0.4

0.2

0
0 5 10 15 20 25 30 35 40

Time (day)

Figure 6.10 Concentration of dissolved lead versus time for the lead corrosion experiment
with aged (A-D) and new (E) lead coupons. The lines are drawn through the individual data
points in plots A-D and through the average of the duplicate data points in plot E. The reactors
contained lab water (10 mM of NaHCO3 at pH 8) amended with 1% R2A medium. The treatments
were as follows: (1) inoculated with bacteria (No Sn), (2) inoculated with bacteria and spiked
with stannous chloride at 0.125 mg/L as Sn+2 (Sn), and (3) no bacteria or tin (Control). Details on
coupon aging are given in Table 3.2.

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Chapter 6: Effect of Stannous Chloride on Lead Corrosion | 63

A B

3.5 3.5
Sn Sn
3 3
No Sn
No Sn
2.5 2.5 Control
Control
Total Pb (mg/L)

Total Pb (mg/L)
2 2

1.5 1.5

1 1

0.5 0.5

0 0
0 5 10 15 20 25 30 35 40 45 50
0 5 10 15 20 25 30 35 40 45 50
Time (day)
Time (day)

C D

3.5 3.5
Sn
3 Sn
No Sn 3
No Sn
2.5 Control 2.5 Control
Total Pb (mg/L)

Total Pb (mg/L)
2 2

1.5
1.5
1
1
0.5
0.5
0
0
0 5 10 15 20 25 30 35 40 45 50
0 5 10 15 20 25 30 35 40 45 50
Time (day)
Time (day)

3.5 Average, Sn
Average, no Sn
Average, Control
3 Sn
No Sn
2.5 Control
Total Pb (mg/L)

1.5

0.5

0
0 5 10 15 20 25 30 35 40 45 50

Time (day)

Figure 6.11 Concentration of total lead versus time for the lead corrosion experiment with
aged (A-D) and new (E) lead coupons. The lines are drawn through the individual data points in
plots A-D and through the average of the duplicate data points in plot E. The reactors contained
lab water (10 mM of NaHCO3 at pH 8) amended with 1% R2A medium. The treatments were as
follows: (1) inoculated with bacteria (No Sn), (2) inoculated with bacteria and spiked with stannous
chloride at 0.125 mg/L as Sn+2 (Sn), and (3) no bacteria or tin (Control). Details on coupon aging
are given in Table 3.2.

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64 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure 6.12 SEM image of biofilm on the surface of a lead coupon from Reactor 5 (Sn treatment)

Figure 6.13 SEM image of biofilm on the surface of a lead coupon from Reactor 15 (Control)

during changes in reactor solutions which could result in release of corrosion solids.
At the end of the experiment, the coupons were removed and examined using SEM.
The collection of images is included in Appendix D and two representative images are shown
in Figures 6.12 and 6.13. Most images show evidence of what appears to be bacterial biofilms,
including bacterial cells and EPS. Even the control reactors showed evidence of biofilm growth
(Figure 6.13) despite efforts to inactivate bacteria by periodically exposing the reactors to UV
light. The coupons from the control reactors, however, appeared to have less cells and EPS. The
accumulation of biofilm was more likely in these experiments as opposed to the coupon aging

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Chapter 6: Effect of Stannous Chloride on Lead Corrosion | 65

(Figure 6.7) because the lab water contained 1% R2A medium. These SEM images do not prove
a direct or indirect role for bacteria in the corrosion of lead, but lend some support to the hypoth-
esis that MIC may be important for lead. Stannous chloride was not effective at limiting biofilm
accumulation or lead corrosion in these experiments at 0.125 mg/L as Sn, likely because there
were substantial nutrients provided to support bacteria growth. Also, no chlorine was added which
would have aided in controlling biofilm development and possibly MIC.

SUMMARY

Lead coupons were aged for 70 weeks with lab and tap water in the presence and absence
of chlorine and presence and absence of stannous chloride. Minerals observed on the coupon sur-
faces at the end of the aging period did not vary significantly among the different treatments and
were dominated by: lead (II) oxide, cerussite and hydrocerussite. Plattnerite was also detected on
a few coupons. During the aging period, the electrical potential values for coupons aged in the
presence of stannous chloride were similar to those for coupons aged without stannous chloride.
Some of these aged coupons were placed in semi-batch reactors containing lab water with 1% R2A
medium and inoculated with Mississippi River water bacteria to investigate the role of MIC and
the effects of stannous chloride on lead corrosion. Bacteria and biofilms were prevalent on the lead
coupons at the end of the lead corrosion experiment for all reactors, including those containing tin
and those without tin. Furthermore, stannous chloride did not affect the concentration of dissolved
or total lead released from the coupons.

