Download The Effects of Magnetic Fields on Free Radical Pairs

Some Effects of Weak Magnetic
Fields on Biological Systems and
Potential Health Effects
Frank Barnes
1.
Department of Electrical, Computer, and Energy
Engineering, University of Colorado, Boulder,
Colorado
Lecture 13
• 1. Test 1 Monday February 27th.
• 2. No paper reviews due on the February 27th.
• 3. Midterm Paper due on Monday March 6th
– Approximately 10+ pages and typically 10 +
references. However it is the content that counts.
Esaka received a Nobel Prize for 3 pages. So 3
pages can be worth an A.
4. Read papers by Bennett and Vijaylaxmi for after
your midterm exam.
Mechanisms for Magnetic Field
Interactions
• Basic Force Equation

  
F = q( E + v x B )
• Induced Electric Fields as low as 10-4V/m in
Neutrophils and 10-7 in sharks can be
detected.
• Magnetic torque on a dipole is given by

  m eff xB


Translational force on a magnetic
material


Fx  M

B
x
M is the magnetization of the material



B  o ( H  M )


= H
χ is the susceptibility tensor per unit volume.
M
   
F
BB
o
Energy Effects of Magnetic Fields
• 1. Shifts in energy with magnetic fields or the
Zeeman Effect
W  l  B  s  B
• where μl = μB L. L is the angular momentum
quantum number in units of Planck’s constant
h, and in states
where S is the
electron spin angular momentum, and is the
Bohr Magneton. B is the magnetic flux density
and
1.39967 x 104 MHz /T.
Energy Shifts with Magnetic Fields
• M, which is the projection of the angular
momentum on the direction of the magnetic
W  g J MB H
field
• The allowed transitions are for
for the
microwave field parallel to the static magnetic
field and
1 when it is transverse
Zeeman Energy Levels
Typical Molecule Has an Even Number
of Electrons
• In a typical molecule, the spins of the
electrons in the outer orbits are paired in a
singlet state. In the singlet state, S, the spins
are aligned in opposite directions or spin up
and spin down parallel to the external
magnetic field. In the triplet state both spins
are aligned parallel to the external magnetic
field or and perpendicular to the magnetic
field
or
Possible Energy Levels.
• In atoms or ions with unpaired electrons, the S-levels
or singlet states correspond to an atom or molecule
with paired spins, one up and one down. The triplet
levels T+, T_ and To,, have an unpaired spin, so that the
energy of the external magnetic field may add,
subtract, or not interact with the internal energy. The
size of the separation between energy levels
corresponds in the classical description to the Larmor
precession frequency, f, for the electron spins, with ,
f = gμBB/h
• a factor that is a function of the angular momentum
and other quantum number,
Hypothesis
Radical Pair Model:
Radical pairs form with electron spins in S or T States.
Changing pair’s energy states and spins polarization
change the recombination rate and radical
concentrations, with downstream consequences.
Proposal:
1. The AC magnetic field, BAC, at frequencies
corresponding, to the Zeeman Energy Level
differences can change in the populations
distribution.
2. Transitions can also occur at level crossings or Zero
External Field.
3. BDC changes can change recombination rates
Free Radicals
1
11
Radical Pairs in S or T States
Pair Spin = 0
Pair Spin = 0
Pair Spin = 1
For Radical Pairs Some Possible
Concentrations.
Background Experiments
1. Decreases in cell growth in fibrosarcoma HT1080 cells for
magnetic fields less than 18 µT.
(Martino, et al. Bioelectromagnetics pp:1 to 7 (2010)
2. Decreases in cancer incidence in mice and rats
(Boorman, et al., Toxicol. Pathol., 27, 267–278, 1999)
3. Increase in cell growth rates with RF and an increase in
H2O2 Concentrations of 50%
(Usselman et al. PLOS ONE 2014)
4. Measured changes in free radical concentrations
(M. Lantow et.al. Radiat Environ Biophys (2006)DOI 10.1007/s00411-006-0038-3)
5. The Interphone Study.
(International Journal of Epidemiology 2010;39:675–694)
On Average, My Temperature Is Just
Right!
• 1
Background In Physical Chemistry and
Magnetic Fields
• 1. Most of the work has been done at magnetic
field levels that are large compared to the earth’s
magnetic field of ≈ 45µT (23 to 65µT)
• 2. Kaptein [1968], Kaptein and Oosterhoff [1969],
many others, and Woodward et al. [2001],
• 3. Reviews by Steiner and Ulrich [1989], Grissom
[1995]
• 4 Book by Hayashi [2004] very good theory
• 5. Adair [1999]
Theory for Allowed Energies
•
•
