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Design, Simulation and Experimental Study of Shape Memory Alloy

and Micro-motor Activated High Pressure Optical Cell for Bio-Physical Studies

A Thesis

Submitted to the Faculty

of

Drexel University

by

Oliver Hongchun Xie

in partial fulfillment of the

requirements for the Degree

of

Doctor of Philosophy

Philadelphia, PA

October 2007

i
Drexel University
Office of Graduate Studies
Thesis Approval Form
(For Masters and Doctoral Students)

Hagerty Library will bind a copy of this form with each copy of your Thesis/dissertation.

This thesis, entitled __Design, Simulation and Experimental Study of Shape Memory Alloy

_____and Micro-motor Activated High Pressure Optical Cell for Bio-Physical Studies___

______________________________________________________________________________

and authored by_________Oliver_Hongchun Xie_________ , is hereby accepted and approved.

Signatures:

Chairman, Examining Committee: Supervising Professor:

________________________________ ________________________________

Committee Members:

________________________________ ________________________________

________________________________ ________________________________

________________________________ ________________________________

Program Advisor: Department Head:

________________________________ ________________________________
Dedications

To my mother, my father in heaven,


my sisters and brothers, and my friends

ii
Acknowledgements
I would like to express my sincere gratitude to my advisors, Professors Jack G.
Zhou from Drexel University and Parkson Chong from Temple University, for their
support and advice throughout the course of this research. Without their encouragement
and guidance, not only in my research and study but also in my new life in United States,
this doctoral research and my other researches would not have been accomplished.

I would like to express my appreciations to my PhD defense committee members,


Professors Kimberley Cook-Chennault, Moses Noh and Bor-Chin Chang for their
valuable time, helpful discussions and useful suggestions.

I would like to thank the Department of Mechanical Engineering and Mechanics,


as a whole, for their financial support in teaching assistantship. This research is founded
and supported by NSF. Thanks NSF to provide me with this challenge project, wonderful
research opportunity, best experimental facilities and working environments. I would like
to thank Dr. Leilkes and his Lab members for helping me to use INSTRON machine. I
would like to thank Berenice and other Dr. Chong’s Lab members for helping me on
fluorescence measurement.

I would like to thank Mark and all other Machine Shop members for fabricating
my experimental tools and modifying them numerous times. Also I would like to thank
glass shop artists, Tom from Drexel University, David from Temple University and
Karen from University of Pennsylvania for making various cuvettes for me.

In addition, I would like to thank my colleagues, Lin Lu, Yi Ma, John Lin, Tie Hu,
Chunlong Hu, for their caring and discussion. I would like to say that I enjoy sharing my
feeling with you. Your warm caring and encouragement, every word during my sad and
happy time, had given me more confidence.

I would like to express my sincerely thanks to my mom for her endless love,
support and encouragement throughout my PhD study and my life, to my brothers and
sisters, to my family members for their love, caring and support. Also I would like to
thank my friends I met in this country, Robert Harcum, Curtis Holloman, Reggie
Garrison, Bryan Skyes, Jingfu Tan, Jay Li and many more who cheered me up when I am
down and shared my joy of my happy time.

Above all, no single word can express my sincerely thanks deeply in my heart. I
would like to say again that this research work cannot be accomplished without your
support, help and caring.

iii
Table of Content

List of Figures.................................................................................................................. viii

ABSTRACT..................................................................................................................... xiv

1 Introduction................................................................................................................. 1

1.1 Problem Statement .............................................................................................. 1

1.2 Research Objective ............................................................................................. 1

1.3 Background and Review ..................................................................................... 2

1.3.1 Significance of HPOC on Bio-Physical Studies ......................................... 2

1.3.2 Interest of Living Organism under Pressure ............................................... 5

1.3.3 Weber’s High Pressure Bomb..................................................................... 7

1.3.4 The Limitations of the Paladini/Weber High-Pressure Optical Cell .......... 8

1.3.5 Zhou’s Effort to Mix Solution under Pressure............................................ 9

1.4 Research Achievements .................................................................................... 10

1.4.1 Design of the New SMA Activated Dual Chamber HPOC ...................... 11

1.4.2 Design of Shape Memory Alloy Activated Stopper and Mixing Systems 11

1.4.3 HPOC Finalization and Fabrication.......................................................... 13

1.4.4 Complete System Assembly and Final Testing ........................................ 13

2 Design of SMA Actuator .......................................................................................... 14

2.1 Review of Shape Memory Alloy Theory.......................................................... 14

2.1.1 History of Shape Memory Alloy: ............................................................. 14

2.1.2 Two Main Characteristics of Shape Memory Alloys ............................... 15

2.1.3 Commercial SME Alloys.......................................................................... 18

2.1.4 Metallurgical Property of NiTi Alloy: ...................................................... 19

iv
2.1.5 Thermoelastic Martensite Transformation................................................ 20

2.1.6 One Way Shape Memory.......................................................................... 25

2.1.7 Two Way Shape Memory ......................................................................... 25

2.1.8 Heat Treating and Shape Setting of SMA................................................. 27

2.1.9 The Limitations of Shape Memory Effect and Superelastic Behavior ..... 30

2.1.10 Application of SMA.................................................................................. 31

2.2 Challenges and Technical Approaches on SMA Based Actuator Design ........ 32

2.3 Various SMA Actuator Design......................................................................... 34

2.3.1 Design 1: Two-way Shape Memory Actuator .......................................... 34

2.3.2 Design 2: Two Commercial Tensile SMA Springs as Actuator ............... 37

2.3.3 Design 3: Actuator with self-developed SMA spring............................... 56

2.3.4 Design 4: SMA Extensive Spring and Biasing Compressive Spring

Actuator 62

2.3.5 Design 5: SMA extensive spring and micro-motor actuator .................... 79

3 Cuvette Design and Mixing Process ......................................................................... 83

3.1 Cuvette Design.................................................................................................. 83

3.1.1 Necking Design......................................................................................... 83

3.2 Mixing under High Pessure .............................................................................. 88

3.3 Turbulent flow................................................................................................... 93

3.4 Mechanism of mixing in turbulent flows.......................................................... 95

4 High Pressure Optical Cell Design ........................................................................... 97

4.1 Weber and Paladini’s High Pressure Bomb...................................................... 97

4.2 ISS’s current HPOC.......................................................................................... 98

v
4.3 HPOC Design: .................................................................................................. 99

4.3.1 HPOC Body Design.................................................................................. 99

4.3.2 Electrical Lead Design............................................................................ 100

4.3.3 Observation Window Design.................................................................. 104

4.3.4 Pressure Inlet Plug Design ...................................................................... 106

4.3.5 Temperature Monitoring under High Pressure Design........................... 106

4.3.6 Machine Finishes .................................................................................... 107

4.4 FEM Analysis of HPOC ................................................................................. 107

4.5 The Final HPOC Design ................................................................................. 110

4.6 Our New HPOC Fabrication collaborated with ISS ....................................... 111

5 Complete System Assembly and Final Testing ...................................................... 115

5.1 The Final Complete System............................................................................ 115

5.2 Leakage Testing of the Stopper ...................................................................... 119

5.3 Mixing Testing................................................................................................ 120

5.4 Conclusion ...................................................................................................... 127

6 Bio-Safety test of Heat Generation of SMA ........................................................... 128

6.1 Bio-safety testing of SMA wire ...................................................................... 128

6.2 Bio-Safety test on SMA heat generation ........................................................ 129

6.3 Avoiding Photo bleaching effect .................................................................... 130

7 Conclusion and future work.................................................................................... 134

Reference ........................................................................................................................ 136

8 Appendix................................................................................................................. 144

8.1 Instron force measurement of SMA springs ................................................... 144

vi
8.1.1 Commercial SMA spring test.................................................................. 144

8.1.2 In house made SMA spring testing......................................................... 146

8.2 ISS measurement test results .......................................................................... 150

8.2.1 Leakage Testing ...................................................................................... 150

8.2.2 Free diffusion .......................................................................................... 151

8.2.3 SMA spring and micro-motor mixing results (1 atm) raw results .......... 153

8.2.4 SMA spring and micro-motor mixing result under 300 bars (normalized)

155

8.2.5 Photo Bleaching ...................................................................................... 160

vii
List of Figures

Figure 2-1 Phase transformation versus temperature curve.............................................. 22

Figure 2-2 Martensite-Austenite transformation s-e (Harrison and Hodgson 1975)........ 23

Figure 2-3 Martensite and Austenite (Ryhänen)............................................................... 24

Figure 2-4 The shape transformation process (Ryhänen) ................................................. 25

Figure 2-5 Two-Way SMA actuator design...................................................................... 35

Figure 2-6 One-Way SMA actuator, dual SMA springs design ....................................... 38

Figure 2-7 Shape Memory Stress/ Strain/Temperature behavior (Warram 1993)............ 39

Figure 2-8 Constant deflection configuration (Warram 1993) ......................................... 40

Figure 2-9 Constant deflection hysteresis behavior (Warram 1993) ................................ 40

Figure 2-10 Constant force loading of a shape memory compression spring (Warram

1993) ................................................................................................................................. 41

Figure 2-11 Force-Deflection behavior for constant force operation ............................... 41

Figure 2-12 Superposition of linear Force-Deflection biasing spring .............................. 42

Figure 2-13 Typical stress-strain behavior of Shape Memory Alloy material ................. 43

Figure 2-14 Measurement parameter for Force-Deflection curve .................................... 44

Figure 2-15 Force of SMA at two phases (N/mm) D=0.135” d=0.019” .......................... 45

Figure 2-16 Force(N) vs Extension at high and low temperature (D=0.156”; d=0.018”) 46

Figure 2-17 Force(N) vs Extension at high and low temperature (D=0.156”; d=0.017”) 46

Figure 2-18 Spring dynamics diagram.............................................................................. 48

Figure 2-19 Spring dynamics Matlab simulation plot ...................................................... 49

viii
Figure 2-20 Heating and cooling time histories of actuators subjected to step voltage

changes of 4 volts with actuator wire diameter of: (a) 0.75mm (b) 0.875mm (Baz, Iman et

al. 1990) ............................................................................................................................ 50

Figure 2-21 Effect of cooling strategy on heating and cooling time histories of 0.4mm

actuators subjected to step voltage of 10 volts (Baz, Iman et al. 1990)............................ 51

Figure 2-22 Heating and cooling time histories of 0.4mm actuators subjected to

sinusoidal voltage excitations with air cooling velocity =7.2 m/s: (a) at 1/36Hz (b) at

1/20Hz (c) at 1/15Hz (Baz, Iman et al. 1990)................................................................... 52

Figure 2-23 Frequency generator circuit layout................................................................ 53

Figure 2-24 (a) Circuit board and relay (b) Two SMA spring testing in water ................ 54

Figure 2-25 Temperature measurement in water close to spring...................................... 55

Figure 2-26 Dual SMA springs actuator with tensile and compressive SMA springs ..... 57

Figure 2-27 Making SMA spring...................................................................................... 58

Figure 2-28 Surface response for Af Temperature and ultimate tensile stress ................. 59

Figure 2-29 Femlab heat transfer FEM Analysis for two SMA springs........................... 60

Figure 2-30 Heat transfer simulation for duo-SMA spring actuator................................. 61

Figure 2-31 SMA actuator with tensile SMA and biasing compressive spring................ 63

Figure 2-32 Heat Transfer analysis result (whole layout) ................................................ 64

Figure 2-33 1A current, 5 seconds heating response ........................................................ 65

Figure 2-34 2A current 5 seconds heating response ......................................................... 65

Figure 2-35 3A current 5 seconds heating response ......................................................... 66

Figure 2-36 4A current, 5 seconds heating response ........................................................ 67

Figure 2-37 5A current, 5 seconds heating response ........................................................ 67

ix
Figure 2-38 0.25 second, 5A heating and cooling process ............................................... 70

Figure 2-39 one coil cross-section of SMA wire close up heating condition................... 68

Figure 2-40 Temperature line plot of crossing SMA center 5A, 5seconds heating.......... 69

Figure 2-41 0.25 second heating, 1Hz and 10 cycles condition ....................................... 71

Figure 2-42 5A 0.25 second heating at 0.5Hz condition .................................................. 72

Figure 2-43 line plot at 5A 0.25 second heating 0.5Hz condition .................................... 73

Figure 2-44 Heat transfer analysis result for two coils of SMA wire close up................. 73

Figure 2-45 Line plot between 2 coils of 5A, 0.25 seconds heating, 0.5 Hz .................... 74

Figure 2-46 Temperature point plot at one wire diameter away from SMA coil (5A, 0.25

second heating, 0.5 Hz)..................................................................................................... 74

Figure 2-47 Temperature point plot at 10 times of wire diameter away from SMA coil

(5A, 0.25 second heating, 0.5 Hz) .................................................................................... 75

Figure 2-48 Temperature point plot at the center of coil (5A, 0.25s heating, 0.5 Hz) ..... 76

Figure 2-49 Temperature point plot (5A 0.33s heating 0.5Hz center).............................. 76

Figure 2-50 Mixing Measurement Setup .......................................................................... 77

Figure 2-51 Mixing Result of SMA Spring Actuator ....................................................... 78

Figure 2-52 Over Night and Manual Mixing Result........................................................ 79

Figure 2-53 SMA Spring Actuator with Micro-motor as a Mixer.................................... 80

Figure 2-54 Mixing result of SMA spring & micro-motor mixer .................................... 81

Figure 2-55 Close-up view at mixing start point .............................................................. 82

Figure 3-1 The Cuvette Design......................................................................................... 83

Figure 3-2 A cuvette of 8cm height and 2cm diameter with 3 quarter necking ............... 84

Figure 3-3 Diffusion time of liquid................................................................................... 85

x
Figure 3-4 Necking examination ...................................................................................... 86

Figure 3-5 Diffusion time for different necking ............................................................... 86

Figure 3-6 Point plots along the center line from necking to bottom of the cuvette ........ 87

Figure 3-7 Location of stirrer with motor within apparatus: 1) motor winding 2) rotor 3)

cooling coil 4) electrical leads 5) packing ........................................................................ 90

Figure 3-8 Power supply of powerful solenoid: 1) interuper 2) magnetic start 3)

transformer 4) autotransformer 5) solednoid .................................................................... 91

Figure 3-9 Drop stirrer: 1) coil 2) head 3) core 4) shaft 5) body of apparatus ................. 92

Figure 3-10 Stirrer with reversing rotation:1) core 2) axle 3) finger 4) tube.................... 92

Figure 3-11 Sample turbulent velocity record .................................................................. 94

Figure 4-1 Weber and Paladini’s Pressure Bomb ............................................................. 97

Figure 4-2 ISS HP Cell ..................................................................................................... 98

Figure 4-3 The design of HPOC, a transparent 3-D view................................................. 10

Figure 4-4 Exploded assembly view and assembly electrical lead................................. 102

Figure 4-5 Kalashnikov electrical lead: 1) body 2) holes 3) conductors 4) thread......... 103

Figure 4-6 Exploded view and assembly a window plug ............................................... 105

Figure 4-7 Stress analysis for cell assembly by Ansys ................................................... 108

Figure 4-8 First principle stress of cell assembly by Femlab ......................................... 109

Figure 4-9 Illustration of final HPOC design ................................................................. 110

Figure 4-10 Electrical lead and inlet plug combo........................................................... 112

Figure 4-11 Cuvette and its cap ...................................................................................... 113

Figure 5-1 Final Complete HPOC Assembly ................................................................. 116

Figure 5-2 Pressure control and cooling system ............................................................. 117

xi
Figure 5-3 ISS K2 Fluorescence Measurement .............................................................. 118

Figure 5-4 Optical diagram of ISS K2............................................................................ 119

Figure 5-5 Leakage ISS CF testing................................................................................. 120

Figure 5-6 Diffusion ISS CF testing ............................................................................... 121

Figure 5-7 Mixing with SMA and biasing spring actuator agitation .............................. 122

Figure 5-8 Intensity vs nM.............................................................................................. 123

Figure 5-9 Mixing result under 1atm first measurement ................................................ 124

Figure 5-10 Mixing result under 1atm second measurement.......................................... 124

Figure 5-11 Mixing result under 1atm third measurement ............................................. 125

Figure 5-12 Mixing result under 300bars, first measurement......................................... 126

Figure 5-13 Mixing result under 300bars, second measurement.................................... 126

Figure 5-14 Mixing result under 300bars, third measurement ....................................... 127

Figure 6-1 Photo bleaching vs light intensity ................................................................. 131

Figure 6-2 Photo Bleaching vs Enzyme effect @ 12k light intensity............................. 132

Figure 6-3 Photo Bleaching vs Enzyme effect @ 100k light intensity........................... 133

Figure 8-1 Low temperature tensile testing force vs time (1-4) ..................................... 144

Figure 8-2 Low temperature tensile testing force vs time (5-8) ..................................... 144

Figure 8-3 Low temperature tensile testing force vs time (9) unplugging force test (10)

......................................................................................................................................... 145

Figure 8-4 Load vs Extension ......................................................................................... 145

Figure 8-5 High Temperature Af force vs step extension loading (1-4) ......................... 146

Figure 8-6 High Temperature Af force vs step extension loading (5-7) ......................... 146

Figure 8-7 High Temperature Af force vs step extension loading .................................. 147

xii
Figure 8-8 High Temperature Af force vs step extension loading (1-3) failed............... 147

Figure 8-9 Current response (3) (by increasing current, the peak of force increases).... 148

Figure 8-10 Stopper force measurement......................................................................... 148

Figure 8-11 Low Temperature force measurement ........................................................ 149

Figure 8-12 Leakage testing............................................................................................ 150

Figure 8-13 Pipet feeding to imamates free diffusion .................................................... 151

Figure 8-14 Pipet feeding to imamates free diffusion .................................................... 151

Figure 8-15 Tube feeding imamates free diffusion......................................................... 152

Figure 8-16 Tube feeding imamates free diffusion......................................................... 152

Figure 8-17 1st result ....................................................................................................... 153

Figure 8-18 2nd mixing result.......................................................................................... 153

Figure 8-19 3rd result....................................................................................................... 154

Figure 8-20 1st result ....................................................................................................... 155

Figure 8-21 2nd Result..................................................................................................... 156

Figure 8-22 2nd result ...................................................................................................... 157

Figure 8-23 3rd result....................................................................................................... 158

Figure 8-24 4th result....................................................................................................... 159

Figure 8-25 Photo bleaching at higher intensity exciting light....................................... 160

Figure 8-26 Photo Bleaching at different intensity......................................................... 161

Figure 8-27 Photo Bleaching on more different intensity .............................................. 161

Figure 8-28 Comparison of Photo Bleaching rates......................................................... 162

Figure 8-29 Intensity vs Concentration (nM) ................................................................. 163

xiii
ABSTRACT
Design, Simulation and Experimental Study of Shape Memory Alloy
and Micro-motor Activated High Pressure Optical Cell for Bio-Physical Studies
Oliver Hongchun Xie
Prof. Jack G. Zhou
Prof. Parkson Chong

During the past two decades, bio-physicists have had an increasing interest in

finding out what happens when two bio-material solutions are mixed under high pressure.

