THE CHARACTERIZATION OF THE ANTIBACTERIAL EFFICACY OF AN ELECTRICALLY ...

178 Pages·2011·3.28 MB·English

by  Rohan Shirwaiker

The Pennsylvania State University

The Graduate School

Department of Industrial and Manufacturing Engineering
THE CHARACTERIZATION OF THE ANTIBACTERIAL EFFICACY OF AN
ELECTRICALLY ACTIVATED SILVER ION-BASED SURFACE SYSTEM
A Dissertation in

Industrial Engineering

by

Rohan A. Shirwaiker
 2011 Rohan A. Shirwaiker
Submitted in Partial Fulfillment
of the Requirements
for the Degree of
Doctor of Philosophy

August 2011
The dissertation of Rohan A. Shirwaiker was reviewed and approved* by the following:

Richard A. Wysk
Professor Emeritus of Industrial Engineering
Dissertation Advisor
Chair of Committee

Robert C. Voigt
Professor of Industrial Engineering

Harriet B. Nembhard
Associate Professor of Industrial Engineering

Paul H. Cohen
Professor Emeritus of Industrial Engineering

Subhashinie Kariyawasam
Assistant Professor of Veterinary and Biomedical Sciences

Paul Griffin
Head of the Department of Industrial Engineering

*Signatures are on file in the Graduate School
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ABSTRACT
There have been growing concerns in the global healthcare system about the
eradication of pathogens in hospitals and other health-critical environments. The problem
has been aggravated by the overuse of antibiotics and antimicrobial agents leading to the
emergence of antibiotic-resistant superbugs such as methicillin-resistant Staphylococcus
aureus (MRSA) and vancomycin-resistant Enterococcus (VRE) which are difficult to
kill. Lower immunity of sick patients coupled with the escalating concurrent problem of
antibiotic-resistant pathogens has resulted in increasing incidences of hospital acquired
(nosocomial) infections. There is an immediate need to control the transmission of such
infections, primarily in healthcare environments, by creating touch-contact and work
surfaces (e.g., door knobs, push plates, countertops) that utilize alternative antibacterial
materials like the heavy metal, silver.
Recent research has shown that it is silver in its ionic (Ag+) and not elemental
form that is antibacterial. Thus, silver-based antibacterial surfaces have to release silver
ions directly into the pathogenic environment (generally, an aqueous media) in order to
be effective. This dissertation presents the study and analysis of a new silver-based
surface system that utilizes low intensity direct electric current (LIDC) for generation of
silver ions to primarily inhibit indirect contact transmission of infections. The broader
objective of this research is to understand the design, and characterization of the
electrically activated silver ion-based antibacterial surface system. The specific objectives
of this dissertation include:
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(1) Developing a comprehensive system design, and identifying and studying its critical
design parameters and functional mechanisms.
(2) Evaluating effects of the critical design parameters on the antibacterial efficacy of the
proposed surface system.
(3) Developing a response surface model for the surface system performance.
These objectives are achieved by formulating the system design, fabricating
prototypes with appropriate design parameters, evaluating the prototypes using various
physical and electrical characterization techniques, and characterizing the antibacterial
efficacy of the prototypes using statistical experiments. The major contributions of this
dissertation include:
(1) Design of a systems focused approach that quantifies the potential effectiveness of
silver ions under various configurations of the surface system design.
(2) Development of meso and micro-scale fabrication methodologies for prototype
fabrication.
(3) Development of microbiological testing protocols utilizing variance reduction
techniques to test the antibacterial efficacy of system prototypes.
(4) Development of empirical models for the surface system using factorial design of
experiments (DOE).
Basic results from the research demonstrate significant antibacterial efficacy of
the surface system against four dangerous bacteria including Staph aureus, Escherichia
coli, Pseudomonas aeruginosa, and Enterococcus faecalis which are together responsible
for more than 80% of nosocomial infections. Results of the DOE characterization study
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indicate the statistically significant contributions of three system parameters – size of
features, electric current, and type of bacteria – to the antibacterial performance of the
system.
This dissertation synergistically utilizes knowledge and principles from three
broader areas of research – industrial engineering, materials science and microbiology –
to model, design, fabricate and characterize an electrically activated silver-ion based
antibacterial surface system with practical applications in improving human health and
healthcare systems. The research is aimed at promoting novel integrative research and
development of technologies utilizing antibacterial properties of silver and other heavy
metals.
vi
TABLE OF CONTENTS
List of Figures ......................................................................................................... ix
List of Tables........................................................................................................... xii
Acknowledgements ................................................................................................. xiii
Chapter 1 Introduction ............................................................................................ 1
1.1 Background ................................................................................................ 1