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©2010 Water Research Foundation. ALL RIGHTS RESERVED
CHAPTER 7
CONCLUSIONS AND RECOMMENDATIONS

CONCLUSIONS

Laboratory experiments were performed to investigate whether stannous chloride is effec-


tive at decreasing lead corrosion and to determine the mode of action. There were three primary
tasks, namely to investigate: (1) Sn chemistry in water, (2) Sn toxicity to planktonic and biofilm
bacteria, and (3) effects of Sn on lead corrosion. The main conclusions from this work are provided
below.
Sn+2 was oxidized to Sn+4 by oxidants typically present in treated water: chlorine, mono-
chloramine, and dissolved oxygen. Dissolved oxygen oxidized Sn+2 to Sn+4 at a relatively slow
rate (k = 0.069 hr–1). Both free chlorine and monochloramine oxidized Sn+2 at an extremely
fast rate (k > 42 hr–1). The oxidation of Sn+2 was typically incomplete, resulting in a mixture of
Sn+2 and Sn+4 in the water even in the presence of residual chlorine. Because the recommended
dose for water treatment is low (~0.2 mg/L as SnCl2), stannous chloride would exert a minor
chlorine demand.
Stannous chloride was highly toxic to laboratory strains of heterotrophic bacteria (P. aeru-
ginosa PAO1 and E. coli K-12) but not as toxic to a laboratory strain of nitrifiers (N. europaea).
Compared to these lab strains, environmental bacteria were significantly more resistant to Sn tox-
icity. For example, an environmental isolate (Acinectobacter sp.) from the Mississippi River was
resistant to inactivation by stannous chloride and stannic chloride. These toxicity and inactivation
experiments were performed with planktonic bacteria in the absence of chlorine to isolate the
effects of the stannous chloride. Additional experiments were performed to evaluate the toxicity of
stannous chloride addition on biofilm bacteria. More work is needed, however, to investigate the
effectiveness of stannous chloride in combination with free chlorine or chloramines for inactivat-
ing planktonic and controlling biofilm accumulation.
Stannous chloride successfully inhibited P. aeruginosa biofilm formation at doses as low
as 0.5 mg/L as Sn+2 and removed established biofilm at a dose of 1 mg/L as Sn+2. Also, because
stannous chloride was only tested on developed biofilm at a concentration of 1 mg/L as Sn in
the absence of chlorine and presence of substantial nutrients, it is unclear how stannous chloride
would impact established biofilm under conditions that better reflect those in water distribution
systems. The observed toxicity of SnCl2 suggests that the chemical may aid in controlling bacterial
regrowth in distribution systems and may inhibit MIC.
Lead coupons were aged for 70 weeks with lab and tap water in the presence and absence
of chlorine and presence and absence of stannous chloride. Minerals observed on the coupon sur-
faces at the end of the aging period did not vary significantly among the different treatments and
were dominated by lead oxide, cerussite and hydrocerussite. During the aging period, the electrical
potential values for coupons aged in the presence of stannous chloride were similar to those for
coupons aged without stannous chloride. Aged and new coupons were employed in semi-batch
reactors to investigate the role of MIC and the effects of stannous chloride on lead corrosion.
Bacteria and biofilms were prevalent on the lead coupons at the end of the lead corrosion experi-
ment for all conditions suggesting that MIC may be important for lead. Furthermore, stannous
chloride did not decrease the accumulation of biofilm on the coupons or the concentrations of

67

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68 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

dissolved and total lead released from the coupons. In fact, in many experiments lead levels were
greater in the tin-containing bottles in comparison to the other treatments. In comparison to typi-
cal tap waters, these experiments were performed under relatively high nutrient levels and without
chlorine. In pipe loop experiments with a treated water containing monochloramine, stannous
chloride use reduced lead levels and heterotrophic plate counts in comparison to an untreated con-
trol, but lead levels were still significantly greater than those for a pipe loop fed orthophosphate.
Certainly the experiments performed in this study covered a small range of possible treated water
qualities, so more work is needed to evaluate the effectiveness of stannous chloride use under a
wide range of conditions. Finally, the bacteria on the coupons were not identified and their role in
lead corrosion, if any, was not elucidated.