Compared to temperature, pressure makes more contributions to our fundamental

understanding of the structure-function relationship of biological systems, because

pressure produces only volume changes under isothermal conditions, and pressure results

can then be interpreted in a more straightforward manner. As pressure brings molecules

closer and thus enhances intermolecular interactions, the previously unattainable

properties of biological molecules may be discovered by using high-pressure optical cells

(HPOC) technology.

Window-type HPOC such as the one designed by Paladini and Weber have

provided biophysicists with a powerful tool to understanding the structure-function

relationships of biological molecules. However, the conventional HPOC is only good for

single solution testing and does not allow for quick mixing and stirring of additional

components while the specimen is under pressure. To mix two solutions under pressure,

Zhou and Chong developed a laser activated dual chamber HPOC. However, the

expensive laser device and its unavailability in most laboratories make the application

difficult. Zhou and Chong also introduced shape memory alloy (SMA) as an actuator to

unplug a urethane stopper with a biasing spring for agitation. The drawback of the

biasing spring is that the spring blocks the observing light beam and creates the creation

xiv
of unwanted reflections, and the SMA spring actuator method has never been put to

practice.

To thoroughly study the feasibility of SMA as an actuator, five types of SMA

actuators were designed, simulated and tested for unplugging and mixing purposes. To

conduct this research, SMA helical springs were fabricated in house according to the

design requirements. With different combinations of SMA tensile springs, SMA

compressive spring and biasing spring, significant ranges of vibration were developed.

To further improving mixing process, a unique hybrid design of SMA as an actuator to

unplug the stopper and micro-motor as a stir device to agitate the solutions was

developed. Rapid mixing of 95% of total solution in 10 seconds was achieved under 300

bars. A new HPOC was designed according to the new cuvette with its new unplug and

mixing mechanism. Our industrial partner, ISS, further modified our design for easy

manufacturing reason and fabricated the HPOC which made SMA actuator mixing test

under pressure possible. Since the inside chamber height was increased to accommodate

the new cuvette and electrical leads were introduced through the cell body to achieve

remote control outside of the HPOC, FEA Method was used to assure the HPOC’s safety

under high pressure. Since the device is designed for bio-physical study, bio-

compatibility of Nitinol SMA with protein and phospholipids was studied. As SMA

actuator generates heat to function, heat transfer simulation was conducted to understand

the heat influence. The enzyme behavior under of applying different current to SMA

spring actuator was tested. A complete testing of the new HPOC system to observe bio-

reagent mixing and reaction under high pressure was conducted and the results were

satisfactory.

xv
1 Introduction

1.1 Problem Statement

The window type pressure cell is commonly used in high-pressure fluorescence

studies and fluorescence measurement is widely used bio-analytical tool with highest

sensitivity and versatility. The current high pressure optical cell (HPOC) doesn’t allow

two or multiple solutions in the same high pressure chamber and mix multi-solution

efficiently under high pressure.

1.2 Research Objective

The objective of this research is to design a smart activation system which

consists of SMA activator and micro-motor mixer and to build a SMA-activated dual

chamber HPOC, which can sustain a pressure up to ~3 kbar. The specific aims are (1) to

shorten the response time of the shape change of the SMA-plug mechanism so that the

initial rates of most enzymatic reactions can be detected under pressure, (2) to minimize

or mitigate the local heat effect caused by the electrically induced activation of SMA so

that the system being studied is subject to negligible perturbation on thermal equilibrium,

(3) to construct a convenient and reliable SMA plug for the use in the final dual chamber

HPOC, (4) if the proposed SMA activator cannot fulfill the mixing requirements, an

alternative micro-motor/mixer system will be introduced. (5) to test the bio-compatibility

of the SMA-plug mechanism and validity of the design for fluorescence measurements

under pressure and to demonstrate the applicability of the final cell in the pressure studies

of sterol superlattice in membranes. The Ni-Ti Alloy composition in the SMA plug will
be systematically varied. Various shapes of the SMA plug will be constructed and tested.

The interacting effects of pressure and cholesterol on the activity of snake venom

phospholipase A2 lipid membrances will be examined in the context of sterol superlattice

model.

1.3 Background and Review

1.3.1 Significance of HPOC on Bio-Physical Studies

The SMA-activated HPOC will have significant contributions to biological

research in at least three different areas: biomedical, biophysical, and bio-hyperbaric

research. Hydrostatic pressure, especially in combination with osmotic pressure and

mutagenesis, is a unique and powerful tool for studying molecular recognition because

pressure produces a volume change in the interfacial region where molecular recognition

occurs. (Robinson & Sligar, 1995). Molecular recognition plays a vital role in cellular

process. Malfunctions in molecular recognition lead to numerous disease states such as

flawed control of gene regulation and cellular profileration (resulting in cancer), and

defects in specific metabolic activities (e.g. defective LDL receptor resulting in

hypercholesterolemia). Through pressure studies, we can learn the forces, energies,

dynamics and structures involved in molecular recognition such as protein-protein,

protein-DNA, and protein-membrane interactions. In view of the close relationship

between molecular recognition and human disease, pressure studies should have a

fundamental and broad impact on our understanding of human health related problems.

The new pressure cell can also make important contributions in understanding

organism living under hyperbaric conditions. Since the late 1980s, several deep-sea

2
hydrothermal vents were discovered where surprisingly large amount of organism are

thriving. Using the type of HPOC proposed here we may eventually gain a fundamental

understanding as to how those organisms sustain high hydrostatic pressures whereas

surface dwelling organism cannot. There are also microorganisms living under varying

depths from soil surface. Clearly, pressure studies have a wide biological relevance.

At present, the use of pressure as a tool to explore these three areas is

underutilized. This situation arises mainly because the current pressure cells are

cumbersome, expensive and having limited versatility (e.g. unable to introduce additional

components into the sample chamber under pressure and unable to provide adequate

mixing and stirring). High-pressure stopped-flow devices (Goldsack et. Al., 1969;

Heremans et al., 1980; Hui Bon Hoa et al., 1982; Balny et al., 1984; Hara & Morishma,

1988; Bugnon et al., 1996) allow for the determination of the initial rate of the

biochemical reactions under pressure. However they are cumbersome to use because they

contain several components, all of which need to be carefully assembled each time before

placing in a high-pressure container. Furthermore, the commercial high-pressure stopped-

flow device is expensive (e.g. ~$70,000) and it not readily compatible with most of the

fluorometers commonly used in research laboratories. Due to the high cost and structure

complexity, only a handful of laboratories used the commercial high-pressure stopped

flow cells.

The SMA-activated HPOC offers a simple and economic way to solve these

problems. The market price of the SMA-activated HPOC is estimated to be below

$10,000. The new cell will allow the addition of one or more bio-chemicals into a sample

while all the components are under the same pressure and temperature. This design also

3
allows for a thorough mixing of those components within a short period of time. In doing

so, the effects of cofactors and regulators on a given biological molecule at high

pressures can be monitored, simultaneously or sequentially, via fluorescence, absorption

or light scattering. Since we use the properties of smart materials, rather than complex

mechanisms, our design makes the mixing and stirring process simple, convenient,

reliable and easy to operate. This design will greatly increase the scope and versatility of

the pressure cell originally designed by Paladini and Weber. Since the Paladini/Weber

cell has already generated so much useful information, as mentioned earlier, the new

pressure cell should become an even more powerful tool in biological research.

High-Pressure Optical Cells has been a very import tool in biophysical studies for

more than twenty years ((Neuman, Kauzmann et al. 1973; Heremans 1982; Silva and

Weber 1986; Wong 1987; Lester 1989). Pressure studies provide a fundamental

understanding in the structure and functional relationship of biological molecules. Under

isothermal conditions, pressure only produces volume changes while under isobaric

conditions, while temperature not only produces volume changes but also changes in

thermal energy. Pressure results are more straightforward and easier to understand. Under

isothermal conditions, the reaction rate of equilibrium of the reaction can be studied

according to volume changes which is yielded by pressure (Royer, Weber et al. 1986;

Royer, Chakerian et al. 1990). The studies of biological functions under different

pressure can demonstrate which functions are dependent on the volume change of the

system and which are independent of it. For example, membrane transport phenomena is

somewhat dependant on volume change (Chong, Cossins et al. 1983; Chong and Weber

1983), but ordinary chemical reactions in which covalent bond exchanges takes place

4
exclusively are relatively insensitive to pressure (or volume change) (Weber and

Drickamer 1983). Since pressure shortens intermolecular distances, it enhances

intermolecular interactions. Thus, formerly unattainable properties of bio-molecules

maybe revealed at high pressure. Wong and Mantsch discovered five gel phases of

dipalmitoylphosphatidylcholine (a major phospholipid found in lung surfactant) at high

pressures (Wong and Mantsch 1985). Without pressure, it would be impossible to reveal

these gel phases.

Pressure studies often involve optical measurements, such as fluorescence,

absorption, Raman and infrared spectroscopy. These measurements provide a convenient

and sensitive method to monitor biological reactions in a continuous manner. A pressure

cell is required for these measurements. Generally, there are two kinds of cells: the

diamond anvil cell is mainly used for high-pressure infrared and two-photon microscopy

studies; a window-type high-pressure optical cell is appropriate for fluorescence,

absorption and scattering measurements in the cuvette. Since fluorescence is used for bio-

analysis due to its highest sensitivity and versatility, this research focuses on window-

type high-pressure optical cells.

1.3.2 Interest of Living Organism under Pressure

The history of high pressure research started from the living organisms found at

600 bars when the discovery of living organism at 6,000 meters below the surface of the

sea in 1882 triggered the curiosity of P. Regnard and A. Certes from France (Paladini,

1980). Following this, Regnard and Certes studied the behavior of whole animals under

pressure, and the effects on some organic materials such as milk, eggs, etc. From 1900 to

1950 more biological materials such as protein, viruses, antigens and antibodies, tumors

5
and tissues were studied under high pressure (Paladini, 1980). The effects of the pressure

on the materials were evident. But temperature was rarely considered in analyzing results.

The experiments usually measured particular biological activities or permanent change

after the pressure was released and the effects were mostly irreversible. However the

actual physical changes in the system under pressure were ignored. The research in that

period only proved that pressure had an effect over the systems studied but didn’t use

pressure as a thermodynamic variable to gather information on the physical nature of

systems subjected to it. Hydrostatic pressure is a scalar magnitude without direction when

it is applied to the media. Real time study when pressure applied to the system is

important to understand pressure influence on the system while is it applied. It has been

found that high pressure creates volume change. Several investigators studied application

of hydrostatic pressure from 10 bar to 20 kbar which produces reversible changes in

dilute protein (Villas-Boas, Silva et al. 1993). These studies mainly relied on the

observation of changes of absorption or emission of visible and ultraviolet light while the

sample was subjected to pressure (Weber and Drickamer 1983).

1974, Weber, et al observed that a series of pressure up to 10 kbars yields a ∆V° =-

4.3 ml/mole (Heremans 1982). It also has been approved that the covalent links between

the partners play no significant role in the interactions that lead to formation of the

stacked complex. Covalent bond lengths and angles remain unchanged over a range of

pressure much larger than the one employed. This un-compressibility of the covalent

bonds lengths is of fundamental importance in any analysis of the effect of pressure upon

proteins on any compact polymer structure determined by the configuration of a

6
convalently bonded chain. Inclusion complexes of benzenic or fluorescence changes

permit their easy study (Paladini 1980).

1.3.3 Weber’s High Pressure Bomb

The interest in measuring fluorescence depolarization of protein solution

motivated Weber to design and construct a high-pressure bomb to meet the experimental

requirements. The bomb was designed to meet 3 criteria: hold pressure up to 2 kbars;

with four optical ports and one hydrostatic port to permit its pressurization; easy loading

and unloading samples and the prevention from making contact with either the inner

metallic bomb walls or the pressure transmitting fluid. For the efficiency of the pressure-

delivery pump consideration, the smallest internal volume is desired. This high pressure

bomb is made of a special steel alloy called Vascomax 300, developed by Teledyne

Vasco Company, Chicago. The steel can be hardened after machining to achieve

Rockwell C58 to C62, thus allowing the pressure cell dimensions to be small in

comparison with the dimensions required for standard steel were to be used instead of

Vascomax. A 2mm diameter conduit with 23 mm depth was drilled to accommodate a

thermocouple to monitor the temperature variation of the bomb. The temperature

monitored is on the inside temperature of the bomb instead of sample temperature since

an extra plug just for temperature monitoring is costly and difficult. The windows were

made of fused quartz T19 Suprasil 1, birefringence. The seal between the window and the

plug port was made by mirror polishing the metal port surface upon which the window

will rest. The seal is achieved when pressure press both surfaces against each other. The

mirror polishing of the plug surface is critical since any gap between quartz and plug port

will result in a leak under pressure.

7
Paladini and Weber’s HPOC has been widely used in fluorescence studies of

proteins, which have led to the discovery of conformational drift of oligomeric proteins

(King and Weber 1986; King and Weber 1986; Weber 1986; Ruan and Weber 1988;

Ruan and Weber 1989; Ruan and Weber 1993). It has also been used in protein folding

(Foguel and Weber 1995; Eftink, Ramsay et al. 1996; J. and A. 1998), protein-DNA

interactions (Royer, Ropp et al. 1992; Villas-Boas, Silva et al. 1993), enzyme kinetics of

membrane proteins (Chong, Fortes et al. 1985) and subunit-subunit interaction of proteins

(Valente-Mesquita, Botelho et al. 1998).

1.3.4 The Limitations of the Paladini/Weber High-Pressure Optical Cell

Despite its great contributions to the biophysical research, the type of pressure

cell made by Paladini and Weber has certain limitations. It does not allow quick mixing

and stirring of additional components under pressure. For example, when using the

Paladini/Weber cell to study the enzymatic activities of (Na,K)-ATPase in membrane

fragments at high pressures (Chong, 1985), the enzymatic hydrolysis reaction was

initiated by mixing the substrates, cofactors and enzyme together at atmospheric pressure.

Then the mixture was loaded into the quartz cuvette and the polyethylene tubing (used as

a cap of the cuvette). The cuvette plus the polyethylene tubing formed a closed system,

which was then transferred into the pressure cell and immediately thereafter pressure was

raised to the desired level. The entire process of sample loading, cuvette assembling, and

pressure generation took about several minutes. Finally, the time dependence of the

optical density from the reaction products was recorded which was reflective of the

apparent enzyme activity at the designated pressure and temperature.

8
The above operation has the following problems: (1) unless the optical change

with time remains in a linear fashion for >10 min., the operation not permit the

observation of the initial rates of the enzyme reactions. This is a serious problem

particularly for the studies of membrane surface acting enzymes because the activities of

those enzymes depend upon the initial membrane lipid composition (Liu & Chong, 1999);

after the reaction proceeds for a prolonged time, the exact lipid composition in the

membrane is not known. (2) This operation does not start with a true thermal equilibrium

and the enzymatic reaction proceeds prior to change become questionable. (3) With this

operation, maintaining a constant temperature for the sample before and after loading it

into the pressure cell is problematic. (4) The Paladini/Weber cell does not allow the

addition of other components into the sample while the sample is under pressure, which

limits the scope and versatility of the pressure experiment.

1.3.5 Zhou’s Effort to Mix Solution under Pressure

To solve current problems, Zhou et al proposed a laser-activated stopper system

(Zhou, Ostrow et al. 1998). This device made it possible to mix two samples under high

pressure. However, this device requires the use of an UV laser which is costly and not

readily available in most laboratories. Another drawback is that this device does not

provide sample mixing. To further deal with above problems, Zhou et al proposed to use

shape memory alloy (SMA) as an actuator for both unplugging stopper and mixing

solutions, they came out two designs: a one-way SMA spring plus a bias extension spring;

and, a two-way SMA spring (Zhou, Chong et al. 2000). Both designs have drawbacks in

that the first design suffers from obstruction of optical observation, due to the use of the

9
bias spring blocked the observation window, and both designs were never materialized,

although theoretically stated.

1.4 Research Achievements

A new HPOC was designed based on Paladini’s high pressure cell (Figure 1-1).

The new cell accommodates a dual chamber cuvette which can separate two solutions

after the system is assembled and pressure is raised to desired level. A smart activation

system resides in the cuvette upper chamber. Electrical lead plug provides electrical

supply to the activation system from outside the HPOC.

Figure 1-1 The design of HPOC, a transparent 3-D view

10
1.4.1 Design of the New SMA Activated Dual Chamber HPOC

The design motivation of the cell is the introduction of biological samples until

the experimenter is ready to mix them. Prior mixing, all the samples experience an

identical pressure and the same rate of pressure increase. A specially designed dual-

chamber quartz cuvette holds two samples and locates at the center of the pressure

chamber. Several cuvettes were designed to fit each of the actuator design. A final HPOC

was designed to fit the latest cuvette. To help us to fabricate the HPOC, ISS Modified our

design to fit with their existing cell body. ISS used their existing HPOC as the base,

modified the top inlet plug to accommodate both Electrical lead and inlet. This design

saved a lot of manufacture cost, and made our final testing under pressure possible.

1.4.2 Design of Shape Memory Alloy Activated Stopper and Mixing Systems

This research designed and fabricated different actuators with different

combination of SMA springs and micro-motor actuators. The main rule is that the design

should be as simple as possible: if less components are introduced to the cuvette, it will

be easier to operate in bio-physical studies. The first design involves the use of a two-way

SMA wire as an actuator. A two-way SMA remembers both a hot (austenite) and a cold

(martensite) shape. This property is used in order to eliminate the extension spring used

in Zhou’s dual spring design. With this design, it is understood that the agitation of the

two samples can be accomplished without the use of extension spring. This two-way

SMA is trained to a hot length and a cold length. The wire will first unplug the stopper

and then act as an agitator by moving up and down using its Two-Way Shape Memory

Effect (TWSME). TWSME of SMA wire can be trained to remember its high

temperature shape and also its low temperature shape (Johnson 1984). The main

11
drawbacks are that the vibration range is too small and TWSME is not reliable. The

second design goes back to SMA springs as actuator with commercially available

resources. This design uses two tensile SMA springs as unplugging and agitation. This

design can do unplugging and provide vibrations, but has its shortcoming of interference

in the lower observation path, since one SMA spring resides in the lower chamber of the

cuvette. This design is mainly to test SMA spring’s possibility as actuator. Both tensile

and compressive SMA springs were bought for testing and tensile SMA spring actuator

worked well both on unplugging and vibration. Unfortunately, the compressive SMA

spring wasn’t usable at all because it lost memory effect very easily. The third design

comes with tensile SMA to pull plug and compressive SMA spring as biasing spring to

create vibration. Since commercially available SMA springs are very limited and

compressive SMA spring is very unreliable, we manufactured our own SMA springs. By

alternatively applying current to either SMA tensile spring or SMA compressive spring,

their phase was switched from their martensite to their austenite phase. Therefore using

both SMA springs we are then able to maximize the vibration range by fully using their

difference in modulus of high or low temperature. Due to heat interfere of Duo-SMA

spring actuator in fourth design which lead to the other cooling spring not being able to

cool down, the fourth design uses a SMA tensile spring with a compressive biasing

spring. As above SMA actuators didn’t achieve rapid and complete mixing, we went with

our alternatively design, the fifth design which introduced a micro-motor as a mixer. The

design combined SMA tensile works as unplugging the stopper and a micro-motor as the

stopper as well as a mixer. The design achieved unplugging and mixing in one. The

12
complete mixing only took 10 seconds. And the heat generation can be controlled to

minimum, since SMA is only heated up on unplugging process.

