Department of Education

Region VI- Western Visayas

Division of Bacolod City

Bacolod City National High School

Bacolod City

APPROVAL SHEET

This research proposal entitled:

EFFECTS OF PROLONGED EXPOSURE OF INFRARED LED TO

Staphylococcus aureus AND Escherichia coli

Is submitted by Mike Daniel T. Arcipe of IV-Newton, Science and Technology and

Engineering Class student, and hereby recommend for acceptance and approval.
CHAPTER I: INTRODUCTION

BACKGROUND OF THE STUDY

Imagine the evolution of technology through time. One of these is the

Infrared LED. Infrared LEDs are mostly used in remote controls. Infrared light can
also be used to transfer data between electronic devices. Mobile phones,
personal digital assistants (PDAs), and some laptops may have an infrared LED
and receiver designed for short-range data transfer. Some wireless keyboards
and computer mice also use an IR LED and receiver to replace a cable.
An infrared light-emitting diode (LED) is a type of electronic device that
emits infrared light not visible to the naked eye. An infrared (IR) LED operates
like a regular LED, but may use different materials to produce infrared light. This
infrared light may be used for a remote control, to transfer data between devices,
to provide illumination for night vision equipment, or for a variety of other
purposes.
Electromagnetic field plays a role in human body function. A study shows
that cells use electromagnetic fields to communicate with each other, thus
maintaining the electromagnetic field balance in our body which is crucial to our
health.
One bacteria found in the skin is Staphylococcus aureus bacteria. This
bacterium is classified as Gram-positive coccus based on its appearance under a
microscope. It can cause skin infections such as Impetigo, Ecthyma, and hair
follicle infections including boils and folliculitis.
The other bacterium found in the skin is Gram-negative Escherichia coli,
that causes skin infection called Necrotizing Fasciitis. Necrotizing Fasciitis is a
type of skin infection that can destroy your skin and muscles as well as
subcutaneous tissue.
This study aims to determine the effect of the exposure of Electromagnetic
Radiation from Infrared LED to Staphylococcus aureus and Escherichia coli. This
study is of value to society of what effect it may have for over exposure from
electromagnetic radiation coming from Infrared LED. This study will add to our
 knowledge concerning what electromagnetic radiation from Infrared LED will
cause to the human body and of course, the bacteria studied with it will greatly
advance. Increasing the knowledge about LED, is significant for many fields,
institutions or people in relation with electromagnetic radiation and its effects.

RESEARCH PROBLEM
A. GENERAL PROBLEM
What is the effect of the prolonged exposure of Electromagnetic Radiation
from Infrared LED to Staphylococcus aureus and Escherichia coli?
B. SPECIFIC PROBLEMS
1. What is the effect of the prolonged exposure of Electromagnetic
Radiation from Infrared LED to Staphylococcus aureus and
Escherichia coli in a span of 0 hours?
2. What is the effect of the prolonged exposure of Electromagnetic
Radiation from Infrared LED to Staphylococcus aureus and
Escherichia coli in a span of 12 hours?
3. What is the effect of the prolonged exposure of Electromagnetic
Radiation from Infrared LED to Staphylococcus aureus and
Escherichia coli in a span of 24 hours?
RESEARCH OBJECTIVES
A. GENERAL OBJECTIVES
To be able to determine the effects of the prolonged exposure of
Electromagnetic Radiation from Infrared LED to Staphylococcus aureus
and Escherichia coli.
B. SPECIFIC OBJECTIVES
1. To be able to determine the number of colonies present after the
exposure.
2. To be able to know if the Infrared LED exposure has an effect to the
number of colonies preset after the exposure.
HYPOTHESIS
A. NULL HYPOTHESIS
 1. There is no significant effect from the prolonged exposure of
Electromagnetic Radiation from Infrared LED to Staphylococcus
aureus and Escherichia coli in a span of 0 hours.
2. There is no significant effect from the prolonged exposure of
Electromagnetic Radiation from Infrared LED to Staphylococcus
aureus and Escherichia coli in a span of 12 hours.
3. There is no significant effect from the prolonged exposure of
Electromagnetic Radiation from Infrared LED to Staphylococcus
aureus and Escherichia coli in a span of 24 hours.
B. ALTERNATIVE HYPOTHES
1. There is a significant effect from the prolonged exposure of
Electromagnetic Radiation from Infrared LED to Staphylococcus
aureus and Escherichia coli in a span of 0 hours.
2. There is a significant effect from the prolonged exposure of
Electromagnetic Radiation from Infrared LED to Staphylococcus
aureus and Escherichia coli in a span of 12 hours.
3. There is a significant effect from the prolonged exposure of
Electromagnetic Radiation from Infrared LED to Staphylococcus
aureus and Escherichia coli in a span of 24 hours.
 FRAMEWORK
Independent Variables