RECOMMENDATIONS

Utilities considering use of stannous chloride for lead corrosion control should proceed
slowly and with caution. In previous pipe loop studies, stannous chloride showed some benefit
with respect to reducing lead and bacterial levels, but the chemical did not decrease lead release or
biofilm accumulation in the batch experiments with lead coupons performed in this work. Coupon
and possibly pipe loop studies are recommended for utilities considering use of stannous chloride
to evaluate whether the chemical might be effective for the given water quality conditions.
More research is needed to identify the bacteria that accumulate on lead surfaces and to elu-
cidate their activity, including a possible role in MIC. Furthermore, additional research is needed
to investigate the effects of stannous chloride on lead corrosion and microbial activity for a wider
range of water qualities including low levels of free chlorine and chloramines. Also, there is a need
for mechanistic studies with more sensitive surface analytical techniques to determine Sn valence
state and surface speciation. In addition, although sensitive techniques are available for measuring
total Sn in water, more sensitive techniques are needed to distinguish between Sn(II) and Sn(IV).
Finally, more information is needed on Sn complexation in water (and related stability constants),
as Sn speciation is likely to have a strong role in bioavailability and toxicity.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


APPENDIX A
METHOD DEVELOPMENT

69

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70 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

MEASURING SN+2 WITH TOLUENE-3,4-DITHIOL

The effect of Sn+2 concentration on the kinetics of reaction with toluene-3,4-dithiol (TDT)
at a fixed concentration of 20 mg/L was investigated (Clark, 1936). The Sn+2 concentrations used
were: 0 mg/L (blank), 0.1 mg/L, 0.5 mg/L, 1 mg/L, 5 mg/L, and 10 mg/L. In an anaerobic glove-
bag, the Sn+2 solutions were prepared and a 10 mL aliquot of each was placed into a 30 mL glass
vial for the analysis. The vials were capped and sealed with parafilm before removing from the
anaerobic environment. Then, TDT was added to the sample vial (final concentration = 20 mg/L)
and the timer started. The vial was then mixed by shaking and an aliquot of the solution was
placed into a 1 cm cuvette for analysis using a spectrophotometer at 550 nm. Absorbance read-
ings were taken every 30 seconds for up to 45 minutes. The reaction results of TDT and Sn+2 at
various concentrations are shown in Figure A.1. More than 95% of the final value was attained
within 30 minutes. Thus, this determined the reaction time (after 30 minutes) for all analyses on
absorbance measurements.

1.2
Absorbance initial/Absorbance final

0.8

0.6
10 mg/L Sn
0.4 5 mg/L Sn
0.5 mg/L Sn
0.2 0.1 mg/L Sn

0
0 10 20 30 40
Time (min)

Figure A.1 Effect of Sn+2 at concentrations 0.1, 0.5, 5, and 10 mg/L on the kinetics of color
development at a constant toluene-3,4-dithiol concentration of 20 mg/L

©2010 Water Research Foundation. ALL RIGHTS RESERVED


APPENDIX B
PHOTOGRAPHS OF PSEUDOMONAS AERUGINOSA PAO1 BIOFILM
GROWING IN CAPILLARY FLOW CELL REACTORS

71

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72 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Table B.1
Control reactor (no Sn added to medium)
Experiment 1
0h 18 h 24 h 48 h

Experiment 2
0h 12 h 24 h 48 h

(continued)

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Appendix B: Photographs of Psuedomonas aeruginosa PAO1 Biofilm Growing in Capillary Flow Cell Reactors | 73

Table B.1 (Continued)


Experiment 3
0h 18 h 24 h 48 h

Experiment 4
0h 24 h 48 h 72 h 96 h

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74 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Table B.2
0.125 mg/L of Sn added to medium
Experiment 1
0h 18 h 24 h 48 h

Experiment 2
0h 18 h 24 h 48 h

Experiment 3
0h 18 h 26 h 48 h

(continued)

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Appendix B: Photographs of Psuedomonas aeruginosa PAO1 Biofilm Growing in Capillary Flow Cell Reactors | 75

Table B.2 (Continued)


Experiment 4
0h 18 h 24 h 48 h

Table B.3
0.5 mg/L of Sn added to medium
Experiment 1
0h 18 h 24 h 48 h

(continued)

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76 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Table B.3 (Continued)


Experiment 2
0h 18 h 24 h 48 h

Experiment 3
0h 18 h 24 h 48 h

(continued)

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Appendix B: Photographs of Psuedomonas aeruginosa PAO1 Biofilm Growing in Capillary Flow Cell Reactors | 77

Table B.3 (Continued)


Experiment 4
0h 18 h 24 h 48 h

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78 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Table B.4
Removal of established biofilm with 1 mg/L of Sn+2 
Before Sn was added
0h 18 h 24 h 48 h