1.4.3 HPOC Finalization and Fabrication

After SMA spring and micro-motor activated dual chamber cuvette functioned, a

new HPOC should be designed and fabricated according the new cuvette design. Since

the dual chamber is used in cuvette design and SMA based actuator needs space to

activate, the new HPOC size is increased. In addition to the ports required for the optical

observation of the samples, two additional ports are required for the transmission of

electrical current into the HPOC in order to activate the SMA actuator. These ports are

called electrical leads which can be achieved using a heat shrinkable plastic tubing (Terry

& Ruoff, 1972). A FEM analysis of stress and distortion will be conducted according to

the changes to guarantee the new cell can still hold pressure up to 3000 kbars.

1.4.4 Complete System Assembly and Final Testing

After the SMA based actuator can function as an unplugger and a stirrer, and dual

chamber cuvette as well as a new HPOC is fabricated, the system will be assembled and

tested under pressure with ISS K2 fluorescence measurement. The assembly of the

complete system involves the sealing of cuvette, the transfer of pressure into cuvette, and

function of SMA based actuator, etc. The final test is basically the whole system function

test, leakage of cuvette sealing under pressure, the unplugging under high pressure,

working of the stirrer on mixing under pressure, and optical observations of mixed bio-

solutions.

13
2 Design of SMA Actuator

This Chapter will explore SMA as an actuator. From two-way shape memory

actuator to SMA spring actuators, five kinds of SMA based actuators were developed in

improving rapid and complete mixing. Before getting into the design and fabrication of

SMA based actuators, some theory reviews of shape memory alloy are very necessary.

2.1 Review of Shape Memory Alloy Theory

SMA is often referred as smart material. Smart material has a prominent role in

the future for self-repairing and intelligent material (Hodgson, 1975). Smart materials are

a class of materials and composite media having inherent intelligence together with self-

adaptive capabilities to external stimuli. Also known as intelligent materials, they

constitute a few subsets of the material family that “manifest their own functions

intelligently depending on environmental changes.”

2.1.1 History of Shape Memory Alloy:

The first reported steps towards the discovery of the shape memory effect were

taken in 1930s. According to Otsuka and Wayman (Otsuka and Wayman 1998), A.

Olander discovered the pseudoelastic behavior of the Au-Cd alloy in 1932. Greninger &

Mooradian observed the formation and disappearance of a martensitic phase by

decreasing and increasing the temperature of a Cu-Zn alloy. The basic Phenomenon of

the memory effect governed by the thermoelastic behavior of the martensite phase was

widely reported a decade later by Kurdiunov & Khandros. Workers at the U.S. naval

Ordnance Laboratory discovered the shape memory effect in an equiatomic alloy of

14
nickel and titanium, which can be considered an alloy named Nitinol (Nickel-Titanium

Naval Ordnance Laboratory). Many efforts had been made since then, but not until the

mid-1990s, that the first breakthrough in medicine utilizing SMA in commercial stent

applications. Nitinol’s completely new properties of superelaticity and shape memory

effect, compare to conventional metal alloys, fascinate many biomaterial scientists.

(Harrison and Hodgson 1975)

2.1.2 Two Main Characteristics of Shape Memory Alloys

Two main characteristics of Shape Memory Alloys are shape memory effect and

pseudoelasticity. SMA is a unique class of alloys which are able to remember their shape

and are able to return to that shape even after being deformed. The phenomenon is called

Shape Memory Effect. Pseudoelasticity (Superelasticity) refers to the ability of NiTi to

return to its original shape upon unloading after a substantial deformation. (Funankubo

1987)

The superelastic effect of memory alloys is characterized by the impressive

amount of possible “elastic” strain, which is more than 20 times higher compared to

conventional materials, it is beyond elasticity, therefore called “superelasticity”. The

effect is based on an internal transformation of the crystal structure, which is called

austenite in the initial condition and which transform to the martensite (this phase is well-

known from steel hardening) if external stresses affect the material. The phase

transformation is related to the appearance of a pseudo-elastic strain, giving the effect its

name superelasticity. After releasing the stress the material recovers the initial shape and

temperature changes are not necessary for the occurrence of the effect. (Suzuki 1976;

Owen and al. 1979; Delaey and Chandrasekaran 1982; Tamura 1987)

15
At present, most interesting applications of the superelastic effect can be found in

medical implants and instruments, where for example the availability of thin-walled

tubing in the market of stents has opened for the memory alloys. Due to high elasticity

the stents can be folded into catheter sheeth and pushed into the vessel, where they

expand and provide a force to keep the vessel open. (Muddle 1990; Liu, Kunsmann et al.

1992; Wayman and Perkins 1993; George, Gotthardt et al. 1997)

A reversible, solid phase transformation known as mastensitic transformation is

the driving force behind shape memory alloys. When nickel and titanium atoms are

present in the alloy, in a given proportion, the material forms a crystal structure, which is

capable of undergoing a change from one form of crystal structure to another. The

transformation from one form of crystalline structure to another creates the mechanism

by which the shape change occurs in SMAs. This change involves transition from a

monoclinic crystal form (martensite) to an ordered cubic crystal form (Austenite). The

Austenite phase is stable at high temperature, and the martensite is stable at lower

temperature. In the martensite phase, atoms orient themselves in rows that are tilted left

or right. This phenomenon is referred to as twining, as the atoms form mirror images of

themselves or twins. The martensite twins are able to flip their orientation, in a simple

shearing motion, to the opposite tilt, creating a cooperative movement of the individual

twins. The atoms reorient themselves to straight up the lattice when the deformed SMA is

heated. Ordinary metals can’t reverse the deformation once deformation occurs by

dislocation motion and atomic planes sliding over one another, taking on a new crystal

position and results in permanent damage to the crystalline order. Unlike regular metals,

SMAs deform by detwinning which changes the tilt orientation of twins and doesn’t

16
cause any dislocation. Detwinning allows the martensite phase to absorb dislocation to a

given extent. Shape recovery only occurs to detwinning, but not to a deformation process

involving slip which is not irreversible. (Chu and Tu 1994; Ren and Otsuka 1997)

SMA in its Austenite phase exhibits a high elastic behavior. This allows the

material to deform up to 7% and then fully recover the resulting strain by simply

removing the load. This is psudoelastic effect or pseudoelasticity, it is also referred as

superelasticity. Deformed psudoelastic material is in the formation of martensite crystal,

it is called stress-induced martensite. Applications using the Shape Memory Effect results

of temperature induced transformation. The transformation temperature depends on the

composition of the alloy and can vary from 100°C down to cryogenic temperatures. (kato

and Mori 1976; Otsuka and Shimizu 1982; Pelton, Hodgson et al. 1995)

A set of characteristic temperatures of transformation are Martensite start

temperature (Ms), Martensite finish temperature (Mf), Austenite start temperature (As)

and Austenite finish temperature (Af). Martensite phase starts to form when temperature

drops below Ms and completes transformation when temperature drops below Mf.

Austenite phase does the opposite and start to form when the temperature is raised up to

As and finish transformation when temperature is above Af. There is a gap between Ms

and Af, it is called hysteresis. The temperature range for the martensite-to-austenite

transformation, i.e. soft-to hard transformation that takes place upon heating is somewhat

higher than that for the reverse transformation upon cooling. The difference between the

transition temperatures upon heating and cooling is called hysteresis. Hysteresis is

generally defined as the difference between the temperature at which the material is 50%

transformed to austenite upon heating and 50% transformed to martensite upon cooling.

17
The difference of heating and cooling temperature can be up to 20-30°C, which means

that an alloy designed to be completely transformed by body temperature upon heating

(Af < 37°C ) would require cooling to about as low as 7°C to fully retransform into

martensite (Mf). (Arneodo and Ahlers 1973)

2.1.3 Commercial SME Alloys

The only two alloys to have achieved level of commercial exploitation are NiTi

Alloys and the copper-base alloys. Properties of the two alloys are quite different. The

Niti alloys have greater shape memory strain (up to 8% versus 4 to 5% for the copper

base alloys), tend to be much more thermally stable, have excellent corrosion resistance

compared to the copper-base alloys medium corrosion resistance and susceptibility to

stress-corrosion cracking, and have much higher ductility. On the other hand, the copper-

base alloys are much less expensive, can be melted and extruded in air with ease, and

have a wider range of potential transformation temperatures. The two alloys both have

their own advantages and disadvantages for particular applications.

The basis of the nickel-titanium system of alloy is the binary, equiatomic

intermetallic compound of NiTi. The intermetallic compound is extraordinary because it

has moderate solubility range for excess nickel or titanium, as well as most other metallic

elements, and it also exhibits ductility comparable to most ordinary alloys. The solubility

allows alloying with many of the elements to modify both the mechanical properties and

transformation properties. Heat treating to impart the desired memory shape is often done

at 500 to 800°C (950 to 1450°F), but it can be done at 300 to 350°C (600 to 650°F) if

18
sufficient time is allowed. The SMA component may need to be restrained in the desired

memory shape during the heat treatment; otherwise it may not remain there.

Commercial copper-base shape memory alloy are available in ternary CuZnAl

and CuAlNi alloys, or in their quaternary modifications containing manganese. Elements

such as boron, cerium, cobalt, iron, titanium, vanadium and zirconium are also added for

grain refinement. The thermal stability of copper-base alloys is ultimately limited by the

decomposition kinetics. For this reason, prolonged exposure of CuZnAl and CuAlNi

alloys at temperatures above 150°C and 200°C respectively, should be avoided. Aging at

lower temperature may also shift the transformation temperatures. In the case of aging the

beta phase, the results from the change is in long-range order (Schofield and Miodownik

1980). When aged in the martensitic state, the alloys exhibit an aging induced martensite

stabilization effect (Ahlers 1986).

2.1.4 Metallurgical Property of NiTi Alloy:

What make SMAs unlike other metallic materials which induce large amount of

deformation without permanent deformation? How can they sustain large deformations

and recover their strain upon temperature change or loading? The unique characteristic of

SMAs is due to their phase transformation mechanism. Some essential metallurgy can

explain these characteristics. When the atomic percentage of Nickel in Nitinol is close to

50%, the structure is called near-equiatomic. The equilibrium Ni-Ti alloys make the

shape memory effect and transformation pseudoelasticity occurs. The atomic structure

undergoes thermoelastic martensitic transformation from parent phase (β) with a B2

structure to the phase with a monoclinic B19’ structure, or to more often the two-step

transformation from the β to a trigonal phase (so called R-phase) and then to the B19’

19
phase. Ni content, aging thermo-mechanical treatment and addition of alloying elements

affect structure and therefore influence on controlling the memory behavior.

Equilibrium Ni-Ti phase diagram in the vicinity of NiTi shows the B2 region is

very narrow at the temperature below 923K (650°C). B2 region below 923K is generally

accepted only between 50.00 and 50.5 at% Ni. When nickel contents exceeds 50.5at%,

Ti-Ni alloys decompose on cooling slowly from a high temperature or on aging at a

temperature below 973K (700°C) after quenching from a high temperature, the final

product of the composition is a mixture of TiNi3 and TiNi. In the early stage of aging at

low temperatures, a metastable phase Ti3Ni4 forms as fine platelets with coherency to the

matrix and affects the properties of the TiNi alloys. Ti3Ni4 has a rhombohedral structure,

the space group R3. The unit cell of this structure contains 18 titanium atoms and 24

nickel atoms. The precipitate particles of Ti3Ni4 are lenticular shape and coherent to the

matrix in early stage and produce strain fields around them. These strain fields produce

the all-round shape memory effect. Precipitation of Ti3Ni4 strengthens the matrix B2

phase and improves the recoverability of the shape memory effect.

Fully annealed near-equiatomic Ti-Ni alloys transform from the B2 parent phase

to monoclinic B19’ phase martensitically; thermo mechanical treated near-equiatomic Ti-

Ni alloys transform in two steps, from the B2 parent to the R-phase and then to the B19’

phase. B19’ martensite has a monoclinic structure and R-phase has a trigonal structure or

a hexagonal lattice. R-phase transformation also causes the resisitivity increase.

2.1.5 Thermoelastic Martensite Transformation

The unique behavior of NiTi is based on the temperature-dependant austenite-to-

martensite phase transformation on an atomic scale, which is also called thermoelastic

20
martensitic transformation. The thermoelastic martensitic transformation causing the

shape memory recovery is a result of the need of the crystal lattice structure to

accommodate to the minimum energy state for a given temperature. In NiTi, the relative

symmetries between the two phases lead to a highly ordered transformation, where the

displacements of individual atoms can be accurately predicted and eventually lead to a

shape change on a macroscopic scale. The crystal structure of martensite is relatively less

symmetric compared to that of the parent phase. If a single crystal of parent phase is

cooled below Mf, then martensite variants with a total of 24 crystallographically

equivalent habit planes are generally created. There is however, only one possible parent

phase (austenite) orientation, and all martensitic configurations revert to that single

defined structure and shape upon heating above Af. The mechanism by which single

martensite variants deform is called twinning, and it can be described as a mirror

symmetry displacement of atoms across a particular atom plane, the twinning plane. Not

like most metals deform by slip or dislocation, NiTi responds to stress by simply

changing the orientation of its crystal structure through the movement of twin boundaries.

A NiTi specimen will deform until it consists only of the correspondence variant which

produces maximum strain. However, deformation beyond this will result in classical

plastic deformation by slip, which is irrecoverable and therefore has no “memory effect”.

If the deformation is halted in the midway, the specimen will contain a parent phase with

an orientation identical to that existing prior to the lattice correspondences between the

original parent phase and each variant martensite has a more complex rhombic structure.

This phenomenon causes the specimen to revert completely to the shape it had before the

deformation.

21
Shape memory alloys (frequently referred to as smart materials), such as nickel-

titanium (Ni-Ti), undergo a phase transformation in their crystal structure when it is

heated from the weaker, low temperature form (martensite) to the stronger, high

temperature form (austenite). Nitinol with 55% Ni and 45% Ti has mechanical properties

as shown in Table 1. It can be seen from Table 1 that nitinol is much easier to deform at

low temperature than that at high temperature due to its low shear modulus and Young’s

modulus. Figure 2-1 shows phase transformation versus temperature curve. Hysteresis of

temperature transformation, T1 is around 20 to 40°C for most SMAs. By heating nitinol,

austenite starts to form when temperature reach As (austenite start temperature) and

finish the transformation process at Af (austenite finish temperature). In cooling process,

martensite start to form at Ms (martensite start temperature) and finish at Mf (martensite

finish temperature).

Figure 2-1 Phase transformation versus temperature curve

(Harrison and Hodgson 1975)

22
Table 1 Nitinol mechanical properties

Temperature Lower than Mf Higher than Af


Yield Strength(Mpa) 100 560
Young’s Modulus 28 75
(Gpa)
Shear Modulus(Gpa) 10.8 28.8

Figure 2-2 shows stress and strain curve of transformation. In heating process, as

temperature increases, strain recovers slightly before As and recovers significantly after

Af .

Figure 2-2 Martensite-Austenite transformation σ-ε(Harrison and Hodgson 1975)

The shape memory transformation has been observed for many years. (Buehler;

Bobrowsky 1968; Hodgson, Wu et al. 1989) This inherent phase transformation is the

basis for the unique properties of these alloys, in particular, shapes memory and super-

elasticity. Generally, these materials can be plastically deformed at some relatively low

temperature, and upon exposure to some higher temperature will return to their original

shape prior to the deformation. The martensite is easily deformed to several percent of its

23
original length and shape at quite a low stress, whereas the austenite possessing a higher

strength is more difficult to deform.

Under uni-axial loading conditions, three different phases can be observed in a

shape memory alloy Figure 2-3, the austenite phase A and two martensitic twin variants

M±. . From crystallographic observations it is known that a macroscopic body under

uniaxial loading forms a layered structure (Chang and Read 1951), each layer being in

one of the three above phases. Motivated by this observation, the SMA shape

transformation process can be described as follows Figure 2-4. The far left is in

martensite phase at low temperature with M+ and M-twined next to each layer. When a

uni-axial force is applied, the twined structure will change to a single direction, either M+

or M- Figure 2-3. By heating up, the phase transformation will be achieved following

Figure 2-1 and Figure 2-2. In the heating process, Nitinol material can generate high

force and strain recovery. In the cooling process, A is simply turned to M+ and M- and

does not produce any force.

Figure 2-3 Martensite and Austenite (Ryhänen)

24
Figure 2-4 The shape transformation process (Ryhänen)

2.1.6 One Way Shape Memory

In most cases, the memory effect is one-way. That is, upon cooling, a shape

memory alloy does not undergo any shape change, even though the structure changes to

martensite. When the martensite is strained up to several percent, however, that strain is

retained until the material is heated, at which time shape recovery occurs. Upon cooling,

the material does not spontaneously change shape, but must be deliberately strained if

shape recovery occurs. Upon recooling, the material does not spontaneously change

shape, but must be deliberately strained if shape recovery is again desired (Foos, Frantz et

al. 1975; Harrison and Hodgson 1975).

2.1.7 Two Way Shape Memory

It is possible in some of the shape memory alloys to demonstrate two-way shape

memory ability in which the shape change occurs upon both heating and cooling. The

25
amount of this shape change is always significantly less than the change obtained with

one-way shape memory, and very little stress can exerted by the alloy as it tries to assume

its low-temperature shape. The heating shape change can still exert very high forces, as

with the one-way memory.

A number of heat-treatment and mechanical training methods have been proposed

to create the two-way shape memory effect (Johnson 1972; Wilson 1972). All these

methods rely on the introduction of microstructural stress concentrations which cause the

martensite plates to initiate particular directions when they form upon cooling, resulting

in an overall net shape change in the desired direction.

Two-way shape memory effect is that the shape memory element will return to a

low temperature shape on cooling as well as to a high temperature shape on heating.

However during cooling with the two-way effect, the material simply recovers its low

temperature shape but can not exert a force to the external mechanical components

(Warram 1993; Takagi, Khovalio et al. 2003). Without the external force, it is difficult to

achieve the two-way shape memory effect. In this case, two-way shape memory is

unreliable.

Johnson published a patent on preparing a two-way memory alloy (Johnson 1984).

