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I N S T I T U T I O N

A Hwa Chong Institution Research Publication

ISSUE : 03/2014

ABOUT
VOYAGER

ABOUT
VOYAGER

Voyager, the brainchild of members from the Science


Voyager,
Students
the brainchild
ResearchofCouncil,
members
was
from
established
the Science Studen
in 2010. The theme varies from year to year and ininthis
2010.
third
The
issue
theme
of Voyager,
varies from
we year
explore
to year
the theme
and in this third is
Discover: Forging Ahead Together for the next
Discover:
Milestone.
Forging
This publication
Ahead Together
aims tofor
serve
the as
next
a Milestone
microcosm of the many facets of research andmicrocosm
seeks to reveal
of thethe
many
research
facets experiences
of research and
and seeks to
extensive support network of Hwa Chong Institutions
extensive
research
support
community.
network of Hwa Chong Institutions resea

Editor-in-Chief

Editor-in-Chief

Chiang Yan Xin 14S7F

Chiang Yan Xin 14S7F

Publication Team

Publication Team

Chiang Yan Li 13S7B

Chiang Yan Li 13S7B

Poh Yong Rui 13S7J

Poh Yong Rui 13S7J

Ng Jun Xuan 13S7G

Ng Jun Xuan 13S7G

Kwan Jing Ling 14S79

Kwan Jing Ling 14S79

Lee Zhi Rui 14S79

Lee Zhi Rui 14S79

Zhou Pei Yu 14S6A

Zhou Pei Yu 14S6A

Teacher Advisor

Teacher Advisor

Dr Kelvin Tan Yong Leng

Dr Kelvin Tan Yong Leng

Education Consultant/ Research

Education Consultant/ Research

www. hc i.e d u. s g

We maintain a culture of continuous transformation. This year, Voyager has evolved to


include Foundations for the Future, which feature articles with a personal and distinct
focus on the research experiences of Hwa Chongs alumni who have embarked on their
various endeavours after graduating from Hwa Chong. Our team has had the privilege of
sharing their insights on their research experiences as well as the potent driving force
behind their choices to embark on their present disciplines with you. We believe that by
sharing both the experiences of our present students as well as our alumni, we can impart
profound meaning to the relevance of Hwa Chongs research experience to the present
and the future.
We hope that this issue of Voyager will be an invigorating and insightful read that serves
as a harbinger of Hwa Chongs distinctive research culture and close-knitted research
community. We also hope that future Voyager editorial committees will continue to
present the best of our students work to you.
Yours sincerely,
Chiang Yan Xin
Editor-in-Chief
On behalf of the Voyager Editorial Committee 2014

VOYAGER 2014

The advent of 2014 marks new beginnings for Voyager. Featuring a plethora of
research-centered articles, we remain committed to bringing to you the finest of Hwa
Chongs research work.

Editorial

EDITORIAL

03

CONTENTS

06
07

SPOTLIGHT:
Nurturing Bright Sparks

09
14

FALL DETECTION SYSTEM BASED ON


KINECT SENSOR USING NOVEL DETECTION
AND POSTURE RECOGNITION ALGORITHM

INVESTIGATION INTO THE RELATIONSHIP


BETWEEN VIOLIN STRING TIMBRE AND
ITS HARMONIC STRUCTURE

57
63

GREEN SYNTHESIS OF SILVER


NANOPARTICLES USING
LALANG EXTRACT

DETERMINING THE AMINO ACIDS


INVOLVED IN INHIBITION OF PDE4B BY
STRUCTURALLY-DIVERSE COMPOUNDS

45
51

USE OF RECYCLED CLAM SHELLS


IN WATER PURIFICATION

QUORUM SENSING INHIBITION


ACTIVITY OF SELECTED TRADITIONAL
CHINESE MEDICINAL (TCM) HERBS TO
INHIBIT BIOFILM FORMATION

32
37

FOUNDATIONS FOR
THE FUTURE

STUDENTS VOICES

17
24

FOREWORD

HISTORIC VENUS TRANSIT 2012


AND INSTRUMENTATION

ACKNOWLEDGEMENTS

This research provides a method to


build a database of different
instrumental timbre. Findings in this
studycan be used in computer
generated music to create more
variance in instrumental timbre.
Yang Liuyi and Wang Jia

Investigation into the relationship between violin string


timbre and its harmonic structure

Our fall detection system is capable of processing


incoming sensor information and displaying of
results concurrently without a loss in processing
speed or performance degradation.

Lee Choon Kiat

Fall Detection System based on Kinect Sensor using Novel Detection and
Posture Recognition Algorithm

Cai Anni

Determining the amino acids involved in inhibition of PDE4B by


structurally-diverse compounds

Four versions of prototype were constructed using lala clam


and blood cockle shells. With each version, improvement
was made to enhance the feasibility of the prototype.
Ong Yinn Jaye, Yeo Kang Le and Gong Wei Jin
Use of recycled clam shells in water purification

VOYAGER 2014

05

Excerpts

The different frequency of stained colonies


for different buffers may be correlated to
the rate and type of mutagenesis that the 6
buffers confer. High rates of mutagenesis
may lead to changes in the essential amino
acids of the catalytic domain, thus
rendering the PDE product non-functional.

FOREWORD
Now in its third issue, Voyager highlights some of our students
exemplary research projects. It is heartening to see the next
generation of scientific talents eager to contribute positively to
the world around them.
Indeed, such passion has been the driving force behind humanitys
efforts to overcome barriers and expand our boundaries. In
todays complex and uncertain world, the challenges are
multifaceted and immense. However, the faith scientists have in
the transformative power of research remains strong. It is research
that will enable us to not just survive, but thrive in the future.

Dr Hon Chiew Weng


Principal
Hwa Chong Institution

Foreword

VOYAGER 2014

06

This year, our students continue to work under the mentorship of


esteemed research institutions and industry partners, such as the
Agency for Science, Technology and Research (A*STAR), National
University of Singapore (NUS), Nanyang Technological University
(NTU), and Singapore University of Technology and Design
(SUTD). Our partners demonstrate the extensive breadth and
depth of cutting-edge research that Singapore has been lauded
for. In addition, our own School Scientist Programme, started in
2012, has shown much promise. The school scientists have taken
students under their wing, proffering their wealth of experience
and knowledge. Under their mentors auspices, our students have
been inspired to think creatively and critically, and to always direct
research for the benefit of mankind. We are thus grateful for their
dedication and guidance.
In this issue of Voyager, you will read about how our students have
striven to help society and the world-at-large overcome critical
obstacles. Under the mentorship of Dr Lee Vwen Yen, from the
Institute for Infocomm Research, A*STAR, Lee Choon Kiat (12S68)
invented a fall detection system for the elderly who live alone. The
system can automatically recognise falls and notify healthcare
services to provide timely assistance, a boon to the elderly in
ageing societies all around the world. Ong Yinn Jaye (13S75), Yeo
Kang Le (13S79) and Gong Wei Jin (13S67) meanwhile explored
the use of clam shells to remove heavy metal ions from water, and
constructed a low-cost water filter. Their project is a significant
breakthrough that might significantly aid developing countries in
their critical quest for clean water. These projects and the other
equally impressive studies all have the potential to shape the
worlds future in a revolutionary manner.
Learn from yesterday, live for today, hope for tomorrow. The
important thing is to not stop questioning. Albert Einsteins quote
encapsulates what we hope to develop our budding scientists into
inquisitive scientific leaders who constantly seek to find new
hopes for the future. After all, science will remain a constructive
and guiding force only if its leaders are driven by two important
values passion and compassion, the very values that we nurture
in our students.

Spotlight:
NURTURING
BRIGHT
SPARKS
think what nobody has thought. Nobel laureate

and the Defence Science and Technology Agency

Albert

great

(DSTA). Similarly, the Nanyang Research Program

scientists, made groundbreaking discoveries in the

(NRP) helmed by Nanyang Technological University

most unsuspecting of circumstances. Hwa Chong too

(NTU) provides a platform for our students to explore

hopes to lay the foundations for such discoveries by

their

developing our students critical, creative and caring

sciences and social sciences. In addition, Hwa Chong

minds, grooming them to become leaders in the

has formed firm partnerships with local research

scientific community of tomorrow.

institutes such as the Agency for Science, Technology

Szent

Gyorgyi,

alongside

many

and

research

Research

interests

(A*STAR)

in

as

engineering,

well

as

natural

overseas

Hwa Chongs own internal research programme, the

universities such as University of Glasgow, Scotland.

