Doctor of Philosophy

University of Pittsburgh

2007

of the requirements for the degree of

the School of Engineering in partial fulfillment

MS, University of Pittsburgh, 2004

Submitted to the Graduate Faculty of

BS, BA, University of Pittsburgh, 2001

Gusphyl Antonio Justin

GENERATING ELECTRICITY WITHIN THE PHYSIOLOGICAL ENVIRONMENT FOR LOW POWER IMPLANTABLE MEDICAL

DEVICE APPLICATIONS: TOWARDS THE DEVELOPMENT OF IN- VIVO BIOFUEL CELL TECHNOLOGIES

by

SCHOOL OF ENGINEERING

This dissertation was presented

by

Gusphyl Antonio Justin

It was defended on

June 4th, 2007

and approved by

Robert Sclabassi, Professor, Department of Neurological Surgery

Mingui Sun, Professor, Department of Neurological Surgery

Harvey Borovetz, Professor, Bioengineering Department

Xinyan Tracy Cui, Assistant Professor, Bioengineering Department

Yingze Zhang, Associate Professor, Department of Pulmonary, Allergy and Critical Care Medicine

David Waldeck, Professor, Chemistry Department

Dissertation Director: Robert Sclabassi, Professor, Department of Neurological Surgery

UNIVERSITY OF PITTSBURGH

ii

Copyright © by Gusphyl A. Justin

2007

iii

GENERATING ELECTRICITY WITHIN THE PHYSIOLOGICAL ENVIRONMENT FOR LOW POWER IMPLANTABLE MEDICAL DEVICE APPLICATIONS: TOWARDS THE

DEVELOPMENT OF IN-VIVO BIOFUEL CELL TECHNOLOGIES

Gusphyl A. Justin, PhD

University of Pittsburgh, 2007

Electrochemical studies were performed to explore electron transfer (ET) between human white

blood cells (WBC) and carbon fiber electrodes (CFE). Currently, an active area of research

involves encouraging ET between microbes and various electrodes in a biofuel cell (BFC). ET

between microbes and electrodes are thought to occur i) directly through plasma membrane-

bound electron transport chain proteins; and/or ii) indirectly through the release of metabolic

products or biomolecules near the electrode surface. An important motivation of this research is

the need for alternative long lasting power sources for implantable diagnostic and therapeutic

devices. A particular interest is reducing the size and weight of implantable devices. Currently

employed internal batteries largely contribute to both. BFCs are promising prospects as they

couple the oxidation of a biofuel (such as glucose) to the reduction of molecular oxygen to water.

Both glucose and oxygen are abundantly present within our body’s cells and tissues. The goal of

this project is to explore the feasibility of utilizing WBCs (a human cell model) to generate

electricity by fostering direct or indirect ET between these cells - or more specifically, between

the metabolic processes of these cells - and the anode of a BFC. ET from the metabolic processes

of whole cells to electrodes had, to the best of our knowledge, only previously been

demonstrated for microbes. The electrochemical activities of WBCs isolated from whole human

blood by red blood cell (RBC) lysis, peripheral blood mononuclear cells (PBMCs) isolated on a

Ficoll-Paque gradient, as well as cells from a BLCL cell line and two leukemia cell lines (K562

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and Jurkat) were all investigated by incorporation of the cells into the anode compartment of a

proton exchange membrane fuel cell (PEMFC). Cyclic voltammetry was employed as an

electrochemical technique to investigate the ET ability of the cells, as it can reveal both

thermodynamic and kinetic information regarding oxidation-reduction processes at the CFE

surface. The results of our studies demonstrate that upon activation, biochemical species, such as

serotonin, are released by PBMCs, which may become irreversibly oxidized at the electrode

surface.

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TABLE OF CONTENTS

TABLE OF CONTENTS........................................................................................................................... VI

LIST OF TABLES .................................................................................................................................. VIII

LIST OF FIGURES ................................................................................................................................... IX

PREFACE..................................................................................................................................................XII

1.0 INTRODUCTION...........................................................................................................................1

2.0 BACKGROUND .............................................................................................................................4

2.1 CHALLENGES IN ENERGY DELIVERY TO IMPLANTABLE DEVICES ....................................................4

2.2 POWER SOURCES ...........................................................................................................................6

2.2.1 Internal Batteries.....................................................................................................................6

2.2.2 Nuclear Batteries.....................................................................................................................7

2.2.3 Piezoelectric Devices...............................................................................................................8

2.2.4 Biological Power Sources .......................................................................................................9

2.3 TRANSCUTANEOUS ENERGY TRANSMISSION................................................................................11

3.0 BIOFUEL CELLS: A REVIEW..................................................................................................12

3.1 INTRODUCTION TO BIOFUEL CELLS ..............................................................................................13

3.2 MICROBIAL FUEL CELLS...............................................................................................................17

3.3 ENZYME-BASED BIOFUEL CELLS ..................................................................................................24

4.0 NADPH OXIDASE .......................................................................................................................28

5.0 LEUKOCYTE-BASED BIOFUEL CELL STUDY ...................................................................33

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5.1 ABSTRACT ...................................................................................................................................33

5.2 INTRODUCTION............................................................................................................................34

5.3 EXPERIMENT I: MATERIALS AND METHODS..................................................................................37

5.3.1 White blood cell isolation......................................................................................................37

5.3.2 Biofuel cell Bioreactor Preparation ......................................................................................40

5.3.3 Cell activation and chemical manipulation ...........................................................................43

5.4 EXPERIMENT I: RESULTS AND DISCUSSION...................................................................................44

5.5 EXPERIMENT II: MATERIALS AND METHODS ...............................................................................58

5.5.1 White blood cell isolation......................................................................................................58

5.5.2 Measurement of open circuit potential and current ..............................................................59

5.6 EXPERIMENT II: RESULTS AND DISCUSSION .................................................................................60

5.7 CONCLUSIONS.............................................................................................................................82

6.0 CYCLIC VOLTAMMETRY AND HPLC OF LEUKOCYTES ..............................................84

6.1 ABSTRACT ...................................................................................................................................84

6.2 INTRODUCTION............................................................................................................................86

6.3 MATERIALS AND METHOD..................................................................................................88

6.3.1 Cyclic Voltammetry ...............................................................................................................88

6.3.2 Microdialysis and High Performance Liquid Chromatography............................................91

6.4 RESULTS AND DISCUSSION...........................................................................................................95

6.4.1 Cyclic Voltammetry ...............................................................................................................95

6.4.2 Microdialysis and HPLC.....