After Sn was added at time = 48 h


48 h 66 h 73 h 96 h

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APPENDIX C
X-RAY DIFFRACTION SPECTRA OF LEAD COUPONS

79

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80 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure C.1 Spectrum of new lead coupon

Figure C.2 Spectrum of lead coupon aged in lab water only

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Appendix C: X-Ray Diffraction Spectra of Lead Coupons | 81

Figure C.3 Spectrum of lead coupon aged in tap water only

Figure C.4 Spectrum of coupon aged in lab water with free chlorine (1 mg/L as Cl2) and
SnCl2 (0.125 mg/L Sn)

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82 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure C.5 Spectrum of lead coupon aged in lab water with SnCl2 (0.125 mg/L as Sn)

Figure C.6 Spectrum of lead coupon aged in lab water with free chlorine (1 mg/L as Cl2)

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Appendix C: X-Ray Diffraction Spectra of Lead Coupons | 83

Figure C.7 Spectrum of lead coupon aged in lab water with monochloramine (1 mg/L as Cl2)

Figure C.8 Spectrum of lead coupon aged in tap water with monochloramine (1 mg/L as Cl2)
and SnCl2 (0.125 mg/L as Sn)

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©2010 Water Research Foundation. ALL RIGHTS RESERVED
APPENDIX D
SCANNING ELECTRON MICROGRAPHS OF LEAD COUPONS

85

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86 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.1 New lead coupon (i.e., reference)

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Appendix D: Scanning Electron Micrographs of Lead Coupons | 87

Figure D.2 Coupon A (aged in tap water with Sn+2 with free chlorine) from Reactor 1
(Sn+2 = 0.125 mg/L)

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88 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.3 Coupon A (aged in tap water with Sn+2) from Reactor 2 (Sn+2 = 0.125 mg/L)

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Appendix D: Scanning Electron Micrographs of Lead Coupons | 89

Figure D.4 Coupon B (aged in tap water with Sn+2) from Reactor 3 (Sn+2 = 0.125 mg/L)

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90 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.5 Coupon B (aged in tap water with monochloramine) from Reactor 4
(Sn+2 = 0.125 mg/L)

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Appendix D: Scanning Electron Micrographs of Lead Coupons | 91

Figure D.6 Coupon C (new coupon) from Reactor 5 (Sn+2 = 0.125 mg/L)

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92 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.7 Coupon C (new coupon) from Reactor 6 (Sn+2 = 0.125 mg/L)

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Appendix D: Scanning Electron Micrographs of Lead Coupons | 93

Figure D.8 Coupon A (aged in tap water with Sn+2 and free chlorine) from Reactor 7 (No Sn)

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94 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.9 Coupon A (aged in tap water with Sn+2) from Reactor 8 (No Sn)

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Appendix D: Scanning Electron Micrographs of Lead Coupons | 95

Figure D.10 Coupon B (aged in tap water with Sn+2) from Reactor 9 (No Sn)

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96 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.11 Coupon B (aged in tap water with monochloramine) from Reactor 10 (No Sn)

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Appendix D: Scanning Electron Micrographs of Lead Coupons | 97

Figure D.12 Coupon C (new coupon) from Reactor 11 (No Sn)

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98 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.13 Coupon C (new coupon) from Reactor 12 (No Sn)

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Appendix D: Scanning Electron Micrographs of Lead Coupons | 99

Figure D.14 Coupon A (aged in tap water with Sn+2 and free chlorine) from Reactor 13
(Control)

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100 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.15 Coupon A (aged with tap water with Sn+2) from Reactor 14 (Control)

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Appendix D: Scanning Electron Micrographs of Lead Coupons | 101

Figure D.16 Coupon B (aged with tap water with Sn+2) from Reactor 15 (Control)

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102 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.17 Coupon B (aged with tap water with monochloramine) from Reactor 16 (Control)

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Appendix D: Scanning Electron Micrographs of Lead Coupons | 103

Figure D.18 Coupon C (new coupon) from Reactor 17 (Control)

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104 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Figure D.19 Coupon C (new coupon) from Reactor 18 (Control)

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Appendix D: Scanning Electron Micrographs of Lead Coupons | 105

Figure D.20 Lead coupon aged in lab water with free chlorine and Sn+2 

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©2010 Water Research Foundation. ALL RIGHTS RESERVED
References