Centre for Talent Development in Science (CenTaD),

These strong alliances provide a vibrant hub for

aims to promote a strong culture and passion for

scientific discoveries and opinions where the passion

research amongst its students. Under the guidance of

and curiosity of Hwa Chong students are cultivated.

experienced mentors and laboratory staff, Hwa


Chong students engage in authentic problem-solving
and gain in-depth knowledge beyond the walls of the
classroom. Equipped with specialised laboratories
with the ability to support projects in the fields of
environmental science, synthetic chemistry, biology

Hwa Chongs own internal research


programme, the Centre for Talent
Development in Science (CenTaD), aims
to promote a strong culture and passion
for research amongst its students.

and photonics, Hwa Chongs Science Research Center


(SRC) is a dynamic nexus for the exchange of

Hwa Chong provides many platforms, such as the

scientific ideas and a facility to promote innovation.

annual CenTaD Research Conference, for budding


student researchers to communicate their research

jointly

projects and receive valuable feedback from their

organised by the Ministry of Education and the

peers as well as mentors. The Singapore Science

National University of Singapore (NUS), provides

Engineering Fair (SSEF) allows for outstanding

opportunities for students to delve into research

student researchers to represent the school in

fields as diverse as robotics and engineering,

showcasing

computer science, microbiology and pharmaceutical

presenting their projects to experts hailing from the

The

Science

Research

Program

(SRP),

their

work

on

the

national

stage,

07
VOYAGER 2014

sciences under the mentorship of scientists from NUS

Spotlight: Nurturing Bright Sparks

Research is to see what everyone has seen, and to

Spotlight: Nurturing Bright Sparks

VOYAGER 2014

08

academia, research institutes and industry. Moreover,

important. Students embark on rigorous learning

Hwa Chong also hosts the annual International

journeys and they end up gaining more than just their

Science Youth Forum (ISYF), where our top science

research findings: they are empowered with the

research students are joined by like-minded peers

ability to think creatively, solve problems and

from across the globe. With activities ranging from

persevere. They see problems in a new light, think out

master classes with Nobel Laureates and Fields

of the box and by applying the valuable skills that

Medallists, to understanding Singapores research

they have learnt, offer refreshing perspectives.

landscape through visits, to dialogue sessions where


ideas are exchanged and experiences shared, ISYF
sets the stage for close interaction between these top
science talents from all over the world, enabling them
to forge valuable friendships and gain new insights
into the global research culture.
Any research student will ascertain the fact that
research is far from a bed of roses. It is rigorous and
the demanding nature of research can be at times
overwhelming, but the process of gaining new
insights and perspectives make research an enjoyable
and exhilarating experience. As Tricia Teo Wei Tian
from 14S7D shared with us, Research is really about
keeping an open mind. There is always a chance for
experimental

results

to

deviate

from

your

expectations so you cannot maintain a fixed mindset.


You have to be able to learn from each attempt, and
be willing to consider and try different ways of
approaching the same problem.
Ultimately, in research, the focus is not solely on the
end-product, as the process is equally if not more

Students embark on rigorous learning


journeys and they end up gaining more
than just their research findings: they
are empowered with the ability to think
creatively,
solve
problems
and
persevere.

FOUNDATIONS
FOR THE FUTURE
The Voyager Editorial Team had the privilege of speaking to three of our distinguished
alumni: Dr Vincent Tan Yan Fu (98S34), Dr Guo Huili (00S73) and Mr Sng Jie Han Timothy
(09S79). Leveraging on the solid foundations built during their time in Hwa Chong, Dr Tan
and Dr Guo, who are married to each other, continued to pursue their passion for research
in their present careers while Mr Sng is currently a Dentistry undergraduate at the
National University of Singapore. They share with us their research experiences and what
influenced them to embark on their present fields and disciplines
ocean and yet barnacles can still stick so sturdily to
rocks and surfaces of ships etc. Interestingly, one of
our team mates was on orthodontic treatment and
somehow we drew parallels between the barnacle
and the orthodontic brackets (braces) that were
adhered to his teeth.

the

reason

for

such

phenomenon.

This

protein-based glue, commonly termed as barnacle


cement, secreted by barnacles was found to be a
bane to the shipping industry and many material
scientists, one of whom was our co-mentor, Dr Gary
H. Dickinson, were exploring ways to develop
anti-fouling surfaces to weaken this robust adhesion
between barnacles and ships.

Could you tell us more about your research project?


My research team was investigating the potential of

With guidance from our 2 mentors (from different

barnacle cement in dentistry, more specifically as a

fields), we continued researching on how we may

form of dental material.

utilise this material for clinical dentistry,

We understand that your initial research topic was

Reflecting on your time as a student, what forms of

not directly related to the application of barnacles in

research exposure did you have and what forms of

the dentistry sector. Your team came up with the

support did you receive from the school?

ingenious idea of applying what was previously

In year 3 and 4 at High School, I was trained under the

undesirable for the shipping industry to dentistry.

Science & Math Research Programme (SMRP). Via

What was your teams inspiration for delving into

SMRP, students learnt about the basics of scientific

this cross-disciplinary research topic and how was

research from guest scientists, teachers & our seniors.

this idea developed?

We

All of us in the team had experiences with dental

projects, eventually presenting and defending our

restorations (fillings) and orthodontic treatment

research paper in the presence of teacher assessors. I

where the dental material used plays a very crucial

believe that the HCI Integrated programme has

role in determining the longevity and success of the

offered me much more liberty to participate actively

specific treatment.

in research projects and research-related activities

then

proceeded

to

conduct

experimental

such as the research collaboration cum exchange


What inspired us was a meeting with our project

programme with the Academy of Science-Virginia,

mentor, A/P Stephen Hsu. He mentioned how the

International Science Youth Forum (ISYF) and various

harsh environment of our oral cavity is akin to the

science fairs. These opportunities to learn beyond the

09
VOYAGER 2014

From there, we were very intrigued and read up on

Foundations For The Future

Our alumnus, Mr Sng Jie Han Timothy


(09S79) was part of a team of 3 HCI
students who undertook a project under
the Science Research Program, which was
co-mentored by a faculty member from the
Faculty of Dentistry as well as a marine
biology expert from the Tropical Marine
Science Institute. This project was awarded
the Gold Award at the 2010 Singapore
Science and Engineering Fair (SSEF), and
the team was selected to represent
Singapore at the 2010 International
Science and Engineering Fair (ISEF).
Timothy is currently an undergraduate at
the Faculty of Dentistry at NUS.

designated O level science syllabus has seeded in me

There were other fun-filled ISEF-organised events

the academic curiosity and drive to apply knowledge

throughout that entire week at San Jose, including

in experimental research.

the pin-exchange, Nobel panel discussion as well as


an exhibition of GOOGLEs technology (robotics and

In addition, the helpful and patient laboratory staff at

satellite technology).

Hwa Chongs Science Research Centre, especially the


laboratory manager, Madam Lim Cheng Fui, have

What challenges did your team face during your

always been pillars of support from the time I was

research and how did you overcome them?

doing experimental research in year 3 till my final year

This project has been a fruitful learning journey as I

in college. I am very grateful for their guidance and

have not only acquired a deeper understanding of the

they are certainly assets to HCI.

multi-faceted aspects of dental research but have


also grown in terms of character. The first bottleneck
came when my group spent one month brainstorming

medallists in the Singapore Science and Engineering

for a research topic but remained undecided on any.

Fair (SSEF) and represented Singapore in the

Nevertheless, with the motivation of our mentors, my

International Science and Engineering Fair (ISEF).

team persevered on and finally chanced upon a

When you first started your research, were you

unique topic that we were all excited about. Despite

expecting such an outcome?

having a slow start, the passion we had for our novel

Personally, I was hoping that our research was, as

and well thought-out project was the very source of

much as possible, conducted to a certain degree of

our motivation. Thus, while we often faced challenges

completion (that we agreed upon before we started

in the experimental phase of our research, we were

on our experiments). Receiving awards in such

always determined to overcome them as a team.

on the cake. Still, I felt that they serve as a form of

Any words of encouragement or advice for your

VOYAGER 2014

science fairs are often stated to be a bonus or icing


recognition and approval of the standards of ones

juniors?

research project. I was certainly exhilarated when we

Take ownership of your project. Be proud of it.

Foundations For The Future

10

We understand that your team emerged as gold

Could

were given this opportunity to participate in ISEF. I


Work on a topic that you are really very keen on and

would say that it is indeed by Gods grace!

this passion will likely be your source of fuel in times


you

share

with

us

more

about

your

experiences at ISEF?

of stress and fatigue as you face various challenges in


your project.

The public viewing day was certainly memorable. As


ISEF finalists, we had the chance to explain our work

If you are unexcited in your research, it is hard to

to the general public and even to one another. The

expect others (be it your peers or judges) to be

depth and scope of the projects at ISEF were

interested in it.

incredible and it was an eye opener to witness how


peers of our age can actually come up with such

Thank you very much and we wish you the very best

detailed methodologies and creative ideas to tackle

in your future endeavours!

scientific problems.

Timothy (right) with fellow Hwa Chong team members Joseph Ang (09S7A) and Clement Lee (09S6K) at the Intel ISEF 2010.

FOUNDATIONS
FOR THE FUTURE
Dr Guo Huili

Dr Vincent Tan Yan Fu


Dr Vincent Tan (98S34) graduated from the
University of Cambridge with an undergraduate
degree in Electrical Engineering as well as a Ph.D.
degree in Electrical Engineering and Computer
Science from MIT. He was a recipient of the Public
Service Commission and A*STAR scholarships.
After working at the Institute for Infocomm
Research at A*STAR, Dr Tan joined NUS as an
Assistant Professor, where he currently holds a
joint appointment at both the departments of
Electrical and Computer Engineering as well as
Mathematics.