Treatment A (3 replicates) Treatment B (3 replicates) Treatment B (3 replicates)

Staphylococcus aureus and Staphylococcus aureus and Staphylococcus aureus and

Escherichia coli Escherichia coli Escherichia coli

Infrared LED exposure for 0 Infrared LED exposure for 12 Infrared LED exposure for 24
hours hours hours

Dependent Variables

Number of colonies present

after exposure

SCOPE AND LIMITATIONS

The experiment will be conducted at the Medical Laboratory of Riverside
College, Inc. in a span of 1 week. Staphylococcus aureus and Escherichia coli
are the bacteria to be used on the experiment. This study is limited in terms of
temperature for optimal growth of bacteria, the humidity of the environment, the
equipment available, weather condition and availability of the laboratory.

SIGNIFICANCE OF THE STUDY

This study is of value to society of what effect it may have for over
exposure from electromagnetic radiation coming from Infrared LED. This study
will add to our knowledge concerning what electromagnetic radiation from
Infrared LED will cause to the human body and of course, the bacteria studied
 with it will greatly advance. Increasing the knowledge about LED, is significant for
many fields, institutions or people in relation with electromagnetic radiation and
its effects.

DEFINITION OF TERMS

CONCEPTUAL

LED - a diode of semiconductor material, such as gallium arsenide, that

emits light when a forward bias is applied, the color depending on the
semiconductor material: used as off/on indicators.
Infrared the part of the invisible spectrum that is contiguous to the red end of
the visible spectrum and that comprises electromagnetic radiation of wavelengths
from 800nm to 1 mm.
Electromagnetic Radiation Radiation consisting of electromagnetic waves
including radio waves, infrared, visible light, ultraviolet, x-rays and gamma rays.
OPERATIONAL
LED two led semiconductor light source
Infrared invisible radiant energy between micro waves and visible light
Electromagnetic Radiation a form of radiant energy released by certain
electromagnetic processes
Chapter II: Review of Related Literature
CONCEPTUAL LITERATURE
INFRARED LED
An infrared light-emitting diode (LED) is a type of electronic device that emits
infrared light not visible to the naked eye. An infrared (IR) LED operates like a regular
LED, but may use different materials to produce infrared light. This infrared light may be
used for a remote control, to transfer data between devices, to provide illumination
for night vision equipment, or for a variety of other purposes.

An infrared LED is, like all LEDs, a type of diode, or simple semiconductor.
Diodes are designed so that electric current can only flow in one direction. As the
current flows, electrons fall from one part of the diode into holes on another part. In
order to fall into these holes, the electrons must shed energy in the form of photons,
which produce light.

The wavelength and color of the light produced depend on the material used in
the diode. Infrared LEDs use material that produces light in the infrared part of the
spectrum, that is, just below what the human eye can see. Different IR LEDs may
produce infrared light of differing wavelengths, just like different LEDs produce light of
different colors.

A very common place to find an infrared LED is in a remote control for a

television or other device. One or more LEDs inside the remote transmit rapid pulses of
infrared light to a receiver on the television. The receiver then decodes and interprets
these pulses as a command and carries out the desired operation

Infrared LEDs can be used for a variety of other purposes. The U.S. Food and
Drug Administration have approved several products with IR LEDs for use in medical or
cosmetic procedures. Robots may use an infrared LED to detect objects, and some
utility meters even have an IR LED to transmit data to a tool for easy meter reading.
 LED

Light emitting diodes, commonly called LEDs, are real unsung heroes in the
electronics world. They do dozens of different jobs and are found in all kinds of devices.
Among other things, they form numbers on digital clocks, transmit information
from remote controls, light up watches and tell you when your appliances are turned on.
Collected together, they can form images on a jumbo television screen or illuminate a
traffic light.

Basically, LEDs are just tiny light bulbs that fit easily into an electrical circuit. But
unlike ordinary incandescent bulbs, they don't have a filament that will burn out, and
they don't get especially hot. They are illuminated solely by the movement of electrons
in a semiconductor material, and they last just as long as a standard transistor. The
lifespan of an LED surpasses the short life of an incandescent bulb by thousands of
hours. Tiny LEDs are already replacing the tubes that light up LCD HDTVs to make
dramatically thinner televisions.