A two-way shape memory alloy can be trained in following process: the alloy is heated

under a first training stress to a temperature which is above the upper operating

temperature of the alloy, then cooled to a cold temperature below the zero-force transition

temperature of the alloy, then deformed while applying a second training stress which is

greater in magnitude than the stress at which the alloy is to be operated, then heated back

to the hot temperature, changing from the second training stress back to the first training

26
stress. Huang and Toh also showed the feasibility of using reheat treatment to obtain two-

way shape memory effect (Huang and Toh 2000). By conducting reheat treatment on

shape memory alloy, two-way shape memory was introduced into initially one-way shape

memory alloy as indicated by the change of the diameter cycling. By embedding SMA

wire with two-way shape memory effect in silicone elastomer cylindrical rods, the shape

of elastomeric structure can be significantly changed. (Blonk and Lagoudas 1998)

Thin-film NiTi SMA membrane exhibits the two-way effect (Ma and Wang 2000;

Gill and Ho 2002). Thin-film NiTi SMA wasis sputtered-deposited onto a silicon wafer in

ultrahigh Vacuum. Transformation temperature of NiTi film was measured by monitoring

the change of residual stress with varying temperature in their research.

2.1.8 Heat Treating and Shape Setting of SMA

Nitinol and other shape memory alloy mill products - bar, wire, ribbon and sheet

are normally finished by cold working to achieve dimensional control and enhance

surface quality. Cold working suppresses the shape memory response of these alloys. It

also raises the strength and decreases the ductility. However, cold work does not raise

the stiffness of the material. Heat treating after cold working diminishes the effects of

cold working and restores the shape memory response of these alloys. Therefore, in

order to optimize the physical and mechanical properties of a Nitinol product and achieve

shape memory and /or superelasticity, the material is cold worked and heat treated.

The mill product supplier normally provides the material in the cold worked

condition. The maximum practical level of cold work will be limited by the alloy and by

the product section size. Binary superelastic NiTi alloy fine wires with As in the range of

27
–25 to +95°C are typically supplied with cold reduction after the final anneal in the range

of 30 to 50%. Higher reductions are sometimes used for very fine wires. These same

alloys will be limited to about 30% maximum cold reduction in larger diameter bar sizes.

Binary NiTi alloys with very low As in the range of –50 to –60°C will not sustain the

higher levels of cold work without cracking.

Both superelastic and shape memory properties are optimized by cold work and

heat treatment. This thermo-mechanical process is applied to all Nitinol alloys although

different amounts of cold work and different heat treatments may be used for different

alloys and property requirements. Shape setting is accomplished by deforming the Nitinol

to the shape of a desired component, constraining the Nitinol by clamping and then heat

treating. This is normally done with material in the cold worked condition, for example

cold drawn wire. However, annealed wire may be shape set with a subsequent lower

temperature heat treatment.

In shape setting cold worked material, care must be taken to limit deformation

strain to prevent cracking of the material. Another approach is to partially anneal the

wire prior to shape setting. Yet another option is to shape set in incremental steps. Smith

et al. reviewed the types of furnaces and fixture hardware or mandrels that have been

used in heat treatment. Many types of furnaces have been used including box furnaces,

continuous belt hearth furnaces, tube furnaces, heated platen presses, vacuum furnaces,

induction heaters, salt baths and fluidized bed furnaces. The electrical resistance of

Nitinol makes it a good candidate for self heating by electric current. Nitinol will be

oxidized when heat treated in air. Therefore, surface requirements and atmosphere

control are important considerations.

28
Shape setting can be done over a wide temperature range from 300°C to 900°C.

However, heat treating temperatures for binary NiTi alloys are usually chosen in the

narrower range of 325 to 525°C in order to optimize a combination of physical and

mechanical properties. Heat treating times are typically 5 minutes to 30 minutes.

Consideration must be given to the mass of the heat treating fixture as well as the mass of

the product. Sufficient time must be allowed in the furnace to get the entire mass to the

desired temperature.

The shape setting heat treatment changes the physical and mechanical properties

of Nitinol. Morgan and Broadly mapped the effect of temperature and time at

temperature on shape set wire properties. (Morgan and Broadley 2003) Their response

curves illustrate that physical and mechanical properties do not always change in the

same direction. Also, some properties are not monotonic functions of time at temperature.

For example, upper plateau stress goes through a minimum as a function of time at

temperature when heat treating a superelastic alloy in the range of 450°C to 550°C. This

can be understood in terms of the complex precipitation response of the nickel rich Ni –

Ti alloys.

Precipitation processes in nickel rich NiTi were studied in detail by Nishida et al.

Their TTT diagram shows that in a Ti – 52 atomic % Ni alloy heat treated below 820°C,

precipitation starts as fine Ti11Ni14 transitions over time to Ti2Ni3 and terminates after a

long time as TiNi3 in equilibrium with the NiTi matrix. All the while, the NiTi ratio in

the matrix is being shifted towards higher Ti content and higher transformation

temperature. Pelton et al. reported on the combined effects of non-isothermal and

isothermal heat treatment on the physical and mechanical properties of Nitinol wire. This

29
work suggests that for a 50.8 atomic % Ni alloy Ti11Ni14 dissolves at about 500°C and

Ti2Ni3 will start to precipitate at 550°C. This results a minimum or maximum in

properties as NiTi ratio in the matrix goes through a peak. Furthermore, this analysis

suggests that the temperature for transition from Ti11Ni14 precipitation to Ti2Ni3

precipitation occurs at higher temperatures for higher Ni content alloys.

Brailovski used measurements of latent heat measured by DSC and Vickers

hardness to map mechanical and physical properties as a function of heat treatment. He

obtained maximum fatigue performance when the combination of transformation

temperature and hardness were optimized.

With different compositions and heat treatment processes, different temperature

SMAs can be attained. Human body temperature (~37°C) SMA is widely used for

medical applications. As show in Figure 2-1, there is a difference of T1 between

martensite and austenite states. Body temperature SMA works best from room

temperature to body temperature. Some companies shortened the temperature

transformation hystersis of Nitinol by adding additives to 15°C, which brought the

application working perfect between room temperature (22°C) to body temperature

(37°C).

2.1.9 The Limitations of Shape Memory Effect and Superelastic Behavior

About 8% strain can be recovered by unloading and heating. Strain above the

limiting value will remain as a permanent plastic deformation. The operating temperature

for shape memory devices must not move significantly away from the transformation

range, or else the shape memory characteristics maybe altered. A shape memory NiTi

implant must be deformed at a temperature below As (usually <+5°C). Moreover, the

30
deformation limit determined by distinctive implant design (sharp angles, etc) and the

intrinsic strain tolerance of NiTi material must not be disregarded.

2.1.10 Application of SMA

Shape memory component can be designed to exert force over a considerable

range of motion, often for many cycles. Such an application is the circuit board edge

connector made by Beta Phase Inc. (Krumme 1987). In this electrical connector system,

the SMA component is used to force open a spring when the connector is heated. This

allows force –free insertion or withdrawal of a circuit board in the connector. Upon

cooling, the NiTi actuator becomes weaker and the spring easily deforms the actuator

while it closes tightly on the circuit board and forms the connections. Based on the same

principle, CuZnAl shape memory alloys have found several applications in this area.

(Waldbusser 1987) Among the SMA force actuator, SMA spring is mostly used. (Baz,

Iman et al. 1990; Lee and Lee 2000)

It is possible to use only a part of the shape recovery to accurately position a

mechanism by using only a selected portion of the recovery because the transformation

occurs over a range of temperature rather than at a single temperature. A device has been

developed by Beta Phase Inc. (Hodgson 1988) in which a valve controls the rate of fluid

flow by carefully heating a shape-memory alloy component just enough to close the

valve the desired amount.

A number of products have been brought to market that use the psudoelastic

(superelastic) properties of these alloys. Eyeglasses frames use superelastic NiTi to

absorb large deformations without damaging the frames are now marketed, and guide

wires developed for steering catheters into vessel in the body using NiTi wire, which

31
resists permanent deformation if bent severely. Arch wires for orthodontic correction

using NiTi have been used for many years to give greater rapid movement of teeth.

2.2 Challenges and Technical Approaches on SMA Based Actuator Design

The first challenge is the transformation temperature of SMA. The transformation

temperature of the Ni-Ti Alloy (composition: 51% Ni and 49% Ti) used in Zhou’s SMA

proposal was 80°C, which was too high for normal biological studies. We have to find

body temperature SMA commercially available with an economic price. The temperature

transformation hysteresis, T1, for most Nitinol alloy is 20°C, which make the control of

temperature before and after activated extremely tight. As body temperature SMA is

selected and the activation temperature is around 37.5°C, the temperature needs to be

controlled under 17.5°C before activation and even on preparation of samples.

Fortunately, body temperature SMA wires were found, and more exciting things is that

with some additives, the temperature transformation hysteresis is reduced to 15°C from

some companies. The 5°C makes a lot of difference since 23.5°C is room temperature.

The second challenge is the reaction/response time and agitation frequency. A

lower transformation temperature of the alloy would also allow for a faster cycling

between heating and cooling, hence facilitating a thorough mixing of the samples.

Specially, in the future experiments on biological systems, we will use Ni-Ti alloys with

a transformation temperature 1~2°C above the desired experimental temperature which

ranges from room temperature to the physiological temperature 37°C. The

reaction/response time and agitation frequency are mainly determined by the

transformation rate at which heat can be delivered to the structure. Since we are using

electrical lead to generate and conduct the heat, it can be very quick. Shape recovers less

32
than one millisecond has been observed (Hodgson, 1988). If in the future study we can

develop a thick SMA wire which has a response/recovery time of 10 miliseconds, then a

thorough mixing and agitation, for example total of 10 agitation cycles can be finished in

100 miliseconds. Since the SMA and the surrounding biological molecules will be

exposed to the transformation temperature briefly, small local temperature perturbations

should have a negligible effect on the thermal equilibrium of the biological samples

examined. Notice that there will be a circulating bath controlling the temperature of the

pressure cell, the local heating problem can be eliminated or greatly reduced by selecting

an appropriate Ni-Ti alloy. The heat transfer analysis will be conducted to find out if the

heating is local and also the heating time and cooling time

The third challenge is two-way SMA. Although the two-way SMA has been

discussed for a long time and it has many advantages and potential applications, the two-

way SMA is still not commercially available. The main reasons are that the material

property and behavior of two-way SMA are still not clear and not easy to control, and

there is not a standard way to fabricate two-way SMA. In this study we will conduct

extensive materials study to find the composition relations, mechanics properties, and

structure behaviors of the two-way SMA. To create two-way SMA in Ni-Ti alloys

involves a somewhat complex training process and can use the following methods

(Duerig, Metton, 1989): overdeformation method, shape memory cycling method, and

combined shape memory and psudoelastic cycling method.

The fouth challenge is pulling-up force. The pulling-up force produced by the

SMA wire can be calculated. The calculation helps to select a right SMA wire size and to

program the SMA change that is required to overcome the weight of liquid on the plug

33
and the frictional force between the urethane plug and the cuvette. In this calculation we

need to consider the sealing pressure and the friction coefficient between the cuvette neck

and the urethane plug. We also need to carefully design a SMA wire in terms of its length,

wire diameter, and material strength, so it will be able to pull off the plug and generate

agitation. Since in the cuvette, the top and bottom of the plug are under equal pressure,

there is no need to consider the pulling-up force at elevated pressure.

2.3 Various SMA Actuator Design

The design of SMA Actuator starts at the Two-Way Shape Memory Effect

(TWSME). The SMA spring comes into the design with different combinations.

Commercial available SMA springs were purchased from Robot Store to test the concept.

Due to its size and available compressive spring wasn’t usable for use, Nitinol wire was

purchased to make our own SMA springs instead.

2.3.1 Design 1: Two-way Shape Memory Actuator

2.3.1.1 Design Concept

The first design (Figure 2-5) is using two-way shape memory effect (TWSME) to

design a simple structure with one SMA string to pull the stopper and then using the

TWSME effect to make the stopper vibrate up and down by supplying square wave

current to the SMA string. While the string is heating up, it will shrink and pull the

stopper up. While the current is cut off and the temperature will drop, the SMA wire will

return to its Martensite phase alternate shape which is longer and drop the stopper

downward.

34
Figure 2-5 Two-Way SMA actuator design

2.3.1.2 How to Train Two-Way Shape Memory Effect

The ability of shape memory alloys to recover a preset shape upon heating above

its transformation temperatures and return to an alternate shape upon cooling is known as

two-way memory. Two-way memory is unique in that the material “remember” different

high temperature and low temperature shapes. Much has been written about this unique

phenomenon in the literature and it has triggered many ideas in the minds of product

designers. Creating two-way memory in NiTi alloys involves a somewhat complex

process using one of the following approaches:

z Overdeformation while in the Martensitic condition.

z Shape memory cycling (Cool->Deform->Heat->Repeat).

z Pseudoeleatic cycling (Load-> Unload -> Repeat).

z Combined shape memory and pseudoelastic cycling.

z Constrained temperature cycling of deformed martensite.

z Constrained aging for long periods of time (to yield the all-round shape

35
effect which is slightly different than two-way memory).

2.3.1.3 The Limitation of Two-Way Shape Memory Effect

However, there are a number of limitations which must be addressed before

attempting to exploit two-way memory behavior, including:

z The amount of recoverable strain is generally about 2%, which is much

lower than that which is achievable in one-way memory (6 to 8%).

z The transformation forces upon cooling are extremely low.

z The memory can be erased with very slight overheating (as low as 250˚C).

z The long-term fatigue and stability characteristics are not well known.

z The inherent temperature hysteresis between the heating and cooling

transformation is still present.

Wherever possible, it is better to modify the device design to make use of one

way memory with biasing force acting against the shape memory element to return it

upon cooling. All of above limitations are addressed by this approach except for the

inherent temperature hysteresis, which remains. Two-way actuators using one-way shape

memory elements acting against bias forces have demonstrated large strains, high forces

in both heating and cooling directions, and excellent long-term stability equal to millions

of cycles.

Two-way shape memory effect is that the shape memory element will return to a

low temperature shape on cooling as well as to a high temperature shape on heating.

However, during cooling with the two-way effect, the material simply recovers its low

temperature shape but can not exert a force to the external mechanical components.

Without the external force, it is difficult to achieve the two-way shape memory effect. In

36
this case, two-way shape memory is unreliable for this design. Another limitation is the

deformation of 2% which is way too less for our application, which requires more than

15% at the least. This brings us back to the SMA springs which provide larger

deformation difference.

2.3.1.4 Conclusion of the Design

This design is not practical main for two reasons: 2% shape change will lead to a

very long cuvette to unplug a 5 mm stopper (at least 250 mm long for the upper chamber);

the shape memory effect is not stable on cooling process and its cooling shape

transformation doesn’t produce any force, which is a big problem for accelerate the

motion of the stopper and the agitation cannot be realized. The failure of this design leads

us back to One-Way Shape Memory Effect designs.

2.3.2 Design 2: Two Commercial Tensile SMA Springs as Actuator

2.3.2.1 Design Concept

To test SMA spring actuator concept, we found some commercial available

product from the robotics store. This design (Figure 2-6) was used only to test the

concept using two tensile SMA springs as an actuator. By apply current alternatively to

either spring, one is heating up while the other one is cooling down, the maximum

vibration range is achieved. The motivation to use two SMA springs instead of one SMA

spring and one biasing spring is to utilize SMA spring’s difference between high Young’s

modulus on high temperature Austenite phase and the low Young’s Modulus on low

temperature Marstensite phase. The difference of Young’s Modulus between two phases

is eight times.

37
Figure 2-6 Dual SMA spring actuator design

2.3.2.2 Background Theory on Spring Actuator Design

The use of a shape memory alloy in the form of a helical spring appears to have

the most promise in actuator applications. A shape memory spring has reasonable

force/motion characteristics, a compact size, and simplicity of design. The force that a

spring of any material produces at a given deflection depends linearly on the shear

modulus (rigidity) of the material. Shape memory alloys exhibit a large temperature

dependence on the material shear modulus, which increases from low to high temperature.

Therefore, as the temperature is increased the force exerted by a shape memory spring

increases dramatically. (Warram 1993)

The mechanical behavior of shape memory alloys can be described by a three

dimensional plot of stress, strain, and temperature, as shown in Figure 2-7. In fact,

simultaneous variation of stress, strain, and temperature results in two surfaces, one for

heating and the other for cooling; although only one surface is shown in Figure 2-7 for

38
clarity. The two surfaces enclose a three dimensional envelope.

Figure 2-7 Shape Memory Stress/ Strain/Temperature behavior (Warram 1993)

When the envelope is cut by a plane perpendicular to the strain axis, the points of

intersection form the stress-temperature hysteresis curve at the particular value of the

strain where the plane lies. In a shape memory actuator, this hysteresis would correspond

to a force-temperature hysteresis, at constant deflection (refer to Table 1 Nitinol

mechanical properties). When the envelope is cut by a plane perpendicular to the stress

axis, the points of intersection form the strain-temperature hysteresis curve for the stress

value where the plane intersects the stress axis.

39
Figure 2-8 Constant deflection configuration (Warram 1993)

Figure 2-9 Constant deflection hysteresis behavior (Warram 1993)

In a shape memory actuator, this would correspond to a moniton-temperature

hysteresis curve, under a constant force loading (refer to Figure 2-10, Figure 2-11 and

Figure 2-12).

40
Figure 2-10 Constant force loading of a shape memory compression spring

(Warram 1993)

Figure 2-11 Force-Deflection behavior for constant force operation(Warram 1993)

41
Figure 2-12 Superposition of linear Force-Deflection biasing spring (Warram, 1993)

When the envelope is cut by plane in an oblique fashion, simultaneous variation

of stress, strain, and temperature occurs, as in the case of a shape memory spring working

against a biasing force with a corresponding variation of force, motion, and temperature

Figure 2-12. For engineering purposes, it is convenient to plot a number of shear stress -

shear strain curves measured at various temperatures, rather than to deal with a three

dimensional plot. These stress – strain curves are referred to as isotherms, since they are

measured at constant temperatures (Figure 2-13).

42
Figure 2-13 Typical stress-strain behavior of Shape Memory Alloy material

(Warram, 1993)

The stress-strain isotherms are constructed by measuring the force-deflection

behavior of a test spring, at several temperatures, and by then converting the force-

deflection data to stress-strain data, using the following formulas:

dδ (2-1)
Shear _ strain = γ =
πD 2 n

2 D ⎛ ∆P ⎞
shear _ stress = T = 3 ⎜
δ + 3P ⎟
πd ⎝ ∆δ ⎠ (2-2)

Where: D = average spring diameter

d = wire diameter

n =number of active turns

P = force

43
δ = deflection

∆P
∆δ = slope of force-deflection curve

Figure 2-14 shows how the data are measured for one point on a force-deflection

curve. Higher values of strain in Figure 2-13 and Figure 2-14, the stress-strain curves are

nonl-linear, thus facilitating the use of the slope parameter to account for the curvature.

The design of shape memory actuators which utilize the non-linear portion of the stress-

strain curve, particularly Cu-Zn-Al springs, is mathematically complex, and an empirical

design technique has been developed.