Dr Vincent Tan

VOYAGER 2014

11

Foundations For The Future

Dr Guo Huili

A recipient of National Science Scholarships from


the Agency for Science, Technology and Research
(A*STAR), Dr Guo (00S73) graduated from the
University of Cambridge in 2005 with a B.A.
degree in Natural Sciences. In 2011, she received
her Ph.D. degree in Biology from the
Massachusetts Institute of Technology (MIT). Dr
Guo was awarded an Institute of Molecular and Cell
Biology Independent Fellowship in 2012. She is
currently an Adjunct Assistant Professor with the
Department of Biological Sciences at the National
University of Singapore (NUS), as well as the Lee
Kong Chian School of Medicine at Nanyang
Technological University. Dr Guo was recently
selected to be one of the ambassadors for the UN
Women STEM (Science, Technology, Engineering
and Mathematics) program.

Dr Guo and Dr Tan, please tell us more about your

combine my love for teaching and mentoring with the

current research field and direction.

discovery of new ideas and theorems.

Dr Guo (HG): I use molecular biology and genomics


to study how protein synthesis regulates cellular

Could you kindly share with us some insights from

differentiation. This may sound abstract but is

your research experience?

actually not: people are familiar with the idea that

HG: I learnt from my professor at MIT to never ignore

genes get turned on and off to make different types

the small details/blips/weird result that come out of

of cells; I am doing essentially that, just that I am

an experiment you never know when that blip could

focusing on a downstream step When a gene is

be telling you something important that has yet to be

turned on, messenger molecule known as the

discovered. Do not be afraid of trying out new

messenger RNA gets made, but this does not

fields/testing out new ideas; you never know when

necessarily lead to the synthesis of a protein. I am

you might find your passion/discover something

studying how this step is regulated, and how it in

because you bothered testing out your ideas (lots of

turns controls the differentiation of different cell

people dont they just say, but dont do. And thats a

types. This is important because proteins are the foot

mistake.)

soldiers that carry out most of the work in our cells.


VT: The main lesson I learned from my professor is to

deciphering how they work will impact upon many

think hard rather than do too much. I remembered

different biological processes and human diseases.

a meeting with him early on my PhD. I told him that I

Gaining insight into these mechanisms, and the

did A, B and C over the past week. He said great but

potential to come up with novel therapeutic avenues,

added that the important thing in research is to think

is what drives my interest in life science research.

deeply rather than be too mired in the details of


advise interested Hwa Chong students to be involved

VOYAGER 2014

doing, which I guess is very Singaporean. I would


Dr Tan (VT): My research is in the area of information
theory. Just as biology is the science behind modern

in research as soon as possible so that you know

medicine, information theory is the science behind

whether this is the right thing for you. To be a

communication systems. It forms the mathematical

successful researcher or academic, you need a certain

basis of why your mobile phones work and why we can

type of personality. You need to be really driven and

compress large files using g-zip efficiently. I establish

have initiative. You cant wait for things to happen.

Foundations For The Future

12

Because these pathways are so basic to a cell,

the fundamental performance limits of transmitting


information through noisy environments. Much of my

What do you find most enjoyable in doing research

work is mathematical in nature and involves using

and what do you find most challenging?

theories

in

HG: Most enjoyable: When you logically plan out an

mathematical statistics and optimization. You can learn

experiment after a lot of troubleshooting, deduce

much more about my work from my homepage.

what you expect to see, and then see the fruits of your

www.ece.nus.edu.sg/stfpage/vtan/

labour (i.e. the results are not uninterpretable

from

and

develop

new

theories

uninterpretable results are worse than negative


What motivated you to embark on research as a

results), that is extremely rewarding. The nature of

career?

research is such that 99% of the time, you are

HG: I have always liked to find out the reason behind

disappointed by what you see, but in that 1% of the

phenomena and explain how things work. When the

time when things work out, the joy is so addictive that

library couldnt answer all the why questions that I

you keep going back for more .

had, I decided that I would go into science to have a

Most challenging: In life science experiments, you

shot at finding things out myself. Growing up, I had

often need to do quite a bit of optimisation and

pretty severe eczema, and wanting to know the

troubleshooting. Knowing when to keep going, and

reason behind my condition was also why I decided

when to let go and try a new approach is not easy, and

to go into the life sciences.

is still something Im learning to do now.

VT: I was motivated to embark on research due to my

VT: The most enjoyable aspect of my job is the

inspirational teachers in school and professors in

discovery of a new result and the proof of a difficult

Cambridge where I did my undergraduate studies. In

theorem. The realization that youre the first to

particular, in the latter part of my studies in

demonstrate

Cambridge, I knew I wanted to be a professor, to

satisfying. It means youre contributed to mankinds

non-trivial

result

is

immensely

knowledge. Future references to your result would


bear your name. Isnt that cool?
The most challenging aspect is the knowledge that
when you try to prove something difficult, you know
that generations of brilliant people have tried and
probably failed so it is hard to imagine that you can
do it. I try not to be affected by this too much and to
persevere or in some cases, work on a completely
new and relatively unexplored area to break new
ground.
In your opinion, what are some qualities that a
researcher should embody?
HG: (i) perseverance, (ii) working smart, (iii) always
thinking of new angles to attack a problem, (iv)
being aware of what is going on in your field, so that
you can do point (iii).
VT: A certain naivety is sometimes crucial and can
prove the difference between success and failure. If
you dont know too much about a certain field,
youre able to bring in tools and techniques from

mathematics in my case is imperative.


Any words of advice for your Hwa Chong juniors
intending to pursue a career in research?
HG:

Have more confidence in your abilities. One

thing that was immediately apparent doing a PhD in


the US is that Americans are very confident.
Conversely, Singaporeans tend to be more reserved
and afraid of voicing out, and in so doing, often
shortchange ourselves.
VT: The training required to be a researcher is a very
long journey. It is important to continually discover
your interests along the way and not to be afraid of
learning new things but if you do choose to do
research, it is important to know the fundamentals
well.
Thank you very much and we wish the both of you
all the best for your careers and family life!

13
VOYAGER 2014

problems. Of course, having a strong background in

Foundations For The Future

some other field youre more familiar in to solve

STUDENTS VOICES
Research is formalized curiosity. It is poking and prying with a purpose
- Zora Neale Hurston

Students Voices

VOYAGER 2014

14

Every year, more than a hundred Hwa Chong Students embark on a new journey to satisfy
their curiosity through the various research programmes such as CenTaD, NRP and SRP.
These programmes provide students with the valuable opportunity to discover the
unknown and embark on an unforgettable learning journey in the realm of research.
As the saying goes, If you want to build a ship, dont drum up people to collect wood and
dont assign them tasks and work, but rather teach them to long for the endless
immensity of the sea. Through their respective research projects, students enrich their
learning experiences and enhance their understanding of their chosen area of study,
going beyond the boundary of textbooks and school curriculum. In the process, students
mature into innovative, independent and resilient individuals with acute and analytical
minds.
Each students research experience is unique and each of them has a different story to tell.
The following are but some of the responses we have collected from students who have
taken their first exploratory steps into the invigorating realm of research.
What have you learnt from your research
experience?

In your opinion, what is the most enjoyable


part of research?

You need to be very patient when conducting the

The most enjoyable part of research is to be able to

experiments, and have to be very persistent and not

learn from your mentor and peers, as well as

give up. You will also need to think out of the box,

interacting with them. When you face problems in

because

or

your experiment, they will come to encourage you

straightforward. It is more than just carrying out

and similarly, when they face problems, you are able

processes and redoing things. You can use the

to help them. This is what makes research enjoyable.

methods others have used before and modify them

It is also exciting when you learn new and interesting

to formulate a better method or simply improve on

things or when your mentor shows you how to handle

those methods.

or solve problems in a different way. It can really

research

is

not

methodological

show you the practicality of science outside of


textbooks and the rest of your academic subjects.

Rachel Ng Li En 12S70

The process of the scientific


method is a meticulous and noble
oneit seems almost too easy to
overlook the fact that mundane
processes contribute significantly
to humanity.

Students Voices

VOYAGER 2014

16

What has your research experience in Hwa


Chong Institution taught you?

Why do you enjoy research?


One of my favourite aspects of research is finding
relationships

and

unlocking

new

possibilities.

Something that seems as predictable as the path of a


shuttlecock

in

badminton

can

hold

hidden

complexities that can be exploited. Simple factors


such as speed and the terminal velocity of a
shuttlecock can dramatically change the shape of its
trajectory and these can be exploited by athletes and
shuttlecock manufacturers in their designs.

Being in Hwa Chong has allowed me to be exposed


to a plethora of research opportunities. I was very

Research is also about adapting to unforeseen

fortunate that Hwa Chong was able to provide me

circumstances.

with the environment as well as the guidance.

conducting our experiments due to unfavourable

Throughre search, I have been able to acquire a

environmental circumstances such as poor lighting

whole range of skills. Last year, I had the privilege of

and bad weather, and had to continuously adapt to

presenting my project at the annual Singapore

these situations. These variables often affected our

Science and Engineering Fair. I had to learn to

sensitive measurements and caused erratic results

present my research in the most succinct manner to

that required us to redo our experiments many times.

my fellow peers as well as the general public. After

Finding solutions to these problems and minimizing

all, what is science if we cant communicate it well?

inaccuracies is a key part of research and is one of


the most valuable skills I have learnt throughout my
research journey.