Electromagnetic Radiation

Electromagnetic radiation is energy that is propagated through free space or

through a material medium in the form of electromagnetic waves, such as radio waves,
visible light, and gamma rays. The term also refers to the emission and transmission of
such radiant energy.

The Scottish physicist James Clerk Maxwell was the first to predict the existence
of electromagnetic waves. In 1864 he set forth his electromagnetic theory, proposing
that light--including various other forms of radiant energy--is an electromagnetic
disturbance in the form of waves. In 1887 Heinrich Hertz, a German physicist, provided
experimental confirmation by producing the first man-made electromagnetic waves and
investigating their properties. Subsequent studies resulted in a broader understanding
of the nature and origin of radiant energy.
 It has been established that time-varying electric fields can induce magnetic
fields and that time-varying magnetic fields can in like manner induce electric fields.
Because such electric and magnetic fields generate each other, they occur jointly, and
together they propagate as electromagnetic waves. An electromagnetic wave is a
transverse wave in that the electric field and the magnetic field at any point and time in
the wave are perpendicular to each other as well as to the direction of propagation. In
free space (i.e., a space that is absolutely devoid of matter and that experiences no
intrusion from other fields or forces), electromagnetic waves always propagate with the
same speed--that of light (299,792,458 m per second, or 186,282 miles per second)--
independent of the speed of the observer or of the source of the waves.

Electromagnetic radiation has properties in common with other forms of waves

such as reflection, refraction, diffraction, and interference. Moreover, it may be
characterized by the frequency with which it varies over time or by its wavelength.
Electromagnetic radiation, however, has particle-like properties in addition to those
associated with wave motion. It is quantized in that for a given frequency , its energy
occurs as an integer times h , in which h is a fundamental constant of nature known as
Planck's constant. A quantum of electromagnetic energy is called a photon. Visible light
and other forms of electromagnetic radiation may be thought of as a stream of photons,
with photon energy directly proportional to frequency.

Staphylococcus aureus

Staphylococcal aureus bacteria are classified as Gram-positive cocci based on

their appearance under a microscope. They may occur singly or grouped in pairs, short
chains or grape-like clusters. They are usually facultative anaerobes, that is, they are
capable of surviving at various levels of oxygenation, and are generally very hardy
organisms.

They are only able to invade via broken skin or mucous membranes, hence intact
skin is an excellent human defense. Once they have invaded they have various ways to
avoid host defenses. They:
 Hide their antigens to avoid an immune response

Kill infection-fighting cells (phagocytes)

Survive within host infection-fighting cells.

Develop resistance to antibiotics

Release toxins (intoxication) these do not require the presence of live bacteria
to have an effect.

Escherichia coli

Escherichia coli (E. coli) bacteria normally live in the intestines of healthy people
and animals. Most varieties of E. coli are harmless or cause relatively brief diarrhea. But
a few particularly nasty strains, such as E. coli O157:H7, can cause severe abdominal
cramps, bloody diarrhea and vomiting.

You may be exposed to E. coli from contaminated water or food especially

raw vegetables and undercooked ground beef. Healthy adults usually recover from
infection with E. coli O157:H7 within a week, but young children and older adults have a
greater risk of developing a life-threatening form of kidney failure called hemolytic
uremic syndrome.

Signs and symptoms of E. coli O157:H7 infection typically begin three or four days
after exposure to the bacteria, though you may become ill as soon as one day after to
more than a week later. Signs and symptoms include:

Diarrhea, which may range from mild and watery to severe and bloody

Abdominal cramping, pain or tenderness

Nausea and vomiting, in some people

FOREIGN LITERATURE

'Far Infrared potential benefits and the scientific basis of far infrared
ray therapy'.
 Written By: Professor Neil Piller, Team Leader Lymphoedema Assessment Clinic,
Flinders Medical Center and Flinders University, Adelaide, South Australia - 2005

2005: "There is still some uncertainty about the benefits of Far Infrared Ray (FIR) use
because there have been no large scale clinical trials detailing its benefits. However,
countless anecdotal observations, experimental work and small scale clinical trials have
strongly suggested a range of benefits (2014 - PubMed.gov now have countless
medical citations regarding far infrared and hyperthermia therapy).

Early in the awareness about FIR it was realized that Far Infrared Ray are released
from the hands (palms) of Qi Gong masters and that this, in part, explained how they
could improve people's health most likely through re-balancing a disrupted physiological
mechanism.