Aiking, H., Govers, H., and Van’t Riet, J. 1985. Detoxification of Mercury, Cadmium and
Lead in Klebsiella aerogenes NCTC 418 Growing in Continuous Culture. Applied and
Environmental Microbiology, 50:1262–1267.
Ashby, J., and Tennant, R.W. 1991. Definitive Relationships Among Chemical and Mutagenicity
for 301 Chemicals Tested by the U.S. NTP. Mutation Research, 257(3):229–306.
Baud, M. 1993. Data Analysis, Mathematical Modeling. In Methods of Immunological Analysis.
Vol. 1: Fundamentals. Edited by R.F. Masseyeff. New York: VCH Publishers.
Cantor, A.F., Park, J.K., and Vaiyavatjamai, P. 2003. Effect of Chlorine on Corrosion in Drinking
Water Systems. Jour. AWWA, 95(5):112–123.
Capone, D.G., Reese, D.D., and Kiene, R.P. 1983. Effects of Metals on Methanogenesis, Sulfate
Reduction, CO2 Evolution and Microbial Biomass in Anoxic Salt Marsh Sediments. Applied
and Environmental Microbiology, 45:1586–1591.
Chang, J.S., Law, R., and Chang, C.C. 1996. Biosorption of Lead, Copper, and Cadmium by
Biomass of Pseudomonas aeruginosa PU21. Water Research, 31(7):1651–1658.
Cheng, K.L., Cleno, K., and Imamura, I. 1992. CRC Handbook of Organic Analytical Reagents.
Boca Raton, Fla.: CRC Press.
Clark, R.E.D. 1936. The Colorimetric Determination of Tin by Means of Toluene-3,4-dithiol. The
Analyst, 62(728):661–663.
Clement, J.A. 1993. Full-Scale Performance Testing of Sodium Silicate to Control the Corrosion
of Lead, Copper, and Iron: York, Maine. In Control of Lead and Copper in Drinking Water.
EPA/625/R-93/001. Washington D.C.: USEPA Office of Research and Development.
Cotton, F.A., and Wilkinson, G. 1980. Advanced Inorganic Chemistry, 4th ed. New York: John
Wiley and Sons.
Crittenden, J.C., and MWH (Montgomery Watson Harza). 2005. Water Treatment: Principles and
Design. Hoboken, N.J.: John Wiley and Sons.
Dantas., F.J.S., Moraes, M.O., Carvalho, E.F., Valsa, J.O., Bernado-Filho, M., and Caldeira-de-
Araujo, A. 1996. Lethality Induced by Stannous Chloride on Escherichia coli AB1157:
Participation of Reactive Oxygen Species. Food and Chemical Toxicology, 34(10):959–962.
Dantas, F.J.S., Moraes, M.O., de Mattos, J.C.P., Bezerra, R.J.A.C., Carvalho, E.F., Bernardo, M.,
and de Araujo, A.C. 1999. Stannous Chloride Mediates Single Strand Breaks in Plasmid
DNA Through Reactive Oxygen Species Formation. Toxicology Letters, 110(3):129–136.
Dantas, F.J.S., de Mattos, J.C.P., and Moraes, M.O. 2002. Genotoxic Effects of Stannous Chloride
(SnCl2) in K562 Cell Line. Food and Chemical Toxicology, 40(10):1493–1498.
Duncan, S.J., and Ganiaris, H. 1987. Some Sulphide Corrosion Products on Copper Alloys and
Lead Alloys From London Waterfront Sites. In Recent Advances in the Conservations
and Analysis of Artifacts. Jubilee Conservation Conference Papers, compiled. Edited by
J. Black. London: Summer Schools Press.
Edwards, M., and Dudi, A. 2004. Role of Chlorine and Chloramine in Corrosion of Lead Bearing
Plumbing Materials. Jour. AWWA, 96(10):69–81.
Edwards, M., and McNeill, L. 2002. Effect of Phosphate Inhibitors on Lead Release From Pipes.
Jour. AWWA, 94(1):79–90.