Furthermore, the process allowed me to crystallize


many of my ideas and the comments from the
experts were invaluable. They provided me with
many extensions and intriguing perspectives for me

We

faced

many

difficulties

in

Ho Kai Lun 14S6G

to further my research. I realised that research was


indeed appealing to me. I felt reinvigorated every
time I entered the laboratory. Research has become a
passion. This year, I decided take up a project with
the Institute of Materials Research and Engineering.
Together with the guidance from my mentors and
support from the school, I believe that I am able to
continue doing what I love.

Han Kai Zhong Terence 14S7F

I had to learn to present my


research in the most succinct
manner to my fellow peers as well
as the general public. After all, what
is science if we cant communicate
it well?

USE OF RECYCLED CLAM


SHELLS IN WATER PURIFICATION
INTRODUCTION

First prize in Singapore Junior Water


Prize Competition 2013
Represented Singapore at the
Stockholm Junior Water Prize
Competition 2013, Sweden

Water is critical to food production. While the


majority of agriculture is rain-fed, irrigated agriculture
provides some 40 per cent of the worlds food and
consumes 75 per cent of worlds freshwater resources
(The Gender and Water Development Report, 2003).
Supplies of freshwater are increasingly threatened by
population growth, changing lifestyles (use of more
water per capita) and pollution. A lack of water has
driven up the use of wastewater for agricultural
production in poor urban and rural communities. More
than 10% of people worldwide consume foods
irrigated by wastewater that can contain chemicals or
disease-causing organisms. Also, current rapid
industrialization in developing countries has caused
levels of heavy metals and pollutants to significantly
increase in the last couple of decades, especially
those from aquatic ecosystems. For example,
according to an investigation conducted by the State
Environmental Protection Administration, much of the
Pearl River Delta in China has been polluted by heavy
metals and that 40 per cent of farms and vegetable
plots in the region had been polluted by these heavy
metals (Wade, 2011).
Heavy metals pose a threat to the environment as
they cannot be degraded and would remain in the soil
and water for a long period of time. They produce
their toxicity by forming complexes with proteins, in

17
VOYAGER 2014

Research Mentor
Mrs Sow-Peh Yoke Keow
Hwa Chong Institution

Access to clean water is a universal human right.


Unfortunately, around 1.1 billion people globally do not
have access to improved water supply sources
whereas 2.4 billion people do not have access to any
type of improved sanitation facility (WHO-UNICEF,
2000). The most affected are the populations in
developing countries, living in extreme conditions of
poverty, normally peri-urban dwellers or rural
inhabitants. The people from these countries have to
travel great distances to collect water every day, from
water sources that are often polluted. The time spent
on the collection of water could have been spent on
more productive work for them to earn a living.
According to the United Nations Development
Programme (2006), 443 million school days are lost
each year due to water-related diseases. Falling ill due
to water related diseases also further reduces
peoples ability to do work and completing education.
According to WHO-UNICEF (2000), about 2 million
people die every year due to diarrhoeal diseases and
most of them are children less than 5 years of age.
88% of the diarrhoeal diseases are due to a lack of
access to sanitation facilities, together with
inadequate availability of water for hygiene and
unsafe drinking water. Globally, the problem is
deteriorating as cities and populations grow, and the
need for water increase in agriculture, industry and
households.

Use Of Recycled Clam Shells In Water Purification

Ong Yinn Jaye 13S75


Yeo Kang Le 13S79
Gong Wei Jin 13S67
Hwa Chong Institution

which carboxylic (-COOH), amine (-NH2), and thiol


(-SH) groups are involved. These modified biological
molecules lose their ability to function properly and
result in the malfunction or death of the cells. When
metals bind to these groups, they inactivate important
enzyme systems or affect protein structure, which is
linked to the catalytic properties of enzymes
(Adepoju-Bello and Alabi, 2005).
Many industries such as mining, electroplating and
piping discharge waste which contains high levels of
metal ions. For example, copper is widely present in
metal-bearing industrial discharges, and if not treated
can cause damage to a variety of aquatic fauna such
as fish and invertebrates (Ng et al, 2002).

Use Of Recycled Clam Shells In Water Purification

VOYAGER 2014

18

The presence of metal ions in domestic drinking water


is also a problem in many countries. Lead
accumulation causes major health problems such as
immune disorders and lead-induced intelligence
impairment, while iron contamination results in poor
tasting and unattractive water with a distinctive
reddish-brown colour (World Health Organisation,
1996). In fact, there was a case in 2009 where
residents in Singapore had reported seeing iron
contaminated reddish-brown coloured water coming
out from taps (Kreamer, 2009). Thus, it can be seen
that contamination of drinking water by corroded
pipes can indeed be a cumbersome problem.
Various kinds of methods have been developed to
purify contaminated water around the world, such as
the invention of desalination plants and sedimentation
(Lalov et al., 2000). However, such methods are not
cost effective. Developing countries cannot afford
expensive filtration systems to treat wastewater it is
difficult for them (and other developing countries) to
obtain pure drinking water.
Hence, scientists have explored alternatives which are
more cost effective. One popular alternative is the
biosorption of metal ions by metabolically inactive
non-living biomass of microbial or plant origin. Due to
their unique chemical composition, metabolically
inactive dead biomass sequesters metal ions, by
forming metal complexes from a solution and
removing the necessity to maintain special
growthsupporting conditions (Ahluwalia and Goyal,
2006).
Another group of researchers found that shells of
crustaceans such as crabs and shrimp can be used to
remove heavy metals. This is mainly because their
shells are made up of the material chitin which has the
ability to adsorb toxic metals (Karthikeyan et al.,
2005). Seashells have also been tested for their
ability to remove zinc, lead, cadmium and iron (Reilly,
2009). Researchers found that flooding ground shells
with highly contaminated water would cause the
metals to be removed by the shell. Shells consist of
aragonite, a mineral that is found to swap its calcium
ions in exchange for heavy metal ions. The metals
trapped are in a solid state and are harmless towards
the environment.

This study explores the feasibility of constructing a


filter from low cost and easily available material which
can be used to remove heavy metal ions. The ability of
three types of clam shell (mussel, lala clam and blood
cockles) in removing copper(II), iron(III) and lead(II)
ion was investigated and the most effective shell was
used to construct the prototype. If successful, such a
filter can be used in developing countries to help
alleviate the problem of access to clean water caused
by heavy metal pollution.

HYPOTHESES
1) All three types of shells are able to remove metal ions.
2) The prototype constructed using clam shell is effective in removing metal ions from contaminated water.

MATERIALS AND METHODS


Materials

Shells of lala clam, blood cockles and mussels were


collected. The shells were washed with deionised
water and dried in an oven at 70C for 24 hours. The
solutions containing various concentrations of
copper(II) ion, lead(II) ion and iron(III) ion were
prepared from copper(II) sulfate, lead(II) nitrate and
iron(III) nitrate respectively.

Preliminary studies:
1) To investigate effectiveness of removing metal ions
with shells:
0.5g of each type of shell was added to 50ml of
50ppm metal ion solution. The mixture was stirred on
a magnetic stirrer for 60 min, after which it was
filtered. The filtrate was analysed for the
concentration of metal ion remaining using a AA
spectrophotometer (AA 6300 Shimadzu) for lead (II)
and a colorimeter (HACH, DR/890) for copper (II) and
iron(III) ion. A control containing same volume of
metal ion solution but without shell was subjected to
the same experimental conditions.
2) Effect of pH on removal of metal ions
The pH of metal ion solution was adjusted to 2 or 5
using Nitric acid (HNO3) or Sodium hydroxide
(NaOH). pH above 5 was not studied as the metal ions
were observed to precipitate out as metal hydroxide
at pH greater than 5.
50ml of the solution with the desired pH was stirred
with 0.5g of shell for 60min. After 60 min, the mixture
was filtered and the metal ion remaining in the filtrate
determined.

Results and discussion

Analysis for removal of Copper (II) ions at pH 2


As shown in figure 1, all three types of shell are able to
remove Cu2+ ions, blood cockles being the most
effective (87.2%), followed closely by lala clam (80.6%)
then by mussels (33.4%). No significant difference is
seen between the effectiveness of removal by lala and
blood cockles.

Analysis for removal of iron (III) ion at pH 2


All 3 types of shells are effective in removing iron(III)
ions, with percentage removal of at least 95% for all
types of shell. The percentage removal is the highest
for mussel (98.9%), followed by blood cockles (98.6%)
and lala clam (97.7%).

Analysis for removal of Copper (II) ions at pH 5


At pH 5 however, all three types of shell show similar
effectiveness in the removal of Cu2+ ions, with mussel
shells showing the highest percentage of removal
(97.8%), followed very closely by blood cockles (97.4%)
and lala clam shells (95.3%). Unlike pH 2, there is no
significant difference among the effectiveness of
removal by the 3 types of shell at pH 5. The removal of
copper (II) ion is more effective at pH 5 than at pH 2.

Analysis for removal of iron (III) ion at pH 5


Lala clam showed the greatest percentage removal
(96.7%), followed by blood cockles (95.8%), and finally
by mussel shells (92.1%). The trend observed here is
similar to that observed for the removal of lead(II) at
pH 5, where mussel shells is least effective.