1. The
Hamiltonian for a radical pair
(1)
• where
and are the electron and nuclear Zeeman terms, depending
upon the interaction between the external magnetic field B and the
electronic and nuclear magnetic moments, respectively. These terms take
the form µ•B, where µ depends on the appropriate net angular
momentum. The total angular momentum is characterized by F for low
external fields, the net electronic angular momentum is J = L + S, the sum
of the orbital and spin angular momenta, and the nuclear spin is
characterized by I. Hex is the exchange term; Hss is the electron–electron
dipolar coupling term between the members of the pair; and Hsi is the
hyperfine coupling term, that is, the interaction of the nuclear moment
with the local field due to the electronic motion, depending on J•S.
• Add
for coupling between nuclear spins in large molecules.
Theory
• 1. For magnetic fields with Bext >>45µT the spins react
with Bext and the electron spins are independent of the
nuclear spins to first order.
• 2. For Bext<< than the magnetic fields generated by the
nuclear spins then the electron and nuclear spins are
coupled and the quantum number is given by F = I +J
where is given by J= L +S ,
Bcoupling ≈ 1mT
• 3. In the case of large B we have noise from coupling to
the background and large line widths
• 4. For small B we have the inertia of the nucleus and
narrow line widths.
More complete Diagram for Transitions between Energy Levels in
Magnetic Fields without Hyperfine States.
Figure 1 Shows a simplified diagram for the energy levels for a free
radical. (Steiner and Ulrich}
In the singlet state, S, the spins are aliened in opposite directions or spin
up and spin down parallel to the external magnetic field. In the triplet
state both spins are aliened or parallel to the magnetic field.
19
Coupling of Energy Level Between States
• 1. Coupling between electronic energy levels with
different angular momentum. RF
• 2. Hyperfine States , Coupling between Nuclear
Energy Levels and the Electron Energy Levels via
the B fields Generated by Nuclear Spins with the
spins and angular momentum of the electrons, RF
• 3. Coupling between nuclear spins of adjacent
nuclei in the molecule.
• 4. Coupling between electrons in different orbits.
20
Energy Levels of Free Radicals
• Energy Levels have electron and nuclear Zeeman terms that
depend on the magnetic moments, of the electrons and
nuclei, the external magnetic field and the alignment of the
angular momentum along the net field at that location,
𝝁 •𝑩
• At low field the nuclear, electronic, and molecular rotational
angular momenta are coupled and Zeeman energy depends
on 𝝁𝑭 •B. Where F is total magnetic momentum; F=J+I, I is the
nuclear angular momentum, J = L + S + j is net electronic one;
L is net orbital, S is net spin, j is rotational.
• At higher fields, L, S, J, I couple separately to B.
• Internal couplings within the radical molecule primarily set
level energies for any B.
Energy Levels for NO
• 1
• Figure 4. Energy level diagram and complete spectrum
of the 𝐽 = 1/2 → 3/2 rotational transition of the 2∏1/2
state of N14O16. From Gallagher et al., [1954]
Energy Levels for NO
•
Figure 5. Energy level diagram of the J=3/2 level of the 2∏1/2 state of N14O16. Stage (a) is in the
absence of magnetic field. Stage (b) shows the magnetic levels considering only molecular effects.
Stage (c) adds magnetic hyperfine splittings. Stage (d) includes the nuclear electric quadrupole IJ
coupling and shows the nine transitions ∆MJ=±1, ∆MI=0. Arabic indices on the transitions
correspond to the labeling of the observed absorption lines. [from Beringer and Castle, 1950]
Hypothesis Theory
1. Relative S and T state of newly-formed free radical pairs
depends on relative orientation of the radicals’ electronic
angular momenta J.
2. Allowed Zeeman transitions include changes between
electron and nuclear spin orientations with respect to the
applied magnetic fields.
3. Applying weak AC magnetic field to radical in one Zeeman
level stimulates transitions between energy levels if
hf = ΔE.
4. If two levels have same energy, spontaneous transitions
can occur subject to some other restraints.
5. Changing relative S or T state of a pair changes
recombination probability, resulting in change in radical
concentration, with downstream consequences.
The Energy Level Diagram for D2
Recombination for Radicals
Background Theory + Experiments
RF Absorption Spectra