1.2 Motivation .................................................................................................. 5
1.3 Research Objectives and Methodology........................................................ 9
1.4 Dissertation Overview................................................................................. 12
1.5 Chapter Summary ....................................................................................... 13
Chapter 2 Literature Review ................................................................................... 14
2.1 Silver as an Antibacterial ............................................................................ 14
2.2 Bactericidal Mechanisms of Silver Ions ...................................................... 16
2.3 Silver-based Antibacterial Surfaces and Coatings ........................................ 20
2.4 Electrically Generated Silver Ions ............................................................... 23
2.5 Chapter Summary ....................................................................................... 25
Chapter 3 Antibacterial Surface System Concept and Design .................................. 27
3.1 Basic System Design Configuration ............................................................ 27
3.2 Silver Ion Release Mechanism .................................................................... 29
3.3 Surface System Design Parameters ............................................................. 32
3.3.1 Physical Design Parameters ................................................................. 33
3.3.2 Electrical Design Parameters ............................................................... 39
3.4 Chapter Summary ....................................................................................... 40
Chapter 4 Antibacterial Surface System Fabrication ............................................... 41
4.1 Fabrication Approach .................................................................................. 41
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4.2 Meso-scale Fabrication Methodology.......................................................... 42
4.3 Micro-scale Fabrication Methodology ......................................................... 46
4.4 Chapter Summary ....................................................................................... 58
Chapter 5 Antibacterial Efficacy Testing Methodology ........................................... 59
5.1 Experimental Testing Procedure ................................................................. 59
5.2 Antibacterial Efficacy Calculations ............................................................. 65
5.3 Chapter Summary ....................................................................................... 67
Chapter 6 System Design and Prototype Validation Studies .................................... 68
6.1 Background of Validation Studies ............................................................... 68
6.2 Physical Inspection of System Prototypes ................................................... 69
6.2.1 Inspection of Micro-scale Prototypes ................................................... 70
6.2.2 Inspection of Meso-scale Prototypes .................................................... 74
6.3 Electrical Characterization of System Prototypes ........................................ 75
6.3.1 Electrical Characteristics of Prototypes in absence of Growth Media ... 76
6.3.2 Electrical Characteristics of TSB Growth Media.................................. 78
6.3.3 Electrical Characteristics of Prototypes in presence of TSB ................. 79
6.4 Evaluation of Surface Tension and Silver Release ....................................... 81
6.5 Evaluation of Antibacterial Efficacy of Proposed System Design ................ 88
6.5.1 Verification of Antibacterial Efficacy of Prototypes ............................ 88
6.5.2 Effect of BFU Width and Current on System’s Antibacterial Efficacy . 91
6.6 Chapter Summary ....................................................................................... 95
Chapter 7 System Characterization using Statistical Analysis ................................. 97
7.1 Design of Experiments (DOE) Study .......................................................... 97
7.1.1 DOE Factors........................................................................................ 100
7.1.1.1 BFU Width (System Scale) ....................................................... 100
7.1.1.2 Partition-Electrode Thickness Ratio........................................... 102
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7.1.1.3 Electric Current ......................................................................... 104
7.1.1.4 Voltage ..................................................................................... 107
7.1.1.5 Type of Bacteria ........................................................................ 108
7.1.2 DOE Response Variable ...................................................................... 110
7.1.3 Design Matrix and Experiments .......................................................... 111
7.2 Statistical Analyses ..................................................................................... 112
7.2.1 Estimating Factor Effects and Analyzing Residuals ............................. 113
7.2.2 Revised Model with Significant Main Effects and Interactions ............ 119
7.2.3 Evaluation of Models with Coded and Uncoded Units ......................... 125
7.3 Chapter Summary ....................................................................................... 127
Chapter 8 Research Contributions and Future Work................................................ 129
8.1 Research Summary ..................................................................................... 129