Figure 2-14 Measurement parameter for Force-Deflection curve

Shape memory alloys, such as nickel-titanium (Ni-Ti), undergo a phase

transformation in their crystal structure when it is heated from the weaker, low

temperature martensite form to the stronger, high temperature austenite form. Nitinol

with 55% Ni and 45% Ti has mechanical properties as shown in table 1. It can be seen

from table 1 that nitinol is much easier to deform at low temperature than at high

44
temperature due to its low shear modulus and Young’s modulus. Different diameter

spring forces are simulated using Matlab Figure 2-15, Figure 2-16 and Figure 2-17. The

minimum force needed to unplug the stopper is 3N. By using safety factor of 1.5, we

need 4.5N minimum to unplug. The bigger the wire diameter, the more heat generated.

Minimum wire diameter is more desirable. From Figure 2-15, to generate 4.5 N of force,

the extension of the spring needed is less than 10 mm for 10 coil spring with diameter of

0.135” of wire diameter of 0.019”. Increasing spring diameter and decreasing wire

diameter can both decrease force generation ability (refer to Figure 2-16). Figure 2-17

shows that 0.017 is a good candidate for our design.

Figure 2-15 Force of SMA at two phases (N/mm) D=0.135” d=0.019”

45
Figure 2-16 Force vs Extension at high and low temperature (D=0.156”; d=0.018”)

Figure 2-17 Force vs Extension at high and low temperature (D=0.156”; d=0.017”)

46
2.3.2.3 SMA Spring Actuator Dynamics

We can illustrate a dual spring actuator dynamic system in following equation.

The equation differs from regular spring dynamic in that its spring constant can change

according to spring temperature change.

⎛ (K − K ) (K ) ⎞ + K
⎜ h l
f ( vt ) + h
⎟ ( x − 10 )
l

⎝ 2 ⎠ 2
(2-3)
2
dx d x
− K biase ( x − 30 ) + λ = m
dt dt 2
Where, Kh denotes spring constant of SMA spring at its high temperature phase,

Kl denotes spring constant of SMA spring at its low temperature phase, and Kbiase denotes

spring constant of biasing spring. And,

Gh d 4
Kh = = 0.538 N / mm
8nD 3

Gl d 4
Kl = = 0.0710 N / mm
3nD 3

K biase = 0.28 N / mm

Young’s Modulus: Austenite ~12 x 106 psi

Martensite ~4-6 x 106 psi

Yield Strength: Austenite 28-100 x 103 psi

Martensite 10-20 x 103 psi

K=Gd/8C3N

Where C=D/d, D – mean coil diameter, d – wire diameter

E
G – shear Modulus µ=
2(1 + ν )

47
D=0.135 d=0.019

D= 0.156; d = 0.018;

D = 0.156 d = 0.017;

Equation below is a simplified version and uses sin(vt) equation to simulate the

force change.

dx d 2x
( K sma sin(vt ) + K )( x − 10) − K biase ( x − 30) + λ =m 2 (2-4)
dt dt

Kh = 0.5348; Kl = 0.0710; K= 0.28 N/mm

a = 4.8 mm; b = 30 mm; m = 1.27g

This system can be illustrated as Figure 2-18. By solving the above equation in

Matlab, vibration can be simulated as Figure 2-19, in which damping ratio of 0.1 was

selected. The simulation gives very encouraging result with 250Hz frequency and 25mm

vibration range. If we can apply input current on its natural frequency, resonance can be

used to achieve huge vibration, which will benefits mixing dramatically.

Figure 2-18 Spring dynamics diagram

48
Figure 2-19 Spring dynamics Matlab simulation plot

2.3.2.3.1 Equipment Limitation

Due to the limitation of current slow relay response and extra electrical lead, the

testing result was with 3mm vibration range and 2Hz frequency. A quick response relay

and elimination of electrical lead will improve the vibration. The following figures and

information show a part of the investigations concerning SMA springs. Different SMA

springs have been produced and tested for several companies. These two images indicate

the load-deformation-behavior of two different SMA springs with different diameters and

transition temperatures. At high temperatures (here 100°C) the spring acts like an

ordinary steel spring, but at low temperatures (here 23°C) the spring yields at a constant

load.

49
2.3.2.3.2 Spring Responses to Electrical Current in the Air

The most common way to heat up SMA wire is applying electrical current. To

understand response of SMA wire to electrical current helps to design SMA actuators.

The extension responses to current before the temperature reach austenite states. Figure

2-20 shows two SMA wires response to a 4V step voltage, and it can be seen that the

bigger diameter travels longer.

Figure 2-20 Heating and cooling time histories of actuators subjected to step voltage

changes of 4 volts (a) 0.75mm (b) 0.875mm (Baz, Iman et al. 1990)

50
Cooling is a very important issue with SMA actuator design. If the application is

in the air, circulation helps cooling efficient. Figure 2-21 shows that by increasing the air

velocity from 0 to 7.2, the cooling time reduces from 58 seconds to less than 10 seconds.

On a repeating heating and cooling cycle, cooling condition is usually set for application

conditions and usually decides the whole cycle time.

Figure 2-21 Effect of cooling strategy on heating and cooling time histories of 0.4mm

actuators subjected to step voltage of 10 volts (Baz, Iman et al. 1990)

51
Figure 2-21 shows 0.4mm actuators subjected to sinusoidal voltage excitations

with air cooling velocity = 7.2 m/s, as the frequency increases, the wire can’t fully cool

to its low temperature and its low temperature phase can’t fully recovered. By increasing

any amount frequency over 1/36Hz, it is sacrificing its full working displacement range.

But since when cooling approach its low temperature it slows down, the selection of

stopping point is critical for design problems.

Figure 2-22 Heating and cooling time histories of 0.4mm actuators subjected to

sinusoidal voltage excitations with air cooling velocity =7.2 m/s: (a) at 1/36Hz (b) at

1/20Hz (c) at 1/15Hz (Baz, Iman et al. 1990)

52
From the above illustration, it seems that cooling is big issue on SMA response

frequency. Also the above result is based on air cooling, our media is water based protein

and a cycling cooling bath. Our cooling is much faster, a heat transfer analysis or

temperature monitoring experiment should be conducted to find out our cooling time.

2.3.2.4 Dual Tensile SMA Spring Actuator Testing

To test two commercial spring actuator concept and to provide power and

frequency, a pulse generator and a thermo-monitor are constructed. A pulse generator is

constructed with a square wave generator chip, Oscilloscope (Tecktronix TDS2024), DC

power supply (Tenna with max 30volt or 30ampere), Relay(Omron 12VDC, 10Ampere),

circuit board, etc. To test the response of SMA spring, a circuit is built as in Figure 2-23.

The frequency can be adjusted by selecting R1, R2 and C values. R stands for resistor and

C stands for capacitor. The ratio of R1 over R2 will control the percentage of peak value.

Figure 2-23 Frequency generator circuit layout

Temperature Measurement is constructed using Oscilloscope (Tecktronix

TDS2024), Amplifier Circuit Board (Figure 2-23), Thermo couple with 0.005 mm in

53
diameter on the tip, DAQ card data collection with Labview analysis. To measure the

temperature, two methods will be used: Oscilloscope monitoring and DAQ card

collecting data with Labview. Both methods will use the following amplifier to convert

measured signal to a measurable value and cold junction compensation built in the circuit.

–1mv is comparable to 1°C.

Figure 2-24 (a) Circuit board and relay (b) Two SMA spring testing in water

It is very hard to measure the temperature of SMA wire, since the ambient cooling

effect is very fast. This experiment tried to get some idea of temperature response to

electrical current. Figure 2-25 shows the temperature in the adjoining water of SMA

54
spring, the temperature response to electrical current is not the temperature of SMA wire,

but the ambient. The result shows the temperature change, but it is far from 15 degree

different. It is not possible to inquire temperature data, so we need to use FEA method to

find out the temperature response to electrical current. FEA simulation will be shown in

following sections. It took carefully designing of FEM model and boundary condition to

get close predictions.

Figure 2-25 Temperature measurement in water close to spring

Using Vh/VL=2/3, the result is to achieve significant vibration (around 5mm)

(Table 1). When the period is decreased to 110 ms, the vibration range can’t be increased

by increasing current, heat is not dissolvable. Heat generation calculation can be

calculated as follow:

Q=Wt=I2Rt-K∆L=m hwater ∆T t (2-5)

55
Where R=0.8ohm; K unknown, but small, ∆L=5mm

Ignoring heat lose in the air. I, applied current; R, SMA spring resistor; K, SMA

spring coefficient; ∆L, spring travel length; h, convective heat transfer coefficient; ∆T,

temperature change; m, mass of water; t, time.

2.3.2.5 Conclusion

The SMA spring can unplug a stopper; the actuator can generate 10 mm vibration

range in open water container; the reaction/response frequency can achieve to 1 Hz; the

ambient water is heated up after 15-20 cycle to 5°C. Measuring temperature of SMA

spring is difficult since water cools the surface of SMA wire much quicker than air does.

Temperature FEM simulation is necessary.

2.3.3 Design 3: Actuator with self-developed SMA spring

To overcome the defect of second design, we put both springs on the same side to

leave the bottom chamber of cuvette clear for observation. The new design includes two

SMA springs, one tensile and one compressive. This will solve the problem of the

previous design in which the bottom tensile SMA spring blocks observation light and

interfere with the readout. We ordered some tensile and compressive SMA springs from

the Robotics Store, the tensile spring which had been utilized on previous design

wouldn’t fit into our small cuvette. The commercial compressive spring didn’t perform

very good SME, and lose its SME easily by a little stretch. Also, its diameter is way too

big for our utilization. After consulting with a few companies, which charge $2000 per

customized SMA springs, we decided to make our own springs according to our surveyed

research paper and patent. Both tensile and compressive springs were made by ourselves

56
and functioned well separately. When they work together, the heat transfer analysis

shows that it was very critical to set both springs position to avoid either of them work

improperly.

2.3.3.1 Design Concept

In this design, we have two self made SMA springs, one is tensile and the other is

compressive. The diagram of this design is show as Figure 2-26. With both spring on one

side, the bottom portion of the cuvette is cleared for observation light to pass through.

Figure 2-26 Dual SMA springs actuator with tensile and compressive SMA springs

2.3.3.2 Making of Our Own SMA Springs

To make SMA springs, a fixture will be used to keep the spring shape and heat

treatment will be conducted. Fixture materials must withstand the temperatures needed

for shape setting to provide the strength needed to maintain required geometries without

losing strength and hardness. At the same time, corrosion loss and oxidation must be

57
minimized. Figure 2-27 a) shows how to make a SMA spring: a SMA wire is wound to a

mandrel fixture according to desired pitch and both ends must be secured to fix the shape

during heat treatment (Smith and Hodgson 2003). After heat treatment, the wire will be

shaped as Figure 2-27 b).

Figure 2-27 Making SMA spring

Heat treatment will influence the Austenite final temperature (Af) and oxidation

(Morgan and Broadley 2003). The longer the heat treatment, the lower the Af and the

more oxidation. Figure 2-28 shows the relationship of Af with both heat treatment

58
temperature and time. The higher the heat treatment temperature, the lower the Af, The

composition of nickel in nitinol determines mostly the Af, but heat treatment temperature

can do small adjustment. Heat treatment time can also slightly adjust the Af in a very

limited range but will create more corrosion and oxidation. Heat treatment temperature

and time can also modify ultimate tensile stress Figure 2-28 (Morgan and Broadley 2003).

Figure 2-28 Surface response for Af Temperature and ultimate tensile stress

2.3.3.3 FEM Heat Transfer Analysis for SMA Tensile and Compressive Spring

The heat transfer process is simulated using FEMLab. A 2-D for 3-D problem

simplified structure is used for the model: two cylinder shape cross-sections represent

two Nitinol springs (the heat generator), quartz cuvette (close to insulator), metal cap,

fluid and Vascomax HPOC (Figure 2-29).

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Figure 2-29 Femlab heat transfer FEM Analysis for two SMA springs

The left is axial symmetric insulation boundary, the right and the top are air

convection cooling and the bottom is cooled by a cooling water pump. By applying

alternative heat flux to either of the springs, each spring will go through its heating and

cooling process. The distance of the tensile spring to the center and the distance of the

compressive spring to the quartz cuvette need to be adjusted carefully to achieve this

periodic high and low temperature pattern. Temperature changes are most likely to occur

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on both springs. Fluid and cell body do not have significant temperature rise. From the

simulation, the spring temperature in some cycles can reach to 370K which is undesirable

and much higher than 313K. The heat source needs to be optimized for each cycle and

variable amplitude of electrical current will be used. The close sine curve is the

temperature change of point on the SMA spring according to current change, the near

point still carry the sine curve with much smaller amplitude, and the far point is almost

not influenced buy cooling down with the cool bath. According to the simulation, the heat

can be dissipated to the environment and the testing solution temperature is kept (Figure

2-30).

Figure 2-30 Heat transfer simulation for duo-SMA spring actuator

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2.3.3.4 Conclusion

In this stage, SMA springs were fabricated by ourselves and their shape memory

response were very stable and repeatable both on tensile and compressive SMA springs.

We also got wide range of body temperature SMA wire from 0.012” to 0.18” and we can

make springs according to our needs and select the best fit diameter. With numerous

testing, we found out 0.014” in diameter wire performs the best in our design. It can

provide enough force to unplug and cooling faster due to its smaller surface area to

volume ratio. The actuator can produce 10mm range vibration with 1 Hz frequency in

open water. Comparing to commercial SMA spring, the spring length is much shorter, 10

rounds compare to 20 rounds, the vibration range was doubled and also the heat

generation is much smaller due to its smaller diameter wire. Assembly test wasn’t very

successful. When the actuator was assembled in cuvette, the heat generation was way

faster, after 5 to 8 cycle, the vibration stopped and this agrees with heat transfer analysis

result which shows that tensile and compressive springs were ever sensitive to their

position to each other and to cuvette too.

2.3.4 Design 4: SMA Extensive Spring and Biasing Compressive Spring

Actuator

2.3.4.1 Design Concept

Due to SMA compressive spring heat interfere with SMA extensive spring from

cooling down and reheat up, SMA compressive spring was replaced with a biasing

compressive spring (Figure 2-31).

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Figure 2-31 SMA actuator with tensile SMA and biasing compressive spring

2.3.4.2 FEM Heat Transfer Analysis (more detailed)

To investigate the heat transfer effect of SMA to the system, a more detailed FEM

model is build with Femlab Figure 2-32. The heat source is from electrical current going

through the SMA spring wire. Water based protein solution conduct convective and

conductive heat transfer process, also ethanol between cuvette and HPOC. HPOC is

cooled by a water bath in the bottom, with air circulation on the top and side. Since this is

2D-axialsymmetric model, the center line is symmetric boundary condition.

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Figure 2-32 Heat Transfer analysis result (whole layout)

The objective of this simulation is to study the heat transfer process and to find an

optimum heat source input to achieve the best performance. Overheating will destroy

protein, and under heating won’t make the spring function. First different currents from

1A to 5A is conducted to find out how fast the temperature of SMA wire can reach from

room temperature(22°C) to body temperature (37.5°C). 1A and 2 A was used for SMA

wire in the air usually (Figure 2-33).shows that using 1A to reach 40°C will take far more

than 5 seconds in water solution. Figure 2-34 shows that using 2A to reach 37.5°C takes a

little more than 5 seconds, still not fast enough.

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Figure 2-33 1A current, 5 seconds heating response

Figure 2-34 2A current 5 seconds heating response

65
3A electrical current is the turning point, Figure 2-38 shows that the heating time

is shorten to 2 second. This is more useful, but not good enough for our application in

that the heating response is not fast enough. 4A electrical current heats up the SMA wire

much faster, refer to Figure 2-36, it takes 0.5 second to get to 37.5°C. 4A electrical

current will be useful to keep the cycle under 1second. 5A electrical current response is

even more promising, refer to Figure 2-37, it takes around 0.25 second to reach body

temperature. We can keep these two current selections in mind for later application.

Figure 2-35 3A current 5 seconds heating response

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Figure 2-36 4A current, 5 seconds heating response

Figure 2-37 5A current, 5 seconds heating response

67
We feel confident that 5A electrical current for heating can produce promising

result. A closer look at the model, Figure 2-38 shows the result after 5 seconds by heating

up with 5A current. Figure 2-39 shows the temperature line plot, which the cross line

resides on the red line as show on Figure 2-38. After 10 seconds heating with 5A

electrical current, the center of the wire reaches 80°C, but its surrounding area

temperature drops dramatically as the distance is a few times of wire diameter. The

heating effect is considerably local. On the other hand, 5 seconds heating up constantly

with 5A current is undesirable, since the center reaches over 40°C which is unfavorable

to protein solution.

Figure 2-38 one coil cross-section of SMA wire close up heating condition

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Figure 2-39 Temperature line plot of crossing SMA center 5A, 5seconds heating

Now, let’s examine the cooling process. The SMA wire is heated up with 5A

electrical current for 0.25 second, and let the wire to cool down. Figure 2-40 shows that

the temperature of SMA wire reaches to 39°C in 0.25 second, the temperature drops

down to 25°C in 0.75 second. It looks that a whole cycle of heating and cooling can be in

exactly 1 second, which means that 1Hz of vibration frequency can be achieved.

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Figure 2-40 0.25 second, 5A heating and cooling process

Now, let’s examine 5A heat transfer analysis result. Another heat transfer model

is to simulate a periodic supply of 5A electrical current for 0.25 second each cycle for 10

second running time. The structure was kept as the previous one, so does boundary

conditions. The change is only the input. Figure 2-41 shows that cooling from 40°C to

25°C needs more than 0.75 seconds, while the heating up needs 0.25 seconds. The total

time comes to 1 second. The cooling time is the limitation to increase agitation frequency.

Referring (Figure 2-1) and (Figure 2-1), the direct current heating and cooling behavior

shows that there is a delay period which SMA absorbs heat to prepare phase

transformation. But the period is not either defined or provided with experimental data

for liquid cooling system. We give 15seconds more for both heating and cooling period.

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Figure 2-41 shows 1Hz heat transfer plot of the wire center temperature. From the second

cycle, the low temperature is higher than 25°C (Martensite Temperature of our Nitinol

Body Temperature wire), which means, that it would not fully return its martensite phase,

but stay in R-phase.

Figure 2-41 0.25 second heating, 1Hz and 10 cycles condition

In Figure 2-42, frequency is reduced to 0.5Hz, the wire is fully cooled down

under Martensite transformation temperature 25°C in the first cycle. After 5 cycles, the

peak temperature is only a little higher than 45°C, comparing 1Hz with reaching almost

50°C instead. The ideal frequency is 0.5Hz instead of 1Hz due to the heat effect.

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Figure 2-42 5A 0.25 second heating at 0.5Hz condition

Figure 2-43 shows the temperature line plot of temperature change of 5 cycle of

5A/0.5Hz/0.25s heating condition. The line plot crosses the cross-section 1 of SMA

spring wire from the center to the ambient as show in Figure 2-44.The ambient

temperature in solution is less influenced comparing to that in Figure 2-39. Figure 2-45

shows line plot of solution temperature between cross-section 1 and cross-sections 2 of

SMA wires parallel to the previous line as show in Figure 2-44. The center of the cuvette

is less influenced, and the high of temperature only reaches 22.7°C, less than 1°C rise.