Discussion:

Lala clam and blood cockle shells are effective in


removing all three types of metal ions at both pH
studied. Mussel shell on the other hand is not as
effective in removing copper(II) ion and lead(II) ion at
pH 2, and not as effective in removing iron(III) ion at pH
5.
Clam shells are able to remove metal ions effectively
due to the presence of aragonite, a biogenic form of
calcium carbonate, which is able to readily swap its
calcium ions in exchange for heavy metal ions. The
equation is:
MCO3 + Ca2+
CaCO3 + M2+

Figure 2: Removal of lead (II) ion at different pH

Analysis for removal of lead (II) ion at pH 2


As figure 2 shows, all three types of shell are able to
remove Pb2+ ions, blood cockles showing the greatest
% removal (99.5%), followed closely by lala clam
(99.4%) then by mussels (95.4%).
Analysis for removal of lead (II) ion at pH 5
Figure 2 shows that the removal of Pb2+ ions by all 3
types of shell at pH 5 is very effective, with the
percentage removed being close to 100%. Blood
cockles showed the greatest % removal (99.3%),
followed by clam shells (99.2%), and finally by mussel
shells (99.0%). No significant difference is seen among
the effectiveness of removal of the three types of shells
as the error bars overlap.

The difference in the ability of different shells in


removing metal ions is likely due to the difference in
the composition of the shell as the amount of calcium
carbonate may vary in different types of shell. To
ascertain whether the ability to remove metal ions is
related to the amount of calcium carbonate,
experiments were carried out to compare the calcium
carbonate content in each type of shell. This was
carried out by first reacting a fixed mass of shell with
excess hydrochloric acid and titrating the unreacted
acid with standard sodium hydroxide. Results show
that mussel shell has the lowest percentage of calcium
carbonate (84.3%), followed by blood cockles (94.4%)
and lala clam (95.2%). Hence it is likely that mussel
shell was less effective as it contains a lower
percentage of calcium carbonate than the other two
types of shell.
To further validate the aforementioned hypothesis,
tests were also carried out with pure calcium carbonate

19
VOYAGER 2014

Figure 3: Removal of iron(III) ion by clam shells at different pH

Use Of Recycled Clam Shells In Water Purification

Figure 1: Removal of copper (II) ion by clam shells at different pH

at pH 5 under the same conditions as that conducted


with the shell. Results are shown in figure 4

Figure 4: Comparing clam shells with pure calcium carbonate in the


removal of metal ions

Figure 4 shows that the effectiveness of pure calcium


carbonate is comparable to that of lala and blood
cockle shells, both of which contain more than 90%
calcium carbonate. This further suggests that calcium
carbonate is the primary agent in the removal of metal
ions.

Version 2:

Figure 7: Version 2 of
prototype

Version 3:

CONSTRUCTION OF PROTOTYPE

Use Of Recycled Clam Shells In Water Purification

VOYAGER 2014

20

4 different versions of prototype were created, with


each version aiming to improve on its effectiveness
and efficiency. Lala clams and blood cockles were
selected because they generally outperformed
mussels at the 2 pH values studied. As this prototype
was developed with third world countries in mind, the
emphasis was on simplicity and ease of construction. A
recycled plastic water bottle was used to contain clam
shell, with sand at the base to prevent turbidity, and a
piece of cloth at the mouth of the bottle to prevent the
shell from flowing out. To test the effectiveness of our
prototype, 100ml of water spiked with Cu2+, Fe3+ and
Pb2+ was poured through the filter. The initial
concentrations of the metal ions were within the range
of 10-40 ppm. The filtrate was then collected and
analysed for the concentration of Cu2+, Fe3+ and Pb2+.
This process was then repeated until more than 90% of
the metal ion was removed, and the number of times to
filter this 100ml of polluted water was then recorded.

Thus, the opening was enlarged


for the second version. However,
though it was again effective with
close to 90% of metal ion being
removed after the first cycle, flow
rate improved only slightly to
10ml/hour. It was then realised
that the slow flow rate was not
due to the opening of the bottle,
but in fact due to the small
particle size of the shells that
were added, which blocked the
flow of water.

Figure 8: Version 3 of
prototype

For the 3rd version, only pounded


but not blended shells were
used. This increased the particle
size of the shells, and thus a
much more practical flow rate of
20ml/min
was
obtained
compared to previous models.
Also, this version was more
eco-friendly as compared to
previous models, only half the
amount of shells was used while
treating the same volume of
water. However, due to this
decrease in the mass of shells,
the water must be passed
through the filter several times
before the metal ions can be
removed.

Figure 9 shows that after the first cycle, the percentage


of metal ion which remains in the filtrate dropped
tremendously. At the end of fourth cycle, percentage
of metal ion has dropped to almost 0% for both
copper(II) and iron(III) and 1.13% for lead(II). Four
cycles of treatment are required before more than 90%
of each metal ion can be removed using version 3 of
prototype.

Version 1:

Finely blended lala


clam and blood
cockle shells in a
mass ratio of 1 : 1
(Total mass: 120g)
Sand

Figure 5: Version 1 of
prototype

Figure 6: Schematic diagram of


Version 1 of prototype

Version 1 of our prototype was able to remove close to


90% of each of the metal ion after the first cycle but it
was impractical as it had a very slow flow rate of
6ml/hour, and it was hypothesized that the slow flow
rate was due to the small opening at the bottom of the
bottle.

Figure 9: Percentage of metal ion remaining after each cycle of filtration


using version 3 of prototype

Charcoal
(10 g)
Clam shell
(300 g)

Sand
(20 g)

Figure 10: Version


4 of prototype

The major shortcoming of


version 3 of prototype is that
water needs to be passed
through the filter 4 times
before more than 90% of the
three types of metal ions can
be removed. This makes the
purification process time
consuming and laborious. To
overcome this problem, 4th
version of prototype was
constructed. The 4th version
of
prototype
adopts
a
modular design. Such a
design has two notable
advantages:

1) Increased interaction time between metal ion


pollutants and the prototype components.
2) Easy and convenient replacement of individual
prototype component. In addition, a layer of charcoal
was introduced to the prototype to further improve its
effectiveness in removing water pollutants as charcoal
is known to adsorb several types of pollutants
including metal ions and organic pollutants.
The mass of shell was also increased from 60 g in
version 3 to 300 g in version 4 to improve its
effectiveness. Lastly sand in the 3rd bottle filters out
big solid particles in water and ensures that the water
after filtration is clear and not turbid.

flow rate of version 4 was 19 ml/min, which was


comparable to the flow rate of version 3. Hence
effectiveness of removing metal ions was enhanced
without compromising the flow rate.
2) Determining the maximum volume of water which
can be purified
100 ml of aliquot of polluted water containing about 5
ppm of each metal ion was poured through the
prototype. Concentration of 5 ppm is a realistic
concentration to investigate as several studies have
reported the concentration of metal ions in
groundwater to be within or less than a few ppm
(Iqbal et al., 2009; Tombul et al., 2005; Minnesota
Pollution Control Agency, 2009). Filtrate was
analysed for the remaining levels of metal ions. The
process was repeated until the metal ion exceeds the
MCL (maximum contaminant level) set by United
States Environmental Protection Agency. An MCL is
the maximum allowable amount of a contaminant in
drinking water which is delivered to the consumer. The
maximum number of 100 ml of aliquots which can be
effectively and safely purified was then determined.
Results are shown in figure 12 and 13

Original 1st

2nd

3rd

4th

5th

6th

7th

8th

21
Figure 12: Contrast between the polluted water and the water filtered
through the prototype, from 1st to 8th aliquot.

Use Of Recycled Clam Shells In Water Purification

Several studies were conducted on the 4th prototype.


Results are as follows
1) Effectiveness of prototype in removing metal ions
of three different concentrations
Water containing 3 different concentrations of metal
ions (15 ppm, 10 ppm and 5 ppm) was filtered
separately using the prototype.

Figure 13: Concentration of metal ions remaining in the filtrate after


passing through version 4 of prototype.

Figure 11: Removal of metal ions of 3 different con centrations using


4 thversion of prototype

Figure 11 shows that the prototype was able to


remove at least 85% of all three metal ions of all three
concentrations after first cycle of filtration. This was a
marked improvement as compared to version 3
which requires 4 cycles of treatment. In addition, the

VOYAGER 2014

Version 4

The MCL of copper and lead are 1.3 ppm and 0.015
ppm respectively. MCL does not apply to iron as it is
regarded as a secondary contaminant which affects
the aesthetics of water such as taste, colour and
odour. Secondary contaminants are not health
threatening (United States Environmental Protection
Agency, 2012).
Figure 12 shows that all the eight 100 ml aliquots that
were filtered using version 4 of prototype was clear
and colourless - a stark contrast to the polluted water

which was yellow due to the presence of iron(III) ion.


Figure 13 shows that the concentrations of copper(II)
and lead(II) ions in all the 100 ml of filtered aliquots
were below the MCL. Although the prototype was
able to filter at least 800 ml of polluted water, it was
observed that the flow rate decreases after each
aliquot was filtered. At the 8th filtration, the flow rate
has decreased from the initial 19 ml/min to 4 ml/min.
The decrease in flow rate was due to clogging of cloth
filter by solid particles which obstruct the flow of
water. It is likely that more water could be purified if
the cloth filter is being replaced. Further investigation
is needed to determine the absolute maximum
volume of polluted water which the prototype can
filter.
3)Comparing 4th version of prototype with
commercial filter
Tests were conducted to compare the effectiveness of
the clam shell filter in removing the three metal ions
with a commercial filter. Results are shown in figure 14.