8.2 Research Contributions ............................................................................... 131
8.3 Future Work ............................................................................................... 133
Appendix A I-V Characterization of External Resistor Circuits............................... 136
Appendix B Minitab Output for Fractional Factorial Design ................................... 141
Appendix C Experimental Results from DOE ......................................................... 143
Appendix D Effects of Electrically Generated Silver Ions on Seven Antibiotic-
resistant/Spore-forming Nosocomial Bacteria ................................................... 147
Bibliography ............................................................................................................ 151
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LIST OF FIGURES
Figure 1-1: Annual hospital admissions with MRSA (based on data published by
CDC in 2007). .................................................................................................. 3
Figure 1-2: Basic concept of the electrically activated silver ion-based anti-
bacterial surface system (adapted from Patent pending PCT/US07/26379). ......... 10
Figure 1-3: A basic summary of the research methodology. .................................... 11
Figure 2-1: Bactericidal mechanisms of silver. ........................................................ 17
Figure 2-2: Internal structure of (a) untreated E. coli cells, and (b-d) cells treated
with Ag+ – (b) A big gap between the cytoplasm membrane and the cell wall,
(c) A cell composed of large electron-dense granules, (d) Seriously damaged
cell wall (Feng, et al., 2000).. ............................................................................ 19
Figure 3-1: Conceptual design of the surface system (a) Isometric view, and (b)
cross-sectional view .......................................................................................... 28
Figure 3-2: Schematic cross-section of the surface system ....................................... 33
Figure 3-3: Effect of basic functional unit width on surface contact area of a
droplet .............................................................................................................. 35
Figure 3-4: Configurations of a droplet on a surface in (a) Normal state on a
smooth surface, (b) Wenzel state on a rough surface, and (c) Cassie state on a
rough surface .................................................................................................... 38
Figure 4-1: Meso-scale fabrication methodology ..................................................... 45
Figure 4-2: Micro-scale fabrication methodology..................................................... 54
Figure 4-3: Geometric profiles of photoresist features when using (a) Image
reversal and (b) Bi-layer photoresists ................................................................ 57
Figure 5-1: Phases of batch culture bacteria growth ................................................. 60
Figure 5-2: Surface testing set-up ............................................................................ 62
Figure 6-1: A micro-scale prototype in comparison with a US dime......................... 70
x
Figure 6-2: Magnified images of a micro-scale prototype (top left and bottom
right corners of the functional area) .................................................................. 71
Figure 6-3: SEM images of a micro-scale prototype. The top image was obtained
with the beam perpendicular to the surface. In the bottom image, the beam
was at a 50° angle ............................................................................................. 72
Figure 6-4: Cross-section surface profiles of a micro-scale prototype. The top
image was taken starting at the left corner. The bottom image was taken
closer to the sample center ................................................................................ 73
Figure 6-5: A meso-scale prototype in comparison with a US dime ......................... 74
Figure 6-6: Current characteristics of a dry (a) micro-scale and (b) meso-scale
prototype .......................................................................................................... 77
Figure 6-7: Effect of change in separation between electrodes on current in TSB ..... 79
Figure 6-8: Current in micro-scale prototypes with an applied voltage of (a) 6 V
and (b) 15 V. The expected current in (a) is 1.5 µA and in (b) is 15 µA ............ 82
Figure 6-9: Current in meso-scale prototypes with an applied voltage of (a) 6 V
and (b) 15 V. The expected current in (a) is 1.5 µA and in (b) is 15 µA ............ 83
Figure 6-10: Edge profile of a TSB droplet on a micro-scale prototype sample
observed under an optical microscope with the sample placed at an angle of
30° with the horizontal ...................................................................................... 84
Figure 6-11: Optical image of a micro-scale prototype after extreme electrical
activation in presence of a TSB droplet ............................................................. 86
Figure 6-12: Surface Profile of micro-scale prototype after electrical activation in
presence of a TSB droplet ................................................................................. 87
Figure 6-13: Examples of blood agar plates from testing of (a) micro-scale (b)
meso-scale (c) acrylic control samples against S. aureus for 30 min .................. 91
Figure 6-14: Examples of MacConkey agar plates from testing of (a) micro-scale
(b) meso-scale (c) acrylic control samples against E. coli for 30 min................. 91
Figure 6-15: Percentage reduction in bacteria versus basic functional unit width ...... 93
Figure 6-16: Percentage reduction in bacteria versus system current ........................ 94