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Figure 2-43 line plot at 5A 0.25 second heating 0.5Hz condition

Figure 2-44 Heat transfer analysis result for two coils of SMA wire close up

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Figure 2-45 Line plot between 2 coils of 5A, 0.25 seconds heating, 0.5 Hz

Figure 2-46 Temperature point plot at one wire diameter away from SMA coil (5A,

0.25 second heating, 0.5 Hz)

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Figure 2-46 shows close area temperature change. The point selected in within

distance of the diameter of the wire. The change of temperature follows the wire center,

but the amplitude is much smaller. Figure 2-47 show the temperature change out of 2

wire diameter distance, the heating process has almost no influence at all. Figure 2-48

shows the center line temperature ramp. The center is heated up by 5°C after 5 cycles

heating.

Figure 2-47 Temperature point plot at 10 times of wire diameter away from SMA

coil (5A, 0.25 second heating, 0.5 Hz)

There is some fine tuning of peak portion to keep the heating to minimum. By

reducing the peak time to 1/3 second, FEMLab won’t cooperate well, it refuse to produce

cycles after 3 rounds Figure 2-49. And the peak temperature doesn’t reduce much. Finer

mesh and smaller time interval was tried, but didn’t produce better result. So we are

going to stick to quarter second peak.

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Figure 2-48 Temperature point plot at the center of coil (5A, 0.25s heating, 0.5 Hz)

Figure 2-49 Temperature point plot (5A 0.33s heating 0.5Hz center)

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2.3.4.3 Mixing Test using Fluorescence Measurement

The heat transfer analysis shows that the best strategy is to use 5A current with

0.25 second heating and 0.5 Hz to create the biggest vibration. The design assembly is

ready for mixing test. Since the HPOC will be fabricate after the finalization of the

cuvette, the mixing test is conducted without HPOC. The set up can be referred in Figure

2-50. Carboxy-fluoresceine (CF) of 300 nM was filled in lower chamber of the cuvette

and the upper chamber was filled with pure water. The exciting light wavelength is 495

nm and the emission wavelength is 520 nm. The excitation light is hit in the middle of the

lower chamber, so is the observation lens is aligned in the middle of the lower chamber.

Figure 2-50 Mixing Measurement Setup

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The mixing test result can be referred as Figure 2-51. Due to the low frequency of

the vibration created by SMA tensile spring and compressive biasing spring, the mixing

result wasn’t very satisfactory. According to the volume ratio of lower chamber and

upper chamber of the cuvette, the drop of the intensity after mixing didn’t reach target

point.

Figure 2-51 Mixing Result of SMA Spring Actuator

To further confirm our prediction, the solution was left over night and continued

to test after 10 hours. Even after 10 hours, the intensity level didn’t drop much. After

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manual mixing, the intensity of observation dropped to predict level (Figure 2-52). This

design can perform unplugging, but can’t reach full mixing.

Figure 2-52 Over Night and Manual Mixing Result

2.3.4.4 Conclusion

The design can provide unplugging and stable vibration range. FEM heat transfer

analysis shows that heat generation is local and controllable. Mixing test was not

satisfactory due to the slow vibration frequency. The cooling time is the bottle neck in

increasing frequency.

2.3.5 Design 5: SMA extensive spring and micro-motor actuator

2.3.5.1 Design Concept

The final design introduced a micro-motor for better mixing result. Since the

electrical lead plug is very costly, the motor shares electrical input with SMA actuator

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Figure 2-53. A resistor is added to control the distribution of current between SMA

spring and micro-motor to prevent the burning of motor by exceeding current

specifications. The biggest concern of micro-motor is if it can work under high pressure.

Different shape and size micro motor was selected and constructed and a vibration motor

commonly used for pager and cell phone was selected. This micro-motor is compact with

4 mm in diameter and 10 mm in length. Since it is widely used in cell phone industry, the

price is incredibly affordable since mass production made the cost to the minimum. A

Teflon O-ring is assembled to the micro-motor to serve as stopper.

Figure 2-53 SMA Spring Actuator with Micro-motor as a Mixer

2.3.5.2 Mixing Testing

The mixing test used the same set up as previous design. With ISS Fluorescence

Measurement, 495 nm excitation wavelength and 520 nm emission wavelength was

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selected according to CF property. The mixing result of this setup can be referred in

Figure 2-54 and Figure 2-55. Figure 2-54 shows the stabilization before and after the

mixing start point and Figure 2-55 shows the close-up view around the mixing starting

point. The result was very promising in that the intensity drop is close to the volume ratio

and the mixing time is with in 10 seconds.

Figure 2-54 Mixing result of SMA spring & micro-motor mixer

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Figure 2-55 Close-up view at mixing start point

2.3.5.3 Conclusion

This design combined tensile SMA spring and micromotor and made both

unplugging and mixing possible. Since SMA spring is only heated for 0.25 second, the

heat generation was controlled to the minimum. Micromotor functions both as a mixer

and stopper, saves space, and keeps design simple. The mixing result was very promising.

Intensity drop shows that the two solutions can be completely mixed in 10 seconds from

totally separation.

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3 Cuvette Design and Mixing Process

3.1 Cuvette Design

3.1.1 Necking Design

The Cuvette is designed for the mixing of two solutions, with its necking to fit a

plug that separatse the two solutions, the cuvette is illustrated as Figure 3-1. The design

concept is to minimize the size but to meet the measurement requirement. The biggest

size of a fluorescence spot is about 3mm in diameter and varies by different experimental

setup. Since some of the assays are very expensive, the minimum size is highly favored.

The current cuvette used by Chong for his ISS HPOC measurement is about 1ml in

volume and the size of the spot light is well centered, so we are going to keep the bottom

part the same volume as the current design. The top part of the cuvette will be decided by

the plug and the travel distance of the SMA actuator.

.60
1.00

Heating Cooling

Heating

(A) (B) (C)

Figure 3-1 The Cuvette Design

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3.1.1.1 FEM Simulation of Free Diffusion vs. Neck Opening

How long it will take the two solutions to diffuse without any agitation? A

Femlab simulation predicts that it takes almost 8000 seconds or more than 2 hours for an

8cm in height and 2cm in diameter cuvette with a three fourth reduction in necking

(Figure 3-2 and Figure 3-3). The diffusion model used water’s diffusion coefficient since

the water is the common base for many solution mix. Water’s diffusion coefficient is

around 1E-8. Material’s diffusion coefficient is proportional to its molecule size. The

bigger the diameter, the smaller the diffusion coefficient and the onger it takes to diffuse.

According to Philip Robert, it takes 30days for salt to be fully mixed in the bottom of

5cm test tube. Molecule diffusion is a long process.

Figure 3-2 A cuvette of 8cm height and 2cm diameter with 3 quarter necking

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Figure 3-3 Diffusion time of liquid

The necking design is based in diffusion analysis. Let’s examine three condition

of necking from one eighth of reduction in diameter to half reduction in diameter (Figure

3-2). Figure 3-5 shows that the different necking will influence the diffusion time. The

point plots are set in the middle of the lower chamber of the cuvette, the same spot where

the observation light will go through. It takes about a hour for half opening and three

quarter opening with different diffusion speed, but it takes almost 2 hour for a quarter

opening. The necking influence is one of the main considerations on cuvette design.

85
Figure 3-4 Necking examination

Cuvette necking vs diffustion rate


1
0.75 opening
0.5 opening
0.9 0.25 opening

0.8
Concentration

0.7

0.6

0.5

0.4
0 1000 2000 3000 4000 5000 6000 7000 8000 9000 10000
Second

Figure 3-5 Diffusion time for different necking

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To further examine the diffusion along the center line, we can find out that the

close necking area can reach the equilibrium fast and moving along the center line, the

diffusion time getting closer and closer (Figure 3-6). The diffusion curve slows down as

it moves to the bottom of the cuvette, but reaches equilibrium about the same time. The

plot is based on cuvette 8cm in height and 2cm in diameter with 3 quarter necking or one

quarter opening.

Cuvette necking vs diffustion rate line plot


1

0.95

0.9

0.85

0.8
Concentration

0.75

0.7

0.65

0.6

0.55

0.5
0 1000 2000 3000 4000 5000 6000 7000 8000 9000 10000
Second

Figure 3-6 Point plots along the center line from necking to bottom of the cuvette

3.1.1.2 Final Cuvette Design

From the simulation above, the bigger the neck opening, the faster the mixing

process takes. Many variations of necking were designed, fabricated and tested. The final

design eliminates necking with “step down” necking design. This necking design will

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maximize the diffusion process. The lower chamber of the cuvette is kept for 1 ml for

observation requirement while the upper chamber is designed according to pulling micro-

motor distance and SMA extension range.

Figure 3-7 Final Design of Cuvette

3.2 Mixing under High Pessure

To understand the mixing process, we need to start from diffusion. When you add

cream or sugar to your coffee, you need a stirrer, a plastic or a wooden stick from

Starbucks or a tiny golden or silver spoon from your granny’s. Without a stick or a spoon,

it takes forever to taste the sweetness or to get rid of the bitterness in coffee. A simple

stir will accelerate the diffusion process dramatically. But how does this happen? What

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make the forever waiting to a few seconds? It seems so simple that nobody will question

it, but answers might not be that easy to reach.

If you work in a chemical or bio lab, many different kinds of stirrer might be

familiar. But to mix some solution under high pressure might only be seen in a science

fiction movie or James Bonds’ adventure. I remember that a big piece of metal block was

enclosed in the glass bell. It is rotating with a suspense background soundtrack. It will

explode at some point and a hero is going to save the scene, but you never know how.

The difficulty of mixing under pressure is space limitation. High pressure will

also increase the viscosity leading to an increased in back force. The choice of proper

methodology is the key component with respect to different application. The main mixing

method under pressure is as followings:

Mixing methods under pressure are mechanical agitators, rotating and rocking

apparatus and electromagnetic stirrers. Mechanical agitators use motors as an agitator.

One of the oldest and least-preferred methods of mixing is to agitate with motors located

outside of apparatus. By introducing a stirrer into high pressure apparatus with a shaft

passing through a gland, the motor drives the stirrer. Although improvements have been

made in the design of glands and friction methods, they now can be reduced without the

breaking the hermetic seal, its use allows friction to be reduced only at low surface

rubbing speeds. (Flom and Porill 1955) Another mechanical agitator is stirrers with

motors located inside apparatus. When the internal dimensions of the apparatus is large

enough, the stirrer and motor can be placed inside the apparatus, introducing the motor

power current through electrical leads. (Kiebler 1945) This requires that the motor

winding can withstand the action of the medium inside the apparatus. A deficiency of this

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design is the possibility of contamination of the medium being mixed by the lubricant of

the motor, as well as the necessity of cooling the motor, which can be easily overheat

(Figure 3-8).

Figure 3-8 Location of stirrer with motor within apparatus: 1) motor winding 2)

rotor 3) cooling coil 4) electrical leads 5) packing

Rotating and rocking apparatus is suitable for apparatus not too cumbersome and

if the general location of the apparatus does not present a problem, mixing maybe

performed by rotating the high-pressure vessel around an inclined axis or by rocking it

about its vertical axis (Payne, Sreed et al. 1958).

Electromagnetic Stirrers are widely used for agitation of the contents of vessels

operating at pressure of a few thousand atmospheres. This design has generally a few

problems: overheating by parasitic currents; dimension of coil, need to be small;

reliability of insulation of the wire, etc(Bruns and Braude 1953). An electromagnetic

stirrer power supply of powerful solenoid has interrupter, magnetic starter, transformer,

autotransformer and solenoid (Figure 3-9). A drop stirrer is attached to the head 2 of the

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apparatus, made of nonmagnetic steel (Figure 3-10). When the current is passed through

the coil, the core returns symmetrically relative to the coil, raising the stirrer. When the

current is turned off, the stirrer drops by its own weight to the bottom of the apparatus.

Thus, agitation is achieved by turning the current on and off. This type of agitation

requires a considerable amount of time, but gives good results, especially if the amplitude

of the stirrer oscillations is rather great. The intensity of the mixing action depends also

on the frequency with which the stirrer is dropped and the form of its blades. A rotating

electromagnetic stirrer is another option of this kind of stirrer. The reciprocating motion

of a stirrer can be combined with rotation. As show in Figure 3-11, core 1 is joined to a

stirrer shaft 2 with finger 3 which moves in a spiral cut in tube 4, fastened firmly into the

apparatus. As the core is draw upward, the motion of the axle will include rotation, due to

finger 3.

Figure 3-9 Power supply of powerful solenoid: 1) interrupter 2) magnetic start 3)

transformer 4) autotransformer 5) solenoid

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Figure 3-10 Drop stirrer: 1) coil 2) head 3) core 4) shaft 5) body of apparatus

Figure 3-11 Stirrer with reversing rotation:1) core 2) axle 3) finger 4) tube

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In the above mixing processes, there are two motions involved: up and down, and

rotation. When drinking a cup of tea made from a tea bag, one will pull the string of the

tea bag up and down and the tea can be dissolved immediately. For coffee with sugar and

milk added, a stir stick will do the trick.

3.3 Turbulent flow

Although the theory of homogeneous isotropic turbulence may have great

mathematical elegance, it is an idealized subject and often not very useful in handling

practical problems involving turbulence in inhomogeneous systems. One of the

consequences of inhomogeneity in a fluid system is that it gives rise to transport

phenomna. Since fluid elements carrying different physical quantities move around

randomly under turbulence, we expect turbulence to enhance the transport processes. The

theory of turbulent diffusion was first formulated by Taylor (1921). Even if you have

never heard of Taylor you make use of the motion of turbulent diffusion when you stir

coffee to mix sugar in it. If one puts sugar in the coffee and does not stir it, then the sugar

mixes with the coffee only by molecular diffusion, which takes a very long time. On the

other hand, if the coffee is stirred, then generates turbulence inside it, turbulent diffusion

mixes up the sugar much more efficiently than molecular diffusion. (Graham).

Turbulence is the main factor to accelerate the mixing process. Despite many

years of intensive research into turbulent diffusion, however, our ability to predict means

contaminant distributions is often quite crude and our attempt to predict statistical

variations of concentration fluctuations even cruder. Turbulence diffusion is very

efficient in rapidly decreasing the concentrations of contaminants that are released into

natural environment (Roberts and Webster). Most mathematical models of turbulent

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diffusion, particularly engineering models, predict only time-averaged concentrations.

Turbulence is difficult to define exactly; nevertheless, there are several important

characteristics that all turbulent flows possess. These characteristics include

unpredictability, rapid diffusivity, high levels of fluctuating vorticity, and dissipation of

kinetic energy. Since the nature of turbulence is unpredictable, statistical measures is the

best method to describe the motion. The velocity is described as a time-averaged value

plus some fluctuation. The average quantities can be denoted as

(3-1)

Where T is a time much longer than the longest turbulent fluctuations in the flow.

A time record such as shown in Figure 3-12 is called statistically stationary if the mean

quantities remain constant over the period of interest.

Figure 3-12 Sample turbulent velocity record

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3.4 Mechanism of mixing in turbulent flows

Turbulent flows contain irregular motions over a wide range of length and time

scales. How do these motions contribute to mixing? Mass transport due to molecular

diffusion can be described by Fick’s Law. In a flowing fluid, the flow itself contributes to

the transport mechanism. The mixing process is therefore due to distortion, stretching,

and convolution of the original patch whereby the original volume is distributed

irregularly over a larger volume, so that the concentration, averaged over some finite

volume, decreases. In the absence of molecular diffusion, however, such a process would

not reduce actual peak concentrations at a point; the reduction of these peaks is therefore

very dependent on molecular diffusion. G. I. Taylor (Taylor 1921) published one of the

most important results in turbulent diffusion theory, which provides a link between the

eddy diffusion coefficients and turbulent flow properties, in 1921. Since turbulent

velocity fluctuations are irregular, the results of each trial differ. On average, however,

the particles wander apart from each other, and the rate at which they wander apart can be

related to a diffusion coefficient. This is easier to imagine by considering individual

particles, released from the coordinate origin at different times. Many scientists and

engineers tried to understand how turbulent flow accelerates mixing processes (McComb

1974; Kesten and Papanicolaou 1979; Eijnden 1998; Tabak 2004).

The expressions for the molecular transport coefficient from kinetic theory have

different values. For example, compare expressions for thermal conductivity and

viscosity. The coefficients for different turbulent transport processes, however, are

believed to be the same. The cause for this contrast between molecular and turbulent

transport is that molecular motions are related to the quantities they transport. Molecules

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carrying more thermal energy move faster. This is not true for a turbulent fluid. The

temperature of a blob of fluid does not have a direct functional relation with the turbulent

velocity of the blob. Hence all the diffusion coefficients for the passive transport of any

quantity in a turbulent fluid have the approximate value as following equation, the

turbulent coefficient is proportional to the average of velocity square.

1
DT = v 2τ cot (3-2)
3

This equation best explains micromotor mixes much better than SMA spring

vibrator, since micromotor can produce a velocity change from 0 rpm to 2500 rpm in

seconds but the SMA spring vibrator can only produce 0.5 Hz to 1 Hz due to cooling

limitation. The equation also shows that velocity fluctuation contributes to accelerate

mixing process, once the flow is fully developed, the velocity won’t help anymore.

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4 High Pressure Optical Cell Design

4.1 Weber and Paladini’s High Pressure Bomb

In order to measuring fluorescence depolarization of protein solution, Weber and

Paladini designed and constructed a high-pressure bomb to meet the experimental

requirements. The bomb can withstand pressures up to 2 kbars, it has four optical ports

and one hydrostatic port to permit its pressurization (Paladini 1980). The cell allow for

easy loading and unloading samples and prevent them from making contact with either

the inner metallic bomb walls or the pressure transmitting fluid. The inner chamber was

kept as small as possible to increase the efficiency of the pressure-delivery pump which

has a total capacity of 11 ml.

Figure 4-1 Weber and Paladini’s Pressure Bomb

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4.2 ISS’s current HPOC

ISS continued Weber and Paladini’s design concept and improved finishes. The

ISS HP Cell is designed for high pressure absorption and fluorescence experiments, as

well as for Raman spectroscopy studies. The cell can be pressurized up to 3,000 bar

(43,500 psi) when using quartz windows and up to 40,000 bar (58,000 psi) when using

sapphire windows. Three 10mm diameter windows are placed in the optical plane; the

10mm clear aperture allows spectroscopy studies using either a laser or a lamp as light

excitation sources.

Figure 4-2 ISS HP Cell

The cell is made of a stainless steel alloy, which has excellent thermal

conductivity. To control temperature, an external temperature bath can be added. The

temperature of the cell can be monitored by a thermocouple inserted in the side of the cell.

Ethanol is the medium to pass hydraulic pressure from the pump to the cell. The ethanol

98
is required with low fluorescence background. A hand pump generates hydrostatic

pressure. The finish of ISS HP Cell is more polished than the Paladini and Weber’s

prototype. The cell shape is rather cubic than cylinder shape, this makes alignment of

light path much easier.