Table 1: Breakdown of the cost of 1 unit of


version 4 of prototype
Cost (in USD)
Fabric

$0.12

Charcoal

$0.010

Clam shell

Recycled

Bottle

Recycled

Sand

$0.064

Total

$0.194

Another feature of the filter is that it is easy to


construct, portable and can be assembled within
minutes anywhere, any time. In addition, clam shells
can be reused by washing it with 1% (W/V) aqueous
NaCl without apparent loss of effectiveness in
removing metal ions as the washed shell was still able
to remove close to 95% of each metal ion.

CONCLUSION

Use Of Recycled Clam Shells In Water Purification

VOYAGER 2014

22

Figure 14: Comparing the effectiveness of version 4 of prototype


with commercial filter. Initial concentration of metal ion is 15 ppm.

Figure 14 shows that the 4th version of prototype is


comparable to the commercial filter in removing all the
three metal ions, with the percentage removal being at
least 95%. The commercial filter is made of commercial
adsorbents such as activated carbon and zeolite. The
clam shell filter on the other hand is made of easily
available and low cost materials charcoal, clam shell,
sand and recycled bottles.
Table 1 reveals the cost of the individual component of
the filter. The cost of the clam shell prototype is very
affordable, with a total cost of less than USD 0.20 per
unit. The low cost of the prototype can be attributed to
the fact that two of the components (clam shell and
bottle) are recyclable materials. Clam shells play a
similar role as zeolite of commercial filter as both serve
to remove metal ions via ion exchange mechanism.
However zeolite costs about USD 76.70 per 100 g while
clam shells are recycled materials with practically no
cost. Also, low cost charcoal is used in place of the
more costly activated carbon which is commonly used
in commercial filters, thus further reducing the cost
without compromising its effectiveness in removing
metal ions.

All 3 types of shells (lala clam, blood cockles, and


mussels) are able to remove metal ions. Lala clam and
blood cockle shells are effective in removing all three
types of metal ions at both pH studied, with the
percentage of removal being above 80%. Mussel shell
on the other hand is not as effective in removing
copper(II) ion and lead(II) at pH 2, and not as effective
in removing iron(III) ion at pH 5. Four versions of
prototype were constructed using lala clam and blood
cockle shell. With each version, improvement was
made to enhance the feasibility of the prototype. The
version 4 of the clam shell prototype was able to
effectively remove metal ions with the following
features:
1) It is eco-friendly as its main components - clam shell
and bottles, are recycled materials.
2) It has zero carbon emission as it utilizes adsorption
(by charcoal), ion exchange (by clam shell) and
filtration (by sand) in purifying the water.
3) It is portable and easy to construct as it makes use
of the modular concept.
4) It is highly effective due to the multi-mechanism
approach.
5) It is cost effective and affordable.
Further studies of the prototype include investigating
whether it can purify more than 800 ml of water by
replacing the cloth filter after it is being clogged and
carrying out on-site studies with the prototype, testing
its effectiveness with real polluted water. In addition,
studies can be carried out to find out whether the
prototype can remove other types of pollutants such
as organic dyes. Finally the flow rate can be further
improved.

REFERENCES
Adepoju-Bello, A.A. and. Alabi, (2005). Heavy metals: A
review. The Nig. J. Pharm., 37, 41-45.
Ahluwalia, S., Goyal, D., (2006). Microbial and plant
derived biomass for removal of heavy metals from
wastewater. Biosource Technology, 98, 2243-225.
Iqbal, M. A., Gupta, S.G (2009). Studies on Heavy Metal
Ion Pollution of Ground Water Sources as an Effect of
Municipal Solid Waste Dumping. African Journal of
Basic & Applied Sciences1 (5-6), 117-122,
Karthikeyan, G, Mandal, N., Anbalagan, K. (2005).
Adsorption studies of iron(III) on chitin. J. Chem. Sci.,
117, 6, 663-672.
Kreamer, J. (2009, July 15). Tap water's clean, but are
the pipes? Straits Times

Nuisance Chemicals. Retrieved April 10, 2013 from


http://water.epa.gov/drink/contaminants/secondarysta
ndards.cfm
Wade, S.(2011). Dangerous elements: Heavy metal
pollution in China. China Digital Times. Retrieved
March 4, 2013 from
http://chinadigitaltimes.net/2011/07/dangerous-eleme
nts-heavy-metal-pollution-in-china/
WHO-UNICEF (2000). Global water supply and
sanitation assessment 2000 Report. World Health
Organisation, 1-2. Retrieved March 4, 2013 from
http://www.who.int/water_sanitation_health/monitorin
g/jmp2000.pdf
World Health Organisation (1996). Iron in drinking
water. Guidelines for drinking-water quality, World
Health Organisation, 1-4. Retrieved March 4, 2013 from
http://www.who.int/water_sanitation_health/dwq/che
micals/iron.pdf

Lalov, I.G, Guerginov, I.I., Krysteva, M. A., and Fartsov,


K.(2000). Treatment of waste water from distilleries
with chitosan. Water Research. 34, 5, 1503-1506.

Reilly, M. (2009). Sea shells used to clean up heavy


metals. Discovery News. Retrieved March 4, 2013 from
http://dsc.discovery.com/news/2009/04/27/shells-met
als-water.html.
The Gender and Water Development Report (2003).
Gender Perspectives on Policies in the Water Sector.
Loughborough, United Kingdom, Water, Engineering
and Development Centre, Gender and Water Alliance.
Retrieved March 4, 2013 from
http://www.genderandwateralliance.org/reports/GWA
Annual Report.pdf.
Tombul, M., Koparal, A.S., Ogutveren, U.B. (2005).
Transport modelling of copper contamination in
groundwater caused by a wood preservation plant.
Water Qual. Res. J. Canada, 40, 4, 469-475.
United Nations Development Programme (2006).
Beyond Scarcity: Power, Poverty and the Global Water
Crisis. Human Development Report. Retrieved March 5,
2013 from
http://hdr.undp.org/en/reports/global/hdr2006/
United States Environmental Protection Agency (2012).
Secondary Drinking Water Regulations: Guidance for

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Ng, J. C. Y., Cheung, W. H., McKay, G. (2002).


Equilibrium Studies of the Sorption of Cu(II) Ions onto
Chitosan. Journal of Colloid and Interface Science. 255,
6474.

23

Use Of Recycled Clam Shells In Water Purification

Minnesota Pollution Control Agency (2009). Cadmium,


Lead and Mercury in Minnesotas Ground Water.
Environmental Outcomes Division, Ground Water
Monitoring & Assessment Program. Retrieved April 10,
2013 from
http://www.pca.state.mn.us/index.php/view-document
.html?gid=6312

QUORUM SENSING INHIBITION


ACTIVITY OF SELECTED
TRADITIONAL CHINESE MEDICINAL (TCM)
HERBS TO INHIBIT BIOFILM FORMATION
INTRODUCTION
Chua Jia Jie 13S7G
Ng Jun Xuan 13S7G
Ng Wei Hao 13S7J
Hwa Chong Institution
Research Mentor
Dr Ng Tze Chiang Albert
Hwa Chong Institution
Singapore Science and Engineering Fair,
Silver Award

Understanding of biofilms, first reported more than 60


years ago (Zobell, 1943), has increased tremendously.
Biofilms are structured communities of bacterial cells
living on the interface of a solid-liquid or liquid-liquid
media, enclosed in a self-produced polymeric matrix.
Such biofilm formation naturally occurs due to the
many survival advantages conferred to individual
bacterium when living in a community. Moreover,
bacteria thrive on synergistic cooperation within the
biofilm environment. As such, bacteria evolved ways to
form communities as layers of biofilm (Simes et al.,
2010).
Biofilm formation is a process that begins from the
adsorption

Quorum Sensing Inhibition Activity Of Selected


Traditional Chinese Medicinal (TCM) Herbs To Inhibit Biofilm Formation

VOYAGER 2014

24

of

molecules

such

as

proteins

and

polysaccharides onto the substratum. After initial


conditioning of the surface with biopolymers, motile
bacteria swim towards the surface and adhere to it.
Such movements occur especially since there is a
greater concentration of nutrients near the surface of
the substratum. Bacterial adherence is followed by the
secretion of extracellular polymeric substances (EPS).
Subsequently colonisation of the surface by the
primary bacteria begins. As the biofilm begins to
swarm along the surface, secondary bacteria are
attracted towards the matrix. Once a mature biofilm is
established, it will actively propagate. Upon reaching a
threshold, where the biofilm becomes overcrowded,
dispersal occurs such that bacteria released are able to
colonise other areas (Lewandowski and Boltz, 2011;
Annous et al., 2008).
Despite the ubiquitous nature of biofilms, this state of
living creates numerous problems. The presence of
biofilms is particularly problematic in food, medical
and environmental related fields. Biofilm contamination
of food products is a serious hygiene issue that will
lead food spoilage and equipment impairment (Simes
et al., 2010). Biofilms found on medical devices and
implants have serious implications. Such internal
biofilms are the cause of 65% of all microbial infections.
Common

infections

like

urinary

tract

infections,

catheter infection, and even common dental plague are


difficult to treat due to the antimicrobial resistance
conferred to bacteria by the structure of the biofilm
(Lewis, 2001). In an industrial setting, biofilms cause

metal corrosion in pipeline and tanks. Microorganisms

Advanced

in biofilms reduce metals and metal alloys as sources of

research fields have shown that biofilm formation could

energy, leading to an accelerated rate of corrosion

be

(Lewandowski and Boltz, 2011). Biofilms found on

Quenchers (QQ), which inhibit bacterial cross-talk or

filtration membranes used in water and wastewater

cell-to-cell communication. This inhibition process is

treatment systems must be periodically removed as

known as Quorum Sensing Inhibition (QSI). QSI may

they cause physical blockage of membrane pores,

proceed in 3 main ways, namely (i) inhibition of

thereby increasing resistance towards water flow

receptors to signal molecules; (ii) inhibition of signal

through the membrane. Biofilms may also release

biosynthesis or inhibition of activity of autoinducer

chemical compounds which subsequently adversely

acyl-homoserine lactone (AHL)-producing enzymes

affects permeate quality (Mafirad et al., 2011). All

and (iii) signal degredation (Khan et al., 2009). The first

biofilm

associated

problems

eventually

lead

biotechnological

controlled

by

techniques

compounds

in

called

other

Quorum

to

method can be carried out by antagonising the binding

economic losses as practitioners have to constantly

of AHL at receptors that resemble those of competitive

combat and remove the bacterial film.