107

©2010 Water Research Foundation. ALL RIGHTS RESERVED


108 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Emde, K.M.E., Smith, D.W., and Facey, R. 1992. Initial Investigation of Microbially Influenced
Corrosion (MIC) in a Low-temperature Water Distribution System. Water Research,
26(2):169–175.
Escobar, I., and Randall, A.A. 2001. Case Study: Ozonation and Distribution System Biostability.
Jour. AWWA, 93(10):77–89.
Gerischer, U. 2008. Acinetobacter Molecular Biology, 1st ed. Ciaster Academic Press.
Guibaud, G., Bordas, F., Saaid, A., D’abzac, P., and Hullebusch, E.V. 2008. Effect of pH on
Cadmium and Lead Binding by Extracellular Polymeric Substances (EPS) Extracted From
Environmental Bacterial Strains. Colloids and Surfaces, 63:48–54.
Hoff, J.C., and Geldreich, E.E. 1981. Comparison of the Biocidal Efficiency of Alternative
Disinfectants. Jour. AWWA, 73(1):40–45.
Holm, T.R., and Schock, M.R. 1991. Potential Effects of Polyphosphate Products on Lead Solubility
in Plumbing Systems. Jour. AWWA, 83(7):76–82.
Hozalski, R., Esbri-Amador, E., and Chen, C. 2005. Comparison of Stannous Chloride and
Phosphate for Lead Corrosion Control. Jour. AWWA, 97(3):89–103.
Javaherdashti, R. 2008. Microbiologically Influenced Corrosion: An Engineering Insight. London:
Springer-Verlag.
Karalekas, P.C., Jr. 1983. Control of Lead, Copper, and Iron Pipe Corrosion in Boston. Jour.
AWWA, 75(2):92–95.
King, R.A. 2007. Microbiologically Induced Corrosion and Biofilm Interactions. MIC—An
International Perspective Symposium, Extrin Corrosion Consultants. Perth, Australia:
Curtin University. Feb. 14–15.
Kipper de Silva, M., Tessaro, I.C., and Wada, K. 2006. Investigation of Oxidative Degradation
of Polyamide Reverse Osmosis Membranes by Monochloramine Solutions. Journal of
Membrane Science, 282:375–382.
Kirmeyer, G.J., and Pierson, G. 2000. Distribution System Water Quality Changes Following
Corrosion Control Strategies. Denver, Colo.: Awwa Research Foundation and AWWA.
LeChevallier, M.W., Lowery, C.D., and Lee, R. 1990. Disinfecting Biofilms in a Model Distribution
System. Jour. AWWA, 82(7):87–99.
Liu, H., Xu, L., and Zeng, J. 2000. Role of Corrosion Products in Biofilms in Microbiologically
Induced Corrosion of Carbon Steel. British Corrosion Journal, 352:131–135.
Loka Bharathi, P.A., Sathe, V., and Chandramohan, D. 1990. Effect of Lead, Mercury, and Cadmium
on a Sulphate-Reducing Bacterium. Environmental Pollution, 67:361–374.
Lytle, D.A., and Shock, M.R. 2008a. Formation of Pb(IV) Oxides in Chlorinated Water. Jour.
AWWA, 97(11):102–115.
Lytle, D.A., and Shock, M.R. 2008b. Pitting Corrosion of Copper in Waters With High pH and
Low Alkalinity. Jour. AWWA, 100(3):115–129.
Lytle, D.A., Gerke, T.L., and Maynard, J.B. 2005. Effect of Bacterial Sulfate Reduction on Iron-
Corrosion Scales. Jour. AWWA, 97(10):109–121.
Mason Corporation. 2008. <www.tinchemical.com/products.html#stannous>. Accessed:
September 4, 2008.
McKubre, M.C.H., and Syrett, B.C. 1986. Harmonic Impedance Spectroscopy for the Determination
of Corrosion Rates in Cathodically Protected Systems. In Corrosion Monitoring in
Industrial Plants Using Nondestructive Testing and Electrochemical Methods. Edited by
G.C. Moran and P. Labine. Philadelphia: ASTM International.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