4.3 HPOC Design:

Our design is based on ISS HPOC. Since the SMA spring is heated up by

applying electrical current, an electrical plug assembly is intruded to the current HPOC.

The two electrical plugs increase the height of the HPOC and require a new pressure

analysis. The high pressure optical cell is made from the cell body, observation windows,

electrical lead plug and pressure inlet plug. The cuvette assembly containing the SMA-

based actuator will be placed inside the inner channel of the cell body. The cell body is

made of Vascomax C-300 which has favorable strength and toughness characteristics and

is commonly accepted as a high pressure cell material. Although the ideal shape of a

pressure cell is sphere (Bobrowsky 1968), a cylindrical shape provides the best

compromise between strength and manufacturability (refer to Figure 1-1).

4.3.1 HPOC Body Design

Stresses are present in a material under load, causing deformation. The design of

the apparatus operating at low pressures entails primarily determining the wall thickness

sufficient for the apparatus to withstand the load to be applied without permanent

deformation. For equipment working at very high pressures, plastic deformation is

included in calculation of wall thicknesses. The occurrence of the type of deformation is

utilized in construction. When the stresses in the metal exceed the yield strength, the

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metal goes over into the plastic state. The internal layer of the wall of the cylinder, where

the pressure exceeds the yield strength, will be in the plastic state, and the equivalent

stresses in it will have a certain value, uniform throughout the entire plastic layer. With a

further increase in pressure, the stress in this layer will be not increased, but the thickness

of the plastic layer will increase. This agrees with the concepts of the nature of plastic

deformation presented above. The high-pressure cylinder is thus, as it were, divided into

two concentric layers; the internal (plastic) layer, and the external (elastic) layer which

protects the plastic layer increases, and the thickness of the elastic layer consequently

decreases until rupture finally occurs. Experiments have shown that rupture of a high-

pressure cylinder begins from the outside.

4.3.2 Electrical Lead Design

To select the right electrical leads is important to the HPOC design. First, the

electrical lead plug must hold 3 kbar of pressure; second, the cost is another issue to

consider; third, easy manufacturability is most desired.

4.3.2.1 Low-pressure Electrical Leads

It is comparatively easy to seal an insulated electrical lead for pressures up to

1000atm using various insulating materials. Sealing wax or other cement, melted onto the

rod to be insulated can be used for this purpose. Good results can be obtained using fibra

as an insulator. The electrical lead design is constructed on principles of ordinary seals.

The insulated lead is compressed by the sealing material, and friction prevents the lead

from being expelled. If the conductor’s diameter is greater than 0.5mm (such as when

100
high current must be passed) and the pressure in the apparatus is over 1000atm, this

method is unsuitable.

4.3.2.2 Unsupported-Area Electrical Leads

A more reliable seal for an electrical lead is the inverted cone. A metal cone with

a conductor, surrounded by insulating material is placed in a carefully fitted seat in the

body. The pressure acts on the large base of the cone and forces it into the seat. The

insulator used may be any one of numerous materials such as ivory, plastic, pyrophyllite,

mica, etc. the selection of the material to be used depends on the pressure and

temperature in the apparatus. Ivory, can be used up to 3000atm while methyl

methacrylate can operate without failure up to pressures of 18,000atm. Mica winding of

the cone is a simple and reliable method allowing the use of an electrical lead at

temperatures up to 800-900˚C and has been successfully used for many years at pressures

of 18,000-20,000atm.

An Electrical Plug Assembly (Figure 4-3) is used to provide the SMA wire with

electricity power under high pressure. A heat shrinkable plastic tubing (Terry 1972) has

been proved to be effective in both gas and liquid systems, and it is capable of

withstanding pressure up to 10kbar (or 1000Mpa). A Kynar tubing is shrunk over a 16°

cone using a heat gun. This assembly is inserted in a cone drilled in the pressure cell,

with the large side of the cone facing the pressure side. This electrical lead design will be

used in conjunction with a Bridgeman (Bridgeman 1931) type plug body.

101
Figure 4-3 Exploded assembly view and assembly electrical lead

4.3.2.3 Multi-Conductor Electrical Lead

A design for brining several continuous conductors out of an apparatus was

suggested by Ya. A. Kalashnikov (Kalashnikov and Zh. 1954). Body, made of 10Kh steel

contains two holes whose length depends on the diameter of the wire to be brought out of

the apparatus and pressure to be contained. In particular, for a pressure of 5000atm and a

conductor diameter of 0.5mm, the channel length should be at least 18mm (Figure 4-4).

This method is not widely used due to its complex process.

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Figure 4-4 Kalashnikov electrical lead: 1) body 2) holes 3) conductors 4) thread

After passing wires 3 through the holes, body 1 is filled with a styrene monomer,

then it is poly-merized in a special bomb for two days at 1500atm and 80°C. As a result

of this, the bond between the leads, polystyrene, and body wall becomes so strong that

the pressure cannot force the conductor out of the plastic or the plastic out of the body.

This force out is hindered also by threaded diameter 4 in which the styrene is

polymerized. The excess polymerized styrene is cut out on a lathe. After this, the

electrical lead is ready to use. This method’s advantage is multi-electrical lead at the same

time, but the process is very complicated.

L. F. Vereshchagin, L. F. Kuznetsove and T. A. Alaev have used an electrical lead

of a somewhat different design, also made in the form of an inverted cone (Vereshchagin,

Kuznetsov et al. 1956). The Inverted cone is the most convenient and popular used

electrical lead method. In our design, we used the inverted cone method.

103
4.3.3 Observation Window Design

The lower window plug assemblies are used for collecting the spectroscopy data.

The port assembly is designed to hold a fused silica or quartz window through which the

light passes. These assemblies are made removable to accommodate window-sealing

surface. A threaded brass cap that provides only light mechanical support for the window

is installed at the end of the assembly. The cap has a 6.4 mm diameter hole to allow the

transmitting fluid to enter and press upon the exposed window surface. The window itself

is made of Corning 7940 fused silica that can withstand the required pressures as well as

pass the UV-visible wavelengths. The window arrangement used in this assembly is

commonly referred to as a Poulter-type window support. The sealing of this type of

support is based on the Bridgeman (Bridgeman 1931) unsupported area principle. In brief,

sealing is accomplished because the viewing aperture creates a surface area imbalance

between the fluid side and the support side of the window, resulting in a pressure

differential, which ensures a strong seal. This sealing method, however, is heavily

dependent upon the accurate machining of an optically flat surface on both the window

and window support and is not reliable for very small apertures. The methods for

checking the surface quality of windows and supports have been reported by Sherman

and Stadtmuller (Sherman and Stadtmuller 1987) (Figure 4-5).

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Figure 4-5 Exploded view and assembly a window plug

Windows for visual observation at high pressures are made of glass, transparent

quartz, sapphire, diamond, and even cooking salt. Glass windows are extremely sensitive

to the action of water. It is known that water is dissolved into the surface layer of glass,

penetrating to depths of up to 200 molecular layers. At high pressure, the solution is

increased and water penetrates even more deeply into the glass. The simplest viewing

window design consists of a cone made of glass or transparent quartz, lapped to the metal

seal. Sometimes, packing is placed between the glass and metal. The large end of the

cone is place toward the inside of the apparatus, so that the pressure forces the cone into

the seat. The reliability of this type of construction depends on the care with which the

lapping is done and the quality of the glass or quartz cone. Even very well-made viewing

windows of the type are burst due to chipping of the edge of the cone. These windows are

usually used at pressure on the order of 1000-1500atm, although in individual cases

quartz cones have withstood pressure up to 10,000atm. A considerably more reliable

viewing window can be constructed on the unsupported-area principle. A glass cylinder

with optically parallel ends is pressed against the polished surface of the nipple. The cap

will retain the glasses. The nipple is screwed into the body of the apparatus and packed

using packing. The pressure created in the apparatus acts on the area. The central hole is

105
drilled into the stock for the nipple and is enlarged with a reamer. The part is place

between the centers of a lathe and turned, after which, while mounted in the chuck, the

thread and final turning are performed in one pass. The surfaces of the parts which are to

be polished are hardened. The end of the nipple is heated until dark red, then tempered

for 2-3 minutes in warm (30-35˚C) water and cooled in oil. After polishing, the viewing

window with its optically flat surface held against the dry surface of metal should show 2

to 3 Newton rings.

4.3.4 Pressure Inlet Plug Design

The inlet plug seal is achieved by using a combination of three sealing rings. One

lead ring, which is sandwiched between two brass rings, deforms to fill any voids created

by wear or poor surface quality. The type of sealing method has been used successfully in

other hydrostatic pressure cells. The pressure inlet plug uses a similar external

configuration and three-ring sealing method as used on the four window assemblies in the

lower chamber.

4.3.5 Temperature Monitoring under High Pressure Design

The methods of measuring temperature in high-pressure apparatus can be divided

into two primary classes: 1) measurement of temperature of the thermostated liquid in

which the high-pressure apparatus is submerged, or measurement of temperature of the

walls of an apparatus heated by the furnace; 2) measurement of temperature within the

high-pressure apparatus.

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4.3.6 Machine Finishes

The parts of high-pressure apparatus must be finished very carefully. The

finishing of areas directly in contact with the medium creating the pressure, as well as

finishing of threads, packing surfaces, etc., must be especially accurate. Burrs and

grooves cannot be tolerated. Sharp angles should be smoothed. Special requirements are

placed on threads. Threads in high-pressure apparatus are usually made short, i.e. with

smoothes sharp angles, with prevents “seizing”. Special attention must be turned to the

centering of threads in holes that are sealed by rods, pins, or shafts. Methods of

mechanical working of parts are described in the corresponding sections.

4.4 FEM Analysis of HPOC

FEM software ANSYS is used for cell body assembly stress and deformation

analysis. Since the cell body assembly is symmetric we only take half for analysis to

save computer time. The pressure is 3000 bar in the chamber and 1 bar outside. As

showed in Figure 4-6, the maximum stress is 8893psi, which converted to metric is

613bars around the window assembly. The yield strength of the Vascomax C-300 is 1915

MPa, which is three times larger than that of the cell’s maximum stress.

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Figure 4-6 Stress analysis for cell assembly by Ansys

To verify ANSYS simulation results, FEM software FEMLAB is used for stress and

deformation analysis of cell body assembly. Since the cell body assembly is axial

symmetric we only take half for analysis to save computer time. Pressure is 3000 bars in

the chamber and 1 bar outside. As showed in Figure 4-7, the maximum stress (824.5 MPa)

and deformation (6.14E-5m) are in the radial direction around the window assembly. The

yield strength of the Vascomax C-300 is 1915 MPa and the safety factor is around 2.35.

The cell design is reasonable and safe.

108
Figure 4-7 First principle stress of cell assembly by Femlab

Difference FEA method showed different results, this bring up the uncertainty of

FEA methods. A thoroughly FEA study can be performed using TRUEGrid and ANSYS

together. Both the safety factors weren’t very high, around 2 to 3. But the highest stress is

below its yield strength which means no plastic deformation will occur. Refer to previous

discussion of high pressure cell design, for very high pressure condition, plastic

deformation will occur in inner layer, the stress in the layer won’t increase according to the

pressure increase, which will lead to plastic deformation layer increase. Our design is under

that stage, a plastic deformation won’t occur for pressure up to 3500bars. The cell thickness

is determined from numerous previous designs. The height increase didn’t introduce higher

stress concentration.

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4.5 The Final HPOC Design

The final HPOC design involves height change to existing design, and also

introduces two new openings to accommodate Electrical Lead Plug design (Figure 4-8).

The height increase is a potential risk of increasing hoop deformation. And new plugs

will lead to an increase in the cost.

Figure 4-8 Illustration of final HPOC design

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4.6 Our New HPOC Fabrication collaborated with ISS

The ISS is a company that specializes in Fluorescence Measurement. They also

manufacture commercial HPOC based on Weber and Paladini’s Original High Pressure

Bomb design. We would like to collaborate with them to have technology transfer our

new designed HPOC to become a product. After receiving our new HPOC design, ISS

proposed some modifications to reduce cost and fit to existing set-up. Basically, the

modified design kept all features of their current HPOC body, but modified the inlet plug

by introducing electrical leads into the chamber. The design eliminated two electrical

lead plugs which are the most costly. Besides the inductions of electrical lead, the top

plug still serves as an inlet plug. With the electrical leads taking the top side of the plug,

and the opening for pressure inlet is set on the side (Figure 4-9 c). To accommodate the

increase of the height of new cuvette, the plug was hollowed in the bottom side. Two

pieces of 3/16 in diameter rod was cut to serve as electrical leads. Heat shrink tubing are

approximately the same length as the brass electrical leads. Using a standard heat gun, the

tubes are shrunk onto the Brass Electrical leads. Then the leads are inserted into the

threaded electrical lead holders as shown in Figure 4-9 a) and the heat shrunk tubes are

trimmed to expose brass electrical lead rods. Using the supplied wrench, we inserted the

leads and holders into the Top Plug as shown, finger tight.

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Figure 4-9 Electrical lead and inlet plug combo

The new ISS design also modified the fixture of the cuvette. The our design and

ISS current HPOC, a fixture was used to hold cuvette in position at the bottom of the

HPOC inner chamber. The fixture positions the cuvette and aligns the observation path

through the same spot of the cuvette on each measurement. Since the electrical lead is on

the top, which will turn as the plug is screwed to tighten up to seal the HPOC, electrical

lead will be tangled for 9 turns if the cuvette stays in current fixture. ISS designed a

fixture which connected to the top plug (Figure 4-10 (a), (b)). The fixture is also served

as cuvette cap (Figure 4-10 (c)). There are two plugs that serve to seal the cuvette: red

silicone plug seals the cuvette fixture cap and pink vinyl plug seals the bottom cuvette.

112
Figure 4-10 Cuvette and its cap

The ISS design has many advantages: budget saving, no tangle of wires, compact,

etc. But it also has some shortcomings compare to our own design. After a few

experiments, the O-rings used to guide both the cuvette and cuvette cap become frictional

which makes mounting and dismounting cuvette to cuvette cap and cuvette cap to inlet

and electrical lead plug combo very difficult. The friction increases because the O-rings

are soaked in ethanol which is the media of transferring the pressure into the cell. The

mounting of cuvette from the top causes problem on the positioning of the cuvette to the

light path. The tilt of the cuvette causes the initial measurement of value to be different

from each measurement. This can be improved upon by some modification on the Cell.

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114
5 Complete System Assembly and Final Testing

5.1 The Final Complete System

The final complete HPOC assembly is shown as Figure 5-1. The new design

HPOC has 3 observation windows, the third window didn’t show in the figure is

perpendicular to the other two windows and at the same level. The excitation light will

pass through the third window and hit on the center of the lower part of the cuvette

chamber. As the assay is excited by certain wavelength light, the emission light is

collected from the right side window and the left side window is used for comparison

purpose mostly. The top plug has two functions: the yellow lines indicate electrical leads

which pass the current to both SMA spring and micro-motor; an opening on the side of

plug passes hydraulic pressure into the chamber of HPOC. Inside of HPOC chamber, a

dual chamber cuvette resides in the cuvette holder connected it to the top plug. Through

the cuvette holder, electrical lead is connected to SMA spring and micro-motor via a

socket. The micro-motor has a Teflon O-ring which functions as a stopper to separate two

different solutions before the pressure reaches desired level. A flexible polyethylene tube

made fan is attached to the shaft of the micro-motor which starts turning once the plug is

opened by SMA spring. A red soft plug seals the cuvette cap from the top to pass

pressure from inner HPOC chamber into the cuvette, so does the pink soft plug in the

bottom of the cuvette.

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Figure 5-1 Final Complete HPOC Assembly

The HPOC assembly is cooled by a thermostatic bath cooling system (refer to

Figure 5-2). Also from the figure shows the hydraulic pressure system. High grade

ethanol is required for eliminating any unwanted fluorescence interference from the

media. Before adding pressure to the HPOC, the manual pressure pump needs to fill with

ethanol. By cutting off the valve to the HPOC, opening the incoming valve and turning

back the screw of manual pump, the whole pressure control system is filled with ethanol.

After closing the incoming valve and opening the valve to the HPOC, turn the manual

screw forward and let ethanol start to flow out from the tip of the tube and this shows the

pressure system is filled with ethanol. Then connect the tip to top plug of the HPOC, the

pumping process can be started. While turning the wheel of manual pressure pump, the

pressure can be monitored from pressure gauge. During the pressurization process,

leakage from all valves and connections need to be checked since the leakage can

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produce pressure drop which can be monitored from pressure gauge. Once the pressure is

stabilized, the fluorescence measurement can be started.

Figure 5-2 Pressure control and cooling system

To measure the diffusion process, ISS K2 fluorescence measurement is used as

Figure 5-3. ISS K2 is widely used for steady-state fluorescence measurement. Its optical

design and automatic instrument control are “state-of the-art” and time-resolved

fluorescence measurements. Full automation and the user-friendly Vinci software

package make the instrument easy to use. The measurement has a light source, light path

through which desired wavelength selection and modulation are made, enclosed HPOC

chamber, light collection and data process system.

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Figure 5-3 ISS K2 Fluorescence Measurement

Figure 5-4 shows its optical diagram. Fluorescent light is generated from a Lamp

and a narrow band of wavelength of fluorescent light is selected through excitation

monochromator. The light is modulated by a light modulator before hitting on sample.

Fluorescent light is modulated to 490nm wavelength to excite Carboxyfluorescein (CF),

CF will emit 520nm wavelength of fluorescent light. The receptor will measure the

intensity of the light, around 520nm wavelength. The wavelength of emission is always

bigger than that of excitation. Without any CF in the cuvette, the intensity is around 1000,

but when 300nM CF solution is measured, the intensity is around 300,000. The

measurement number is relative, it can be adjusted. The upper limit is above 1,000,000.

To receive the most sensitivity of comparison, the higher the intensity, the more desired.

But high intensity can cause photo bleaching which cause the intensity to drop gradually.

Our measurement is set around 300,000 which will be a good resolution and minimum

photo bleaching.

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Figure 5-4 Optical diagram of ISS K2

5.2 Leakage Testing of the Stopper

The plug design has two considerations: to seal the two solutions well before

activation; and keep the size compact. Since the diffusion process takes a long time for

free diffusion, once the two solutions are separated, the leakage won’t influence much.

To keep the plug compact reduce the travel distance of the plug which will reduce the

upper part of the cuvette as well. The leakage testing is set up with ISS fluorescence

measurement. Carboxyfluorescein (CF) is used for this testing. CF is diluted to 150nM

and filled at the bottom part of the cuvette and water is filled in the top part. The

measurement result is show as Figure 5-5. Raw data is normalized and fitted with

polynomial method. The measurement put water solution in the lower cuvette chamber

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and CF solution in the upper chamber. There is more fluctuation on the measurement

since the value is very small, around 1000 intensity. But if leakage happens, the CF will

increase the intensity dramatically. There is no leaking found in 1800s or 30mins. We

keep the measurement running for a few hours, no leakage found either.