From Woodward et al., 2001
Population Saturation
Population difference of states in AC field is
n1
n12 
1   2 B1T2T1
Where 𝒏𝟏 is population of one state, 𝜸 =
𝝁𝑭
𝟐 𝝅 is the gyromagnetic ratio, B1 is the AC
𝒉𝒇
magnetic flux density, T1 is the relaxation time
between states and T2 is the nuclear spin
relaxation time (Bovey et al., 1988).
Free Radical Concentrations
• T2 may be seconds long
• T1 is typically in the range from 10-6 to
• 10-10sec
• Assuming T2 is seconds or longer, T1 is 10-6 10-10 sec for transition between the S and T
states of the radical pair, and if γ = 1.933 x
107 for the nuclear spin transition, then
B1 = 2.68 x10-5 to 2.68x10-9 T to reduce Δn12
by a factor of 2.
Some Examples of Radical
Concentration Changes
• Usselman et al. show that for rat pulmonary
arterial smooth muscle cells (rPASMC).
• 1. RF exposures at 7MHz and 10µTRMS for
3days in a BDC= 45µT lead to decreases of
45% in O2 -*and an increase in H2O2=50%
• 2. Enhanced cellular proliferation by up to
40% on day 2 and 45% on day 3 in
proportion to the SMF control group
Growth of P815 Mastacytoma Cells
Bdc=38µT f= 60Hz (Bingham 1996)
Some Effects of H2O2
• 1. H2O2 is both a signaling molecule and can be
destructive by conversion to radicals that lead
to the modifications of DNA and lipids.
• 2. It can stimulate growth in cancer cells at low
concentrations and lead to killing them in
high concentrations.
• 3. The generation of ROS and H2O2 is a normal
part of the metabolic process.
• 4. ROS produced via normal cell metabolism
modify approximately 20,000 bases of DNA
per day in a single cell.
A Positive Example of Concentration
Changes
• 1. Arendash et.al show exposures of
transgenic mice destined to develop
Alzheimer’s-like cognitive impairment to
0.25 W/kg at MHz reduced brain amyloidβ (Aβ) deposition through Aβ antiaggregation actions and increased brain
temperature during exposure periods
Reduced Damage from
by pre exposure RF at 900MHz
Uses of Radicals
• 1 Destroying bacteria by enveloping them and
releasing ROS.
• 2. Use NOx for control of ROS and
vasodilation of blood vessels.
• 3. Activation of the immune system.
Variations in growth rates of E.coli
with Static Magnetic Fields
Effects Of Periodic Signals
• 1. Cell often communicate with periodic pulses
or signals. This gives a better signal to noise,
S/N, than a straight amplitude signal for
things like gene expression.
• 2. Examples Ca+2 and the circadian rhythms with
temperature pulses
• 3. Signals close to the natural oscillation
frequency can pull the period and the closer
you are to the natural frequency the smaller
the signal needed to get phase locking.
Variations In the Growth Rate of Human Fibrosarcoma HT1080
Cells with ΔT=+/- 1.25oC and ΔB≈+/- 100µT
1.2
Normalized cell counts for 4 hour exposure with varying temperature pulses
Cell counts of exposed samples normalized
using counts of control sample
1
0.8
0.6
0.4
0.2
0
7s oscillation
13s oscillation
Normalized Initial Count
20s oscillation
Normalized Final Count
25s oscillation
Variations In the Growth Rate of Human Fibroblast Cells with
ΔT=+/- 1.25oC and ΔB≈ +/- 100µT
1.2
Normalized cell counts for 4 hour exposure with varying temperature pulses - Fibroblast cells
Cell counts of exposed samples normalized
using counts of control sample
1
0.8
0.6
0.4
0.2
0
7s oscillation
13s oscillation
Normalized Initial Count
20s oscillation
Normalized Final Count
25s oscillation
Variations in NADPH Concentrations
for HD1080 Cells
ΔT=+/- 1.25oC and ΔB≈ +/- 100µT
Variation in NADPH levels with frequency for temperature oscillations
18
NADPH concentration (pmol/well)
16
14
12
10
8
6
4
2
0
7s oscillation
20s oscillation
Seeding control
Incubator control
25s oscillation
Exposed
Methods
Blood collection
Centrifuge
Buffy Layer
Drawing Sample
Cell Separation
Making the Slides
• Make C-AMP sample to known concentration
(120 mM/L)
Small Needle
C-AMP
SLIDE
• Using a small needle, draw a tiny stripe of the CAMP solution on the slide
• Place a small drop of the WBC sample on the
slide
Micro Pipette
• Sample at least ½ cm away from the stripe
Sample Drop
Slide
• Cover Slip placed on sample without covering
the stripe
• Use a needle to push the slip over the stripe
until it is fully covered.
Needle
• Apply Vaseline around slip to keep moist
Cover Slip
Push Cover slip
Typical concentrations as a function of time and
distance from the strip.
Random Motion of Human Neutrophils
Positive Chemotaxis