inhibitors in enzymatic activity. Signalling molecule


production is currently undeveloped and only a few

inhibiting microbial growth. The most common method

biosythesis in Pseudomonas aeruginosa. The approach

is primarily based on introducing antibiotics or

to destruction of signalling molecules, however, is

antimicrobial agents to kill bacteria cells. However,

accomplished via controlling local pH or by specific

rampant and indiscriminate use of such compounds led

enzymes (Kociolek, 2009), such as AHL lactonase and

to increased resistance by bacteria through the

AHL acylase (Decho et al., 2009). Studies have shown

evolution of new resistance mechanisms and pathways

that plants, such as pea plants, are able to secrete

(Huerta et al., 2008). Even with introducing stronger

AHL-signal mimic compounds that overwhelm the

agents, bacterial cells are advantaged by the physical

effects of the actual QS mechanism in plant-associated

architecture of the biofilm environment (Flemming,

bacteria (Teplitski et al., 2000).

2009). Therefore, there is a need to develop strategies


that control the very basis of biofilm formation.

Traditional Chinese Medicine (TCM) is characterised as


a potent combination multiple compounds that work

Quorum Sensing (QS) is the process that is believed to

synergistically. It has been used, up till today, to treat a

cause biofilm formation. Occurring in many species

variety of diseases. Studies have shown that some

across gram-negative and gram-positive bacteria, it is a

herbs possess antimicrobial and antifungal properties,

mechanism

for example, cinnamon (Prabuseenivasan et al., 2006).

for

intra-species.

cross-talk
Microbial

between

inter-

communities

and
utilise

As

different

herbs

express

varying

quenching

extracellular signalling molecules called autoinducers,

compounds,

as the main mode of communication. As population

significant QSI properties that can be harnessed. QSI

density of a microbe increases, concentration of

achieved using TCM plant compounds, is expected to

autoinducers increases up to a given threshold in the

be a more ideal solution in controlling biofouling as it is

surrounding

environment

(Kociolek,

there

may

be

herbs

that

possess

2009).

inexpensive and environmentally friendly. Such natural

Autoinducers are detected by, and binds to receptors

products already being consumed as food or medicinal

on neighbouring bacteria, transmitting signals, which

herbs are expected to create less downstream

are transduced by receiving molecules into intracellular

problems should they be used in water treatment or

biochemical signal, stimulating a series of physiological

medical industries. Specifically towards membrane

adaptations, such as the formation of a biofilm layer

bioreactors (MBR) for water reclamation, this method

(Raffa et al., 2004). Autoinducers serve as precursors

will reduce reliance on physical and chemical control

to biofilm formation, by acting as chemical signals

strategies that are largely inefficient and laborious,

communicated between bacteria. These compounds

providing an effective method to control biofouling as

are actively transmitted throughout the stages of

well as a pathway to a continued supply of freshwater.

biofilm formation, including bacterial motilities such as

As most herbs are found to have bactericidal

movement in a medium or a solid surface, or

properties, they will be able to destroy bacteria and

agglomeration. By quenching the signals produced by

prevent biofilm formation (Koh and Tham, 2009).

the molecules, information cannot be passed between

However, in order to co-opt this treatment into

bacterial cells, inhibiting motility and the initiation of

biofouling control in MBRs, only herbs with QSI

biofilm formation.

properties are required.

25
VOYAGER 2014

QQs have been identified for use in targeting signal

Quorum Sensing Inhibition Activity Of Selected


Traditional Chinese Medicinal (TCM) Herbs To Inhibit Biofilm Formation

Current methods of biofilm control are based on

The objectives of the research were to investigate the

Antibacterial Assay

QSI activity of various TCM herb extracts for reducing

Bacteria cultures were each inoculated across the

biofilm formation on membranes in an MBR. This was

surface of the agar. 9-mm diameter wells were bored

done by incorporating bacteria, membrane as well as

into the agar of each plate using a glass borer. 50 L of

extracts in a suspension. This study further determined

the sterile herb extracts solutions were added into the

the inhibition process on various bacterial motilities,

wells of each plate with different bacteria. Sterile water

since the motilities, complimented by autoinducers,

and bleach were also added into the wells of each plate

serve as the basis of biofilm formation. By targeting the

with different bacteria as the negative and positive

different aspects of biofilm formation separately, the

control,

different

appropriate temperatures overnight and diameter of

extracts

were

then

assessed

for

their

effectiveness in greater depth.

respectively.

Plates

were

incubated

at

the zones of inhibition measured after the incubation


period.

MATERIALS AND METHODS


Modified Microtitre Plate Assay

Quorum Sensing Inhibition Activity Of Selected


Traditional Chinese Medicinal (TCM) Herbs To Inhibit Biofilm Formation

VOYAGER 2014

26

Plant Material and Extraction Method

Sodium alginate solution was prepared and mixed with

The extraction methods used in this experiment were

the herb extracts to obtain a 3% alginate solution at a

adapted from Yeo and Tham (2012). 6 TCM herbs were

final concentration of 30 mg/mL of herb extract. The

chosen for their potential QSI ability (Table 1). 5 g each

solution was then added drop-wise to 2% calcium

of the TCM herb was ground into small pieces with

chloride solution to produce alginate beads containing

mortar and pestle. Dried ground plant material was

extracts.

soaked in 50% acetone at a ratio of 1:6 (wt:vol) on a

incorporated with a 10 x 5 mm Millipore polyvinylidene

shaker overnight. The mixtures were filtered with filter

difluoride (PVDF) membrane and 2 calcium alginate

paper, of nominal pore size of 5

beads

m, to remove

in

mL

of

each

bacterial

micro-centrifuge

culture

tube.

Tubes

was

were

particulate matter. Filtrates were vacuum-dried and

incubated and shaken for 5 d, before biofilms on the

stored at 4oC. Extracts were reconstituted in deionized

membrane were stained with 1% crystal violet solution

water to obtain final working concentrations of 25, 50,

and its absorbance value read at 600 nm.

and 75 mg/mL. Extracts were filter-sterilized using 0.22


m (pore size) Iwaki filter disks before application.

Swimming Motility
Sterile herb extracts were individually mixed with 10

Bacterial Strains and Culture Conditions

mL of 0.3% LB agar to final concentrations of 25, 50

Both Gram positive and Gram negative bacteria were

and 75 mg/mL, and poured immediately over an agar

used as model strains in this study. Gram negative

plate. Bacteria cultures were stab inoculated at the

bacteria used were Escherichia coli (ATCC 25922), and

centre of the agar after solidification. Plates were

Agrobacterium tumefaciens (ATCC 33970). Gram

incubated at appropriate temperatures for 3 d.

positive bacterial strain used was Staphylococcus

Swimming

epidermidis

(ATCC

12228).

Each

bacterium

migration

distance

was

assayed

by

was

following the colony fronts of the bacterial cells.

cultured in 10 mL of DifcoTM LuriaBertani (LB) media,

Swimming assays were performed in eighteen identical

and incubated overnight on a shaker spinning at 200

plates on three independent occasions. Measurements

rpm at 28 C. Bacterial cultures were diluted with LB

were taken every 24-h throughout the course of 3 d.

broth to an absorbance value of 0.1 at 600 nm before

Bacterial cultures used for swimming motility tests

being used in subsequent studies.

included E. coli and A. tumefaciens.

Table 1. Traditional Chinese Medicinal herbs used.

Name (Scientific)
Angelicae Sinensis Radix
Asini Corii Colla
Panax notoginseng
Persicae Semen
Rhizoma Curcumae Longae
Salviae Miltiorrhizae Radix

Swarming Motility
Sterile herb extracts were individually mixed with 10

Name

Name

(Chinese Characters)

(Hanyu Pinyin)

mL of 0.75% molten LB agar to final concentrations of

Dang Gui
E Jiao
Tian Qi
Tao Ren
Jiang Huang
Dan Shen

25, 50 and 75 mg/mL and poured immediately over an


agar plate. 1 L of E. coli culture was point inoculated
onto the surface at the centre of the agar plate after
solidification, and the surface was air-dried for 10 min
and incubated at 35oC. Extent of swarming was
determined by measuring the area of the colony in
square millimetre. Swarming assays were performed in
eighteen identical plates on three independent occasions,

with measurements taken every 24-h throughout the

Modified Microtitre Plate Assay

course of 3d.