Leakage Testing

0.8 raw data


fitted
Intensity Normalized

0.6

0.4

0.2

0
0 200 400 600 800 1000 1200 1400 1600 1800
Time (s)

Figure 5-5 Leakage ISS CF testing

5.3 Mixing Testing

The diffusion test was set up with water in the bottom of the cuvette and 300nM

CF on the top. A plug is sealed in the middle. After stabilize, the plug is pushed down,

the diffusion process starts. Figure 5-6 shows the diffusion results. The results show that

it takes more than 3 hours to reach the equilibrium point. Since CF molecule is much

larger than water molecule, its diffusion coefficient is much smaller since diffusion

120
coefficient is reverse to its size. The simulation of water diffusion shows that it takes

around 2 hours to reach equilibrium. The CF diffusion takes much more time than water.

In the experiment, the push down of the plug gave a turbulence push and the process is

more unpredictable. Turbulence diffusion will be discussed in the agitation mixing

chapter.

5-1

Where DO2-H2O denotes diffusion coefficient of O2 in H2O, µ denotes viscosity,

VO2 denotes molecule volume of O2. From the equation, we can see that diffusion

coefficient is proportional to the inverse of viscosity and molecule volume.

Diffusion Testing
800

700

600

500
Intensity

400 raw data


fitted
300

200

100

0
0 2000 4000 6000 8000 10000 12000 14000
Time(s)

Figure 5-6 Diffusion ISS CF testing

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.

5
x 10 Mixing with SMA and biasing spring actuator agitation
5
data 1
4.5 6th degree

3.5

3
Intensity

2.5

1.5

0.5

0
0 1000 2000 3000 4000 5000 6000 7000
time(s)

Figure 5-7 Mixing with SMA and biasing spring actuator agitation

After introducing micro-motor, the mixing process is improved dramatically. The

experiment designed to start the measurement from high intensity to low intensity. By

putting 300 nM CF in lower chamber of the cuvette, the starting intensity is set around

300,000. With known volume ration of CF solution in lower chamber to water in higher

chamber, the final intensity of well mixed solution can be predicted. To test the dilution

and intensity relation, we did some dilution test. By doubling the volume with water each

time, the intensity from 300 nM and stopped at 37.5 nM is measurement. The result

almost follows the linear trend (Figure 5-8). Using this result, with known volume ratio,

the target intensity can be estimated.

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Figure 5-8 Intensity vs nM

First, the mixing fluorescent measurement under 1 atm pressure is recorded and

the result is shown in Figure 5-9, Figure 5-10, and Figure 5-11. The target was set to 10

second with 90% drop in intensity. All 3 cases reach the target. The mixing ratios

according to Figure 5-8 are from 90% mixing to 100%.

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Figure 5-9 Mixing result under 1atm first measurement

Figure 5-10 Mixing result under 1atm second measurement

124
Figure 5-11 Mixing result under 1atm third measurement

When the pressure is raised to 300bars, about 300 times atmosphere, the mixing

result is as Figure 5-12, Figure 5-13 and Figure 5-14. The mixing trends varied a little

from each other, 90% drop all achieved in three cases, but all reached the equilibrium

around 25 seconds. The trends difference is the turbulence diffusion’s unpredictable

feature. The diffusion coefficient is related to the flow velocity, but only can be described

by average velocity. Instant relationship is not well established, and remains

unpredictable. This experiment proved that SMA spring with micro-motor actuator can

perform unplugging and mixing work under pressure. More experiments are undergoing

for higher pressure such as 1000bar or 3000bar. Higher pressure experiments encounter

some unstable problem, the cause and solution are under investigation.

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Figure 5-12 Mixing result under 300bars, first measurement

Figure 5-13 Mixing result under 300bars, second measurement

126
Figure 5-14 Mixing result under 300bars, third measurement

5.4 Conclusion

The complete system was assembled, tested and ready for bio-physical study. The

new designed HPOC can hold up to 3 kbar with easy assembling, loading and unloading.

The connection between HPOC to cuvette achieved full function of positioning,

providing electrical supply. A dual chamber cuvette can separate two solutions before the

pressure was added up to desired level. SMA spring and micromotor achieved

unplugging and rapid mixing under pressure.

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6 Bio-Safety test of Heat Generation of SMA

6.1 Bio-safety testing of SMA wire

The compatibility of the Ti-Ni shape memory alloy (SMA) with biological

molecules such as phospholipids and proteins have been tested. After the SMA Flexinol

wires were immersed to a solution of liposome’s composed of 1-palmitoyl-2-oleoyl-

phosphatidylcholine (POPC) and dehydroergosterol for 30 minutes at room temperature,

the concentration of phospholipids remained in the bulk solution was determined using

the Bartlett method [15]. To our surprise, there was a small increase in phospholipids

concentration with increasing the surface area of Flexinol wires (Table 2). At present, the

origin of the increase in [POPC] is unclear. It is unlikely due to wire contamination. The

wires were thoroughly rinsed with light detergent and deionized water prior to use, and

no phosphate-containing buffer or reagent was used. Nevertheless, we demonstrate here

that we can make corrections for this unexpected increase in phospholipid concentration

once the surface area of the SMA wire is known.

Table 2 The change of phospholipid concentration with SMA wire surface area

Wire surface Average Standard


area, square [POPC], mM deviation of
inches [POPC], mM
0 0.0826 0.0006
3.77 x 10-2 0.0872 0.0005
6.29 x 10-2 0.0895 0.0006
1.13 x 10-1 0.0918 0.0005

There are no significant changes in the concentration of the protein samples after

the addition of the shape memory alloy Flexinol wires. Three proteins, namely, human

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albumin, bovine serum albumin, and bovine gamma-globulin, were tested. The protein

concentration was determined using the Lowry (Lowry, Rosbrough et al. 1951)method

with bovine serum albumin as the standard.

6.2 Bio-Safety test on SMA heat generation

The SMA spring is actuated by applying electrical current, electrical energy

generates heat. The heat generation can damage protein that’s around when temperature

reach too high. The experiment of heat study is conducted with interplay between

membrane cholesterol lateral organization and the activity of membrane surface-acting

enzymes. The effect of the membrane cholesterol mole fraction on the initial rate of

cholesterol oxidation catalyzed by COD was investigated at 37°C using cholesterol/1-

palminoyl-2oleoyl-L-α-phosphatidylcholine (POPC) large unilamellar vesicles. The

experiment is conducted in 3 conditions: 5A in 2 seconds; 5A in 5 second and 5A in 10

seconds. 2 seconds is for SMA spring to unplug the stopper. And 5A is the effective

current to heat SMA up in a bathed controlled HPOC to body temperature. After

unplugging, the current can be dropped to 2A which heating effect is dramatically

reduced due to heat transfer is very efficient in a bathed controlled cell. From Table 3, it

shows that the 5A applied for 2 seconds reduce the dropping time factor by 19% and 5A

applied for 5 seconds reduce the dropping time factor by 25%. From the testing result,

COD enzyme is very sensitive to temperature increase caused by electrical current

heating. To control the temperature minimum less than 40°C is very critical to many

membrane proteins. Referring to Figure 2-37, heat can be controlled to a minimum, less

than 2 seconds. By applying current quarter second periodically and giving the cooling

time for a second, it can minimize the heating effect and provides desired force. For 5A

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10 seconds testing, the result wasn’t consistent at 2 seconds and 5 seconds condition, the

influence was less. But when the solution was examined, residue was found in enzyme

solution. Residue can be the influence on the curve.

Table 3 Bio-safety testing according current and time


A Tau Y0 S^2 Tau ave %
5A 2sec 1 0.53572 405.515 0.42317 0.0216
2 0.5144 399.345 0.44761 0.02303
3 0.53552 401.526 0.44914 0.02043
4 0.54826 402.35 0.42632 0.02112 402.184 19.40%

5A 5sec 5 0.52503 383.539 0.45777 0.0245


6 0.53503 374.35 0.43773 0.0269
7 0.50894 369.154 0.45475 0.02273
8 0.51054 369.085 0.46752 0.02419 374.032 25.04%

5A 10sec 9 0.52397 462.3 0.45332 0.02839


10 0.53516 480.261 0.4375 0.02606
11 0.55631 437.369 0.43856 0.03121
12 0.56295 431.947 0.41343 0.02963 452.969 9.22%

5A 10sec 13 0.48766 481.116 0.4632 0.02963


repeat 14 0.48324 439.213 0.48993 0.0386
15 0.4802 414.542 0.48694 0.02864
444.957 10.83%

Control 16 0.59914 530.898 0.37337 0.04541


17 0.57204 505.74 0.39571 0.0438
18 0.60774 483.255 0.38146 0.04748
19 0.57506 476.077 0.40264 0.02415 498.993 0.00%

6.3 Avoiding Photo bleaching effect

Photo bleaching was a big issue on fluorescent measurement of bio-solution, since

the light emission intensity of most protein, lipid, etc was low, photo bleaching misleads

130
the result of measurement. When bio-safety experiment was conducted, photo bleaching

effect was encountered. The Experiment Design was using 1.8ml Buffer (Water); 10µl

DHE; 10µl Enzyme (Cox). The ISS setup: Excitation Wavelength: 325nm; Right

Emission Wavelength 396nm. Steady state is selected at 37˚C with 15 minutes

stabilization.

Figure 6-1 Photo bleaching vs light intensity

The first comparison is for different light intensity. From 350K to 12k, photo

bleaching reduces while light intensity drops. Figure 6-1 shows the red line with intensity

of 350k at its starting point; blue with 100k and pink with 12k.

131
The second test was comparing photo bleaching and enzyme effect. Fig. 2 and Fig.

3 are 2 runs with different start point. Fig. 2 starts from 12k for the photo bleaching test;

just after 15min, 10µl Enzyme is added in. From Figure 6-2, the drop of photo bleaching

is more significant than enzyme effect to DHE.

Figure 6-2 Photo Bleaching vs Enzyme effect @ 12k light intensity

Figure 6-3 is another photo Bleaching and enzyme effect comparison at higher

light intensity 100k. It produces similar results as Figure 6-2 result, but larger scale.

132
Figure 6-3 Photo Bleaching vs Enzyme effect @ 100k light intensity

We are going to compare enzyme treated under SMA influence with control

enzyme. It seems to me that the control enzyme doesn’t produce a good steep slope and

the photo bleaching effect is more significant. We ran different ratio between DHE and

enzyme to increase the slope drop with ratio of DHE to enzyme from 100:10; 25:10 and

10:10. Ration of 10:10 produced the best result.

The photo bleaching effect was minimized by reducing the light passing amount.

Photo bleaching always exist in fluorescent measurement, and can’t be eliminated. But it

can be minimized to the level where it can be neglected within the measurement frame.

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7 Conclusion and future work

In this project, SMA as an actuator was studied and designed. SMA spring was

made and tested. Self made SMA springs both tensile and compressive performs

according to body temperature SMA properties. Tensile SMA can produce reliable force

for unplugging. SMA spring actuator can produce 5mm to 10mm range vibration with

1Hz to 0.5Hz under water based media cooling. Mixing process was studied both

theoretically and experimentally. Free Diffusion (Molecular Diffusion) process and

Turbulent Diffusion process were studied to help with the understanding of the mixing

process. CarboxyFluoresceine (CF) was used to measure mixing process. ISS K2

Fluorescence Measurement provided a very good understanding how well the mixing

process were done. 95% mixing were achieved both in 1atm and 300bars. The mixing

time to 90% took less than 10 seconds for both conditions. 25 seconds is the time for

reaching equilibrium. Compare to free diffusion and unreliable perturbation, which can

take a few hours or days, it is a very reliable mixing under pressure. It is a huge

achievement.

A new HPOC with electrical lead was designed and manufactured. A complete

HPOC with smart activation mechanism were tested with fluorescence measurement. The

whole system is tested under pressure. It proved SMA spring with micro-motor stirrer

work for high pressure mixing. The mixing of 95% was achieved under 300 bar.

There are still some problems existing in the current design. The mixing of two

solution wasn’t 100%. The performance of SMA spring under pressure wasn’t reliable.

The SMA spring doesn’t pull the micro-motor which function as both a stopper and a

stirrer to its highest position. These problems are due to our manufacture facility

134
limitations. Professional manufacture process will improve the performance. Quartz

cuvette fabrication has the similar problem, individual glass shop cannot control cuvette

shape as our requirement and a professional glass factory can improve the dimension and

shape control and produce unified products. Optimum design is needed to compromise

functions and reliability.

Other solutions such as ultrasonic, piezoelectric motor and stir bar are very

promising on mixing process. From our investigation, the problem of ultrasonic is that

commercially available is hard to fit in HPOC due to its size. Piezoelectric motor was

very attempting to me through my research in that no coils will be involved, less

contamination. Stir bar is a widely used in bio-chemical labs, the difficulty of using stir

bar is how to construct magnetic field around or inside HPOC. Heat generation is a

common problem for all above applications and the minimization of heat generation with

properly controlling input power is critical.

135
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143
8 Appendix

8.1 Instron force measurement of SMA springs

8.1.1 Commercial SMA spring test

Specimen 1 to 4

3.0

2.0
Load (N)

Specimen #

1.0 1
2
3
4
0.0

-1.0
0 10 20 30 40 50 60 70 80 90 100 110
Time (sec)

Figure 8-1 Low temperature tensile testing force vs time (1-4)

Specimen 5 to 8

3.0

2.0
Load (N)

Specimen #

1.0 5
6
7
8
0.0

-1.0
0 10 20 30 40 50 60 70 80 90 100 110
Time (sec)

Figure 8-2 Low temperature tensile testing force vs time (5-8)

144
Specimen 9 to 11

3
Load (N)

2 Specimen #
9
1 10
11

-1
0 10 20 30 40 50 60 70 80 90 100
Time (sec)

Figure 8-3 Low temperature tensile testing force vs time (9) unplugging force test

(10)

Specimen 1 to 4

10

6
Load (N)

Specimen #

4 1
2
3
2 4

-2
-1 0 1 2 3 4 5 6 7 8 9 10 11 12

Extension (mm)

Figure 8-4 Load vs Extension

145
8.1.2 In house made SMA spring testing

Specimen 1 to 4

8
7

6
5
Load (N)

Specimen #
4
1
3 2
3
2 4
1

0
-1
0 10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 160
Time (sec)

Figure 8-5 High Temperature Af force vs step extension loading (1-4)

Specimen 5 to 7

6
Load (N)

5 Specimen #
5
4 6
7
3

1
0 10 20 30 40 50 60 70 80 90 100 110 120 130 140 150
Time (sec)

Figure 8-6 High Temperature Af force vs step extension loading (5-7)

146
Specimen 5 to 8

4
Load (N)

Specimen #

3 5
6
7
2 8

0
0 10 20 30 40 50 60 70 80 90 100 110 120 130
Time (sec)

Figure 8-7 High Temperature Af force vs step extension loading

Specimen 1 to 4

4
Load (N)

Specimen #
3
1
2
2
3
4
1

-1
0 10 20 30 40 50 60 70 80 90 100 110 120 130

Time (sec)

Figure 8-8 High Temperature Af force vs step extension loading (1-3) failed

147
Specimen 1 to 3

12

10

8
Load (N)

Specimen #
6 1
2
4 3

0
0 20 40 60 80 100 120 140 160 180 200 220

Time (sec)

Figure 8-9 Current response (3) (by increasing current, the peak of force increases)

Specimen 1 to 1

3
Load (N)

2
Specimen #

1 1

-1
0 10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 160

Time (sec)

Figure 8-10 Stopper force measurement

148
Specimen 1 to 1

1.6
1.4
1.2

1.0
Load (N)

0.8
Specimen #
0.6 1
0.4

0.2
0.0
-0.2
0 10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 160

Time (sec)

Figure 8-11 Low Temperature force measurement

149
8.2 ISS measurement test results

8.2.1 Leakage Testing

Plug Leakage Testing

1200

1000

800
Intensity

600

400

200

0
0 500 1000 1500 2000
Time

Figure 8-12 Leakage testing

150
8.2.2 Free diffusion

Diffusion 062906 pipet feeding 1st time

600000

500000

400000
Intensity

300000 Series2

200000

100000

0
0.01
314
673
1274
1587
1897
2211
2525
2835
3149
3463
3773
4087
4400
4714
5246
time(s)

Figure 8-13 Pipet feeding to imamates free diffusion

Diffussion 062906 pipet feeding 2nd time

600000

500000

400000
Intensity

300000

200000

100000

0
311.38 909.84 1246.87 1584.06 1920.96 2254.06 2590.9 3093.39 3426.85
time(s)

Figure 8-14 Pipet feeding to imamates free diffusion

151
diffusion tube feeding 1st time

600000

500000

400000
Intensity

300000

200000

100000

0
139.69
306.63
476.32
654.95
821.73
988.45
1155.2
1321.67
1488.75
1655.32
1818.29
1984.95
2151.64
2318.75
2485.55
2652.31
2819.1
2991.99
time(s)

Figure 8-15 Tube feeding imamates free diffusion

Diffusion Testing 1st Trial

800000
700000
600000
500000
Intensity

400000 Series1
300000
200000
100000
0
0 200 400 600 800
Time

Figure 8-16 Tube feeding imamates free diffusion

152
8.2.3 SMA spring and micro-motor mixing results (1 atm) raw results

Figure 8-17 1st result

Figure 8-18 2nd mixing result

153
Figure 8-19 3rd result

154
8.2.4 SMA spring and micro-motor mixing result under 300 bars

(normalized)

Figure 8-20 1st result

155
Figure 8-21 2nd Result

156
Figure 8-22 2nd result

157
Figure 8-23 3rd result

158
Figure 8-24 4th result

159
8.2.5 Photo Bleaching

Figure 8-25 Photo bleaching at higher intensity exciting light

160
Figure 8-26 Photo Bleaching at different intensity

Figure 8-27 Photo Bleaching on more different intensity

161
Figure 8-28 Comparison of Photo Bleaching rates

162
Figure 8-29 Intensity vs Concentration (nM)

163
Vita

Mr. Oliver Hongchun Xie was born on February 9, 1972 in Qixingpao, Baoqing

County, Heilongjiang province, People’s Republic of China. He is currently a citizen of P.

R. China. He received his bachelor’s degree on Mechanical Engineering from Harbin

University of Science and Technology in 1994. After college graduation, his first job was

as a process engineer in a state owned twenty thousand employee company, Harbin

Electric Machinery Works. In 1996, he joined Philips Optical Storage in Shanghai,

China which brought the booming industry of VCD market and established Chinese new

consumer electronics era. He served as a process engineer for Compact Disc Mechanism

production, then as project leader on CD-Rewritable Module project.

Oliver Hongchun Xie came to the United States for his M.S study at Villanova

University in 2001. In July 2003, he finished his M.S. and started his Ph.D. study in the

Mechanical Engineering and Mechanics Department of Drexel University. During his

graduate study, he worked as a teaching assistant in Machine Design, Manufacturing

Process I & II. His research includes squeeze film bearing force analysis; FIB method in

making single digit nano scale hole for bio-applications; Shape Memory Alloy actuator

design, etc.

164

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