FIR can penetrate deeply into the tissues, generating the sensation of warmth as they
do so. There is a belief that the Far Infrared Rays set up a resonance in the body, thus
generating this feeling of gentle warmth and repair. It was later discovered that certain
ceramic substances could emit FIR naturally and when stimulated by an electric
current.

There have been many studies about the benefits of various wavelengths of laser, of
intense polarized and non-polarized light on the human body, but it is those of the Far
Infrared Rays which seem to have the widest and most systematic effect, and more
work is underway to substantiate this (PubMed.gov as well as German cancer clinics
can substantiate this).

Some recent good quality experimental and clinical scientific studies (2003) from Japan
have shown FIR to result in more rapid wound healing which was independent of
changes in blood flow and skin temperature. FIR helped this healing by stimulating a
group of cells called fibroblasts to make more collagen which is a very important part of
good wound healing and tissue repair. FIR and the gentle tissue warming has also been
shown to help improve the 'health' of the cells that line our blood vessels, which if left
untreated may increase the risk of heart disorders.

Some other exciting work in progress in Japan strongly suggests that when certain
herbal plants are subjected to FIR that their anti-cancer activity is enhanced.

We have a long way to go in terms of our knowledge and understanding of the

beneficial effects of FIR. It does seem clear however, that there are good benefits in the
use of FIR on the skin, blood circulation and other conditions where the homeostatic
mechanisms of body balance have been disrupted. FIR is what is believed to be the
basis of the healing powers of Qi Gong masters but now there are some aids which can
uniformly deliver FIR, which can be used in your home.

We all have a responsibility to ourselves to ensure good health, but to do this we must
be well informed about the best ways we can attain and maintain this and its associated
vitality. A search of "PubMed" and a reading of the booklet "The Scientific Basis and
Therapeutic Benefits of Far Infra-Red Ray therapy" by Dr. Kyuo, which our team
helped translate (for HTE), will help you in this quest.

Chapter III: Methodology

A. Procedural Design

Gathering of Materials

Preparation of Infrared LED box

Sterilization of Materials

Preparation of Culture Media

Preparation of 96-well plates

Colony
Data Gathering
Counting
 Streaking the bacteria and
incubation
B. Materials and Equipments

B.1 Gathering of Materials

- 1 piece 25.4 x 15.24 cm for the base, 1 piece 33 x 20 cm for the top
cover, 2 pieces of 25.4 x .70 cm, 2 pieces of 15.24 cm for the side and 1 m
Infrared LED strip, autoclave, detergent, sponge, distilled water, old newspapers,
test tube brush, petri dishes, L-rod, electronic balance, laboratory water bath,
graduated cylinder, alcohol lamp, glass tube, 96-well plates, wire loop,
Erlenmeyer flasks, beakers, laboratory rotator, micropipette, Mueller Hinton agar,
normal saline solution, Suntex Colony Counter 570

B.2 Preparation of LED box

- 1 piece 25.4 x 15.24 cm for the base, 1 piece 33 x 20 cm for the top
cover, 2 pieces of 25.4 x .70 cm, 2 pieces of 15.24 cm for the side and 1 m
Infrared LED strip

B.3 Sterilization of Materials

 - Autoclave, detergent, sponge, distilled water, old newspapers, test tube
brush

B.4 Preparation of Culture Media

- Mueller Hinton agar, distilled water, Erlenmeyer flask, electric water bath,
petri dish, refrigerator

B.5 Preparation of 96-well plates

-96-well plates, normal saline solution

B.6 Preparation of inoculum

-Tryptic soy agar, sterile wire loop, 0.5 McFarland

B.7 Exposure of Infrared LED

-LED box, 96-well plates, normal saline solution

B.8 streaking the bacteria and incubation

-test tube, micropipette, petri dish, L-rod, incubator

B.9 Colony Counting

-Suntex Colony Counter

B.10 Data Gathering

-Paper, pencil

C. Procurement and Preparation of Materials

For the sterilization of laboratory apparatus, the apparatus will be

washed with powdered detergent and water, then it will be allowed to dry. Each
apparatus will be covered using used bond papers in a manner of wrapping a
hamburger. The unprinted area of the bond paper was in contact with apparatus.
It will be autoclaved for 30 minutes.
 In the preparation of culture media, the Mueller Hilton agar will be weighed
and mixed with distilled water in an Erlenmeyer flash. It will be then placed in a
water bath for 15 minutes. The opening of the lids of dishes and flask will be
heated. The agar will be put into dishes at uniform depth and will be allowed to
cool and solidify. After it solidified, the petri dish will be inverted and wrapped
with paper and put into refrigerator.