References | 109

McNeil, M.B., and Little, B.J. 1990. Mackinawite Formation During Microbial Corrosion.
Corrosion, 46(7):599–600.
McNeil, M.B., and Odom, A.L. 1994. Thermodynamic Prediction of Microbially Influenced
Corrosion (MIC) by Sulfate-Reducing Bacteria (SRB). In Microbiologically Influenced
Corrosion Testing. Edited by J.R. Kearns and B.J. Little. Philadelphia: ASTM International.
Motlagh, A.R.A. 2008. Application of Cross-Flow Hollow-Fiber Membranes in Water and
Wastewater Treatment. Ph.D. Diss., University of Minnesota, Minneapolis.
Neil, T.K., Abraham, N.G., Levere, R.D., and Kappas, A. 1995. Differential Heme Oxygenase
Induction by Stannous and Stannic Ions in the Heart. Journal of Cellular Biochemistry,
57(3):409–414.
NSF International. 2001. NSF/ANSI Standard 61. Drinking Water System Components—Health
Effects.
NSF International. 2008. NSF/ANSI Standard 60. NSF Certified Drinking Water Treatment
Chemicals. Stannous Chloride (Anhydrous and Dihydrated).
Oldenburg, P.S., Regan, J.M., Harrington, G.W., and Noguera, D.R. 2002. Kinetics of Nitrosomonas
europaea Inactivation by Chloramine. Jour. AWWA, 94(10):100–110.
Piyachaturawat, P. 2005. Potential N-nitrosodimethylamine (NDMA) Formation From Water
Treatment Polymers. MS thesis, Georgia Institute of Technology, Atlanta.
Pontius, F. 2007. First Draw on Lead and Copper Rule Revisions. Jour. AWWA, 99(12):13–20.
Prasad, M.N.V. 2001. Metals in the Environment: Analysis by Biodiversity. Boca Raton, Fla.: CRC
Press.
Pungartnik, A.C., Viaua, C., and Picada, J. 2005. Genotoxicity of Stannous Chloride in Yeast and
Bacteria. Mutation Research, 583:146–157.
Rahal, J. 2006. Novel Anitbiotic Combinations Against Infections With Almost Completely
Resistant Pseudomonas aeruginosa and Acinetobacter Species. Clinical Infection Disease,
43(Suppl. 2):S95–S99.
Renner, R. 2004. Plumbing the Depths of D.C.’s Drinking Water Crisis. Environmental Science
and Technology, 38(12):224A–227A.
Sand, W. 1997. Microbial Mechanisms of Deterioration of Inorganic Substrates—A General
Mechanistic Overview. International Biodeterioration and Biodegradation, 40(2):183–190.
Shock, M.R. 1980. Response of Lead Solubility to Dissolved Carbonate in Drinking Water. Jour.
AWWA, 72(12):694–704.
Shock, M.R. 1989. Understanding Corrosion Control Strategies for Lead. Jour. AWWA,
81(7):88–100.
Shock, M.R., Lytle, D.A., Sandvig, A.M., Clement, J., and Harmon, S.M. 2005. Replacing
Polyphosphate With Silicate to Solve Lead, Copper and Source Water Iron Problems. Jour.
AWWA, 97(11):84–93.
Silva, C.R., Valsa, J.O., Caniné, M.S., Caldeira-de-Araujo, A., and Bernardo-Filho, M. 2002.
Biological Effects of Stannous Chloride, a Substance That Can Produce Stimulation or
Depression of the Central Nervous System. Brain Research Bulletin, 59:213–216.
Singh, I. 1983. Induction of Reverse Mutation and Mitotic Gene Conversion by Some Metal
Compounds in Saccharomyces cerevisae. Mutation Research, 117:149–152.
Stewart, P.S., Hamilton, M.A., Goldstein, B.R., and Schneider, B.T. 1996. Modeling Biocide
Action Against Biofilms. Biotechnology and Bioengineering, 49:445–455.

©2010 Water Research Foundation. ALL RIGHTS RESERVED


110 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

Stoffyn, P., and Buckley, D.E. 1992. The Titanic 80 Years Later: Initial Observations on the
Microstructure and Biogeochemistry of Corrosion Products. In Proceedings of the 50th
Annual Meeting of the Electron Microscope Society of America. San Francisco: San
Francisco Press.
Taheri, R., Nouhi, A., Harnedi, J., and Javaherdashti, R. 2005. Comparison of Corrosion Rates of
Some Steels in Batch and Semi-Continuous Cultures of SRB. Asian Journal of Microbiology
and Biotech Environmental Science, 7(1):5–8.
Teitzel, G.M., and Parsek, M.R. 2003. Heavy Metal Resistance of Biofilm and Planktonic
Pseudomonas aeruginosa. Applied and Environmental Microbiology, 69:2313–2320.
Tokajian, S.T., Hashwa, F.A., Hancock, I.C., and Zalloua, P.A. 2005. Phylogenetic Assessment of
Heterotrophic Bacteria From a Water Distribution System Using 16S rDNA Sequencing.
Canadian Journal of Microbiology, 51(4):325–335.
Uchida, M., and Okuwaki, A. 1999. Dissolution Behavior of Lead Plates in Aqueous Nitrate
Solutions. Corrosion Science, 41:1977–1986.
USFDA (U.S. Food and Drug Administration). 1982. Listing of Specific Substances Affirmed as
GRAS Section. 184.1845 Stannous Chloride (Anhydrous and Dihydrated). Code of Federal
Regulations, 47 FR 27816, June 25, 1982.
USFDA (U.S. Food and Drug Administration). 2003. Canned Asparagus Deviating From Identity
Standard; Temporary Permit for Market Testing. Federal Register, 68(117):36567.
Vasquez, F.A., Heaviside, R., Tang, Z.J., and Taylor, J.S. 2006. The Effects of Free Chlorine and
Chloramines on Lead Release in a Distribution System. Jour. AWWA, 98(2):144–155.
Videla, H.A. 1996. Manual of Biocorrosion. Boca Raton, Fla.: CRC Press.
Waara, K.O. 1991. Effects of Copper, Cadmium, Lead and Zinc on Nitrate Reduction in a Synthetic
Water Medium and Lake Water From Northern Sweden. Water Resources, 26:355–364.
Zhang, M., Semmens, M., Schuler, D., and Hozalski, R. 2002. Biostability and Microbiological
Quality in a Chloraminated Distribution System. Jour. AWWA, 94(9):112–122.