15
20
5
1
10
White cell movement
Positive chemotaxis
3
White cell movement
without the effect of RF
radiation,
4
2
5
1
6
7
8
10
11
12
9
13
14
Few Data Samples : Speed
DC
ELECTRIC
FIELD
REVERSE
DIRECTION
AC
ELECTRIC
FIELD (V/mm)
Field Strength
(V/mm)
Speed With
(µm/min)
Speed Without
(µm/min)
6.67
12
20
-12
-20
13.3
16.67
1.12
1.2
2
2.65
3.3
0.9
1.1
3.0
3.0
3.2
3.0
3.32
3.2
3.2
20
3.97
3.32
DC magnetic field versus WBC velocity
BDC( µT)
Velocity (µm/min)
with BDC
4.93
15
w/o BDC
4.11
w/o BDC again
4.69
22.89
5.02
6.83
5.81
25.33
31.2
35.73
3.22
3.89
4.65
3.68
6.8
5.13
2.79
5.42
4.87
39.2
52.27
63.47
3.0
5.78
4.58
4.40
6.65
6.48
3.37
2.06
4.50
71.43
78.4
101.43
3.24
5.65
3.03
4.38
7.34
4.34
2.85
6.15
2.13
Neutrophil Motion with RF at 900MHz, E≈ 1V/M
Parallel to Chemical Gradient and B≈20nT
Positive Chemotaxis
43 - Ending
35- RF Removed
33
1 – Starting Point
22
'
5
8
15
Speed with and Without RF at 900MHz, E≈ 1V/M
Parallel to Chemical Gradient and B≈20nT
Under RF Field
Velocity (microns/min)
Without RF Field
Temperature (oC)
Radicals in Biology
• Reactive oxygen (ROS) and nitrogen (NOS) free radicals
occur as a part of the metabolic processes .
• NO may be used as a signaling molecule regulating ROS
that leads to a cascade of events that amplify the signal.
• NO is used in the control of vasodilation
• The body has a normal operating range for free radicals
and magnetic fields can lead to biologically significant
events when they take these concentration ranges outside
their normal range.
• This may mean we see health effects only when the
concentrations go outside the range where the feedback
and repair process can compensate for the induced
changes.
Free Radical Concentrations
• Free Radical concentrations vary by more than an order of
magnitude in time as show below.
• Normally these concentrations return to the baseline level. Resistive
stimulations over long periods of time can lead to a rise in the
baseline level we can in turn lead to aging, cancer etc. (Droge 2002)
Some Observation on Exposures to
weak EMF
• 1. Weak electric and magnetic fields can be
sensed by biological systems.
• 2. The responses are time dependent and
there are adaptive responses that can
decrease the sensitivity.
• 3. Both positive and negative effects have
been observed including the activation of the
bodies defense mechanisms, and reduction in
sensitivity high energy radiation.
Conclusions -1
1. Low level magnetic fields can lead to both
increases and decreases in the concentration
levels of free radicals such as ROS and NOS.
2. These effects will be a function of the frequency
of the of the AC fields, the angle between the AC
and DC magnetic field, the amplitudes and the
pulse repetition rates.
3. The biological effects of these fields will be a
function of time and depend on other stress in the
biological system
Conclusions -2
1. This hypothesis is consistent with experimental and
theoretical results, including both observed increases
and decreases in free radical concentration and
experimental changes in the growth rate of some cancer
cells, E.-Coli and some tumor growth rates.
2. It may also explain why we see little or no health
effects for short term exposure and different effects for
long term exposures.
3. More work will need to confirm or disprove this
hypothesis. Additional mechanisms may also be
operative.
Loss of life expectancy and total number of lost days
Cause
Days
Cause
Days
Being unmarried-male / female
3,500 / 1,600
Average job accident
74
Cigarette smoking-male / female
2,250 / 800
Drowning
41
Heart disease
2,100
Falls
39
Being overweight 30% / 20%
1,300 / 900
Fire-burns
27
Being a coal miner
1,100
Generation of electricity
24
Cancer
980
Suffocation
13
Low Socio-economic status
700
Fire arms accidents
11
Living in unfavorable state
500
Natural radiation / From nuclear industry
8 / 0.02
Army in Viet Nam
400
Medical x-rays
6
Smoking Cigar / Pipe
330 / 220
Poisonous gases
7
Job Dangerous / with RF exposure / safe
300 / 40 / 30
Coffee
6
Accidents Motor vehicle / to pedestrians
207 / 37
Oral contraceptives
5
Pneumonia, influenza / Diabetes
141 / 95
Reactor accidents-UCS / RCS
2 / 0.002
Alcohol (US average)
130
PAP exams
-4
Accidents in home
95
Smoke detector in home
-10
Suicide
95
Air bags in car
-50
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•
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•
•
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