For the preparation of the bacterial inoculum, E. coli and S. aureus will be
streaked in tryptic soy agar using Streak Plate Technique. Three to five isolated
colonies will be obtained using sterile wire loop and inoculated in 5 mL sterile
normal saline solution (NSS) and adjusted to 0.5 McFarland in which the
approximate cell count density is 1.5 x 108.
For the Infrared LED exposure process, 100 L of bacterial inoculum will
be inoculated to each well containing NSS to make a bacterial suspension. The
plates will be incubated with irradiance of LED blue light for 24 hours. Bacterial
density will be determined in 0Hr, 2Hr, 10Hr, and 24 Hr. incubation. 50 L of
suspension will be obtained from each well and spread in Nutrient Agar and
incubated for 24 hours at 37C.

For the preparation of streaking the bacterial culture, the researchers will
take 100 L of solution from the 2nd test tube using micro pipette, then it will be
transferred into the petri dishes and then the solution will be streaked using the
disinfected L-rod, in which the streaking direction must be horizontal, vertical and
diagonal, overlaying each direction and fully dispersing the culture into the petri
dish.

For the preparation of the 96-well plates, the wells will be filled with 100 L
sterile normal saline solution (NSS). It will be then divided into eight groups. The
first four groups for E. coli. 1st group for 0 hour set-up, 2nd group for 2 hour set-
up, 3rd group for 12 hour set-up and 4th group for 24 hour set-up. The remaining
four groups are for S. aureus. 5th group for 0 hour set-up, 6th group for 2 hour set-
up, 7th group for 12 hour set-up and 8th group for 24 hour set-up. In the counting
of the number of colonies, the researchers will divide the plate into four
quadrants and choose one quadrant. Then the researchers will mark every
colony present and then it will be automatically counted by the suntex colony
counter.

For the final procedure, the researchers will collect and compare the
results taken and observed from the two different set-ups.

In this study, data will be gathered, treated and analyzed statistically with
the use of the mean and the standard deviation were employed to determine the
average values gathered from the determination and the degree of variability of
the individual values from their means, respectively. One-Way ANOVA, set to
0.05 Alpha level of significance, determined the effect of Electromagnetic
Radiation from LED.

D. Research Locale

The experiment will be conducted at the Medical Laboratory of Riverside

College, Inc. where the needed materials are available.

E. Research Design

Preparation of Materials

Preparing of the box

Preparing the culture media

Preparing the inoculum

Preparing the well plates

Exposure of the bacteria

Streaking of the bacteria

Testing of Results

F. Experimental Procedure
 F1. Test for the number of colonies present after the exposure

This test was done by dividing the cultured bacteria into 4

quadrants so that it would be counted easily. Then the researchers will use the
Suntex Colony Counter to count the number of colonies.

G. Research Lay-Out

Set A: 0 hours of Set B: 12 hours of Set C: 24 hours of

exposure exposure exposure
Test for Trial 1 X Trial 1 X Trial 1 X
the
number
of
colonies
present

H. Variables

H.1 Independent Variable

-Time of exposure
 H.2 Dependent Variable

-Number of colonies present after exposure

H.3 Constant Variables

-Amount of inoculum, temperature, humidity, agar solution, Infrared LED

strip, distance of exposure

I. Data Gathering

-Test on the number of colonies present after exposure

J. Statistical Data Analysis

-The data gathered from the data will be analyzed with the use of the
following statistical tools:

J.1 Mean

To determine the average of the data gathered from the statistical tools.

Appendix

A. Schedule of Activities

Activities October November December January February

Title
defense
Chapter 1 7-8
Chapter 2 7-8
Chapter 3 7-8
Typing 7-8
Editing 7-8
Printing
Research
Proposal

B. Budget Estimate

Items Price
A. Communication
1. Transportation Fare Php 200
B. Materials
1. Petri Dish Php 400
2. Infrared LED strip Php 150
3. Cold-pressed board Php 300
C. Consultation Php 500
1. Consultant + Bacteria Php 2700
D. Bond Paper and Print Php 150
TOTAL Php 4400
Bibliography

http://www.wisegeek.com/what-is-an-infrared-led.htm
http://electronics.howstuffworks.com/led2.htm
http://abyss.uoregon.edu/~js/glossary/electromagnetic_radiation.html
http://www.mayoclinic.org/diseases-conditions/e-coli/basics/causes/con-
20032105
http://www.chimachine4u.com/fir.html
http://www.astrosurf.com/luxorion/qsl-em-radiation3.htm