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ABBREVIATIONS

AAS atomic absorption spectrophotometry


AL action level
AOB ammonia oxidizing bacteria
ATCC American Type Culture Collection
AWWA American Water Works Association

BOD biochemical oxygen demand

C concentration
°C degrees Celsius
CBE Center for Biofilm Engineering
Cd cadmium
CFU colony forming unit
Cl– chloride ion
Cl2 chlorine
cm centimeter
CO2 carbon dioxide
CO32– carbonate ion
Cu0 zero-valence copper
Cu2+ cupric ion
CuO cupric oxide
CuS copper monosulfide

DIC dissolved inorganic carbonate


DNA deoxyribonucleic acid
dNTP deoxyribonucleotide
DO dissolved oxygen
DRDFAS N diethyl-p-phenylenediamine-ferrous ammonium sulfate
DS-Red fluorescent marker
D change in
DG0 Gibbs energy at ideal conditions

e– electron
EPS extracellular polymeric substances
exp exponential

Fe0 zero-valent iron


Fe2+ ferrous ion
FeOOH iron(III) oxide-hydroxide
FeS iron(II) sulfide
FeS2 pyrite

111

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112 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

GFP green fluorescent protein

h hour
H+ hydron ion
H 2 hydrogen
HCl hydrochloric acid
Hg mercury
H2O water
HOCl hypochlorous acid
HPC heterotrophic plate count
hr hour
hr–1 per hour
HS– anionic form of hydrogen sulfidet

ICP-MS ICP-mass spectrometry


ICP-OES inductively coupled plasma-optical emission spectrometry

k rate constant
kV kilovolts

L liter
LB Luria-Bertani
LCR Lead and Copper Rule
LD50 lethal dose at which 50% of subjects will die
LW lab water
l wavelength

M molar
mA milliamp
MDL method detection
mg milligram
MIC microbially-influenced corrosion
min minute
mL milliliter
mm millimeter
mM millimolar
MN Minnesota
mV millivolt
mg microgram
mL microliter
mm micrometer
mM micromolar

N nitrogen
N 2 nitrogen gas

©2010 Water Research Foundation. ALL RIGHTS RESERVED


Abbreviations | 113

Na2CO3 sodium carbonate


NaOH sodium hydroxide
NH3 ammonia
NH4+ ammonium ion
NH2Cl chloramine
nm nanometers
NO2– nitrite ion
NO3– nitrate ion
NOM natural organic matter

O 2 oxygen
OD600 optical density at 600 nm
OH– hydroxide ion
OUR oxygen uptake rate

P phosphorus
Pb lead
Pb0 zero-valent lead
Pb2+ plumbous ion
Pb(II) plumbous ion
Pb(IV) plumbic ion
PbCO3 cerussite
Pb3(CO3)2COH2 lead hydroxycarbonate
Pb3(CO3)2(OH)2 hydrocerussite
PbO lead oxide
PbO2 plattnerite
PBS phosphate buffered saline
PCR polymerase chain reaction
P.E. Professional Engineer
ppm parts per million
PTFE plytetrafluoroethylene

QA/QC quality assurance/quality control

R gas constant
rpm revolutions per minute
rRNA ribosomal RNA

S2– sulfide ion


SCE standard calomel electrode
SEM scanning electron microscopy
SiO2 silicon dioxide
Sn tin
Sn2+ stannous ion
Sn4+ stannic ion

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114 | An Investigation of Stannous Chloride as an Inhibitor of Lead Corrosion

SnCl2 stannous chloride


Sn(II) stannous ion
Sn(IV) stannic ion
Sn(OH)2 stannous hydroxide
Sn(OH)4 stannic acid
SO42– sulfate ion
sp. species
SPRWS St. Paul Regional Water Services
SRB sulfate-reducing bacteria

T temperature
TDT toluene dithiolate
TW tap water

U.S. United States


U.S.A. United States of America
USFDA U.S. Food and Drug Administration
UV ultraviolet

XRD X-ray diffraction

Zn zinc

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