Extracts immobilised into alginate beads were added


into a bacterial broth containing membrane pieces for

RESULTS AND DISCUSSIONS

delivering QQ in a controlled manner into the broth


culture. The modified microtitre plate assay tested the

Antibacterial Assay

effectiveness of the various extracts on an actual

Preliminary tests were conducted to identify herb

membrane. Bacteria were allowed to form biofilms on

extracts containing antibacterial compounds after the

membrane surfaces under the influence of the extracts.

extraction process. Table 2 shows results of the

A lower absorbance value recorded signified an

well-diffusion tests, with sterile water and bleach as the

inhibition of the bacteria to form extensive biofilms on

negative and positive controls, respectively. None of

membrane surfaces.

the TCM extracts exhibited any antibacterial activity


A.

Figure 1a shows the effect of herbs on E. coli biofilm

tumefaciens. However, herb extracts Angelicae Sinensis

Gram-negative

formation on PVDF membrane slices. With the

Radix and Salviae Miltiorrhizae Radix were seen to be

exception of Persicae Semen, all extracts showed

antibacterial

varying effectiveness in biofilm control. In order of

against

bacteria,

E.

Gram-positive

coli

and

bacterium,

S.

epidermidis.

increasing efficacy, Panax notoginseng, Angelicae


Sinensis Radix, Asini Corii Colla, Salviae Miltiorrhizae

Table 2. Zones of inhibition recorded for antibacterial assay.

Radix, and Rhizoma Curcumae Longae were found to

Zones of Inhibition (mm)

inhibit biofilm concentration as evident from the lower

E. coli
9.0 0.0
9.0 0.0
9.0 0.0
9.0 0.0
9.0 0.0
9.0 0.0
9.0 0.0
17.0 0.0

absorbance values compared to the control. Both

* Well diameter = 9.0 mm. Antibacterial property was


not observed when a value of 9.0 was recorded.

tumefaciens. Surprisingly, less extracts were now

Panax notoginseng and Angelicae Sinensis Radix were


able to marginally suppress bacterial attachment and
agglomeration of E. coli on the membrane surface by
approximately 17%. Salviae Miltiorrhizae Radii and
Rhizoma Curcumae Longae were able to inhibit and
restrict biofilm formation of E. coli by 46% and 57%,
respectively.
Figure 1b shows the effect of herb extracts on A.
effective in preventing biofilm formation even though
both E. coli and A. tumefaciens were Gram negative

Bacteria present within MBRs function to break down

bacteria. This was likely due to different autoinducers

waste in wastewater influent. The metabolic processes

being expressed in each bacterial strain. Among Gram

of bacteria are responsible for the biodegradation of

negative bacteria, two main types of AIs have been

bioorganic

in

classified: AI-1 and AI-2 (Niu et al., 2006). E. coli

wastewater, leading to the overall purification effect

expresses AI-2 autoinducers while A. tumefaciens

(Gallert and Winter, 2005). Therefore, TCM extracts

secrete

possessing antibacterial compounds are discouraged

highlights the different abilities of the extracts in

from use due to its bactericidal effects on such useful

targeting a wide range of autoinducers. Panax

bacteria, which can possibly lead to a reduction in

notoginseng and Angelicae Sinensis Radix were found

treatment

compounds

to be ineffective, to the extent the biofilm growth was

exhibiting strong antibacterial activity, any reduction in

promoted by 15% in the latter. Asini Corii Colla and

biofilm formation or motilities as described may not be

Persicae

attributed

especially

concentration by up to 20%. Salviae Miltiorrhizae Radii

important since the effect of biofilm reduction may be

and Rhizoma Curcumae Longae were the most

attributed to the toxicity of the extracts to the

effective herbs in controlling A. tumefacians biofilm

bacterium tested, as such Modified Microtitre Plate

growth where bacterial growth on membranes was

Assay results for herb extracts of Angelicae Sinensis

restricted by 50% as compared to the control.

and

organic

efficacy.

to

QSI.

compounds

Moreover,

This

present

for

distinction

is

AI-1

autoinducers,

Semen

were

able

and

to

this

discrepancy

control

bacterial

Radix and Salviae Miltiorrhizae Radix on S. epidermidis


should not be treated as QSI effect.

In preventing biofilm formation of S. epidermidis


(Figure 1c), only Salviae Miltiorrhizae Radix and

27
VOYAGER 2014

S. epidermidis A. tumefaciens
9.0 0.0
9.0 0.0
9.0 0.0
9.0 0.0
9.0 0.0
9.0 0.0
11.3 3.2
9.0 0.0
32.0 1.0
9.0 0.0
9.0 0.0
9.0 0.0
Sterile water
9.0 0.0
9.0 0.0
Bleach
23.7 1.2
18.3 0.6

Quorum Sensing Inhibition Activity Of Selected


Traditional Chinese Medicinal (TCM) Herbs To Inhibit Biofilm Formation

against

Persicae Semen were effective. Both herbs were able to


control the amount of biofilm formed on the membrane
by 20% on average. However, as compared to the
previous results, the other herbs were unable to inhibit
bacterial agglomeration of S. epidermidis. This could
again be due to the difference in autoinducers utilised
by the bacteria. Gram positive S. epidermidis expresses
autoinducer oligopeptides (AIPs), and effectiveness
against preventing biofilm formation of S. epidermidis
on membranes correlated to the ability of the extracts
in preventing cell communication via AIPs.
(a)
However, Salviae Miltiorrhizae Radix was found to
possess an antibacterial effect against S. epidermidis.
Therefore, results of relating to the effect of biofilm
inhibition by Salviae Miltiorrhizae Radix may not be due
to QSI but due to its toxicity towards the bacterium. In
contrast, Persicae Semen has not been found to be
antibacterial against S. epidermidis, thus the QSI
properties of this herb extract can be ascertained in the
Modified Microtitre Plate Assays since it was not found
to be toxic towards the bacterium.

Quorum Sensing Inhibition Activity Of Selected


Traditional Chinese Medicinal (TCM) Herbs To Inhibit Biofilm Formation

VOYAGER 2014

28

Results from the Modified Microtitre Plate Assays


(b)

indicated that specific herbs were effective to only


specific autoinducers. This indicated that while each of
the extracted contained QQ, in order to eliminate
environmental biofilms, which contain a mix of
Gram-negative and Gram positive bacteria, a cocktail
of extracts should be used. Understanding the
mechanism to which each of the extracts prevented
biofilm formation could yield more insights towards
producing such a cocktail. Motility studies were
conducted to determine which stages of the biofilm
process were the autoinducers inhibited.

(c)

Inhibition of Swimming Motility


Figure 1. Effect of herb extracts on biofilm formation on PVDF
membrane slices. Error bars represent standard error of data; (a) E.
coli; (b) A. tumefaciens; (c) S. epidermidis.

Swimming motility refers to the ability of bacteria to


travel

through

liquid

medium.

In

biofilm

developmental mechanism, swimming motility was


postulated to be the first course of action wherein
bacteria swim towards a solid stratum, which will be
the membrane surface in an MBR. Such actions were
thought to be due to a higher concentration of solutes
near the solid stratum. Effects of herb extracts and
their concentrations on swimming motilities of E. coli
and A. tumefaciens are shown in Figures 2 and 3,
respectively.

(a)

(a)

(b)

(b)

(c)

(c)

Figure 2. Effects of herbs on swimming of E. coli. Error bars represent

Figure 3. Effects of herbs on swimming of A. tumefaciens. Error bars

standard error of data; (a) Day 1, (b) Day 2, (c) Day 3.

represent standard error of data; (a) Day 1, (b) Day 2, (c) Day 3.

Rhizoma Curcumae Longae and Persicae Semen at all

Figure 3 shows the effect of TCM herb extracts on

concentrations inhibited the swimming motility of E.

swimming motility of A. tumefaciens. Similarly, it was

coli throughout the 3 days (Figure 2). Their inhibitory

observed that Rhizoma Curcumae Longae, Persicae

properties were further enhanced in Days 2 and 3,

Semen and Asini Corii Colla inhibited swimming

where there was a significant difference in the

motility of A. tumefaciens throughout the 3 days. The

swimming radii of E. coli in the negative control plate

properties were further enhanced in Days 2 and 3.

and in the plates with Rhizoma Curcumae Longae and

However, Persicae Semen was the most effective

Persicae Semen added. This difference in swimming

amongst these 3 herbs, showing up to 90% inhibition of

motility of E. coli was approximately 80% on Day 3.

the swimming motility of A. tumefaciens by Day 3.

Panax notoginseng at low concentrations was also


effective in inhibiting the swimming motility of E. coli

Both sets of results showed that Rhizoma Curcumae

throughout

Panax

Longae and Persicae Semen were effective against

notoginseng at high concentrations only revealed its

swimming motility of E. coli and A. tumefaciens. These

swimming inhibitory abilities on Day 3.

results reflected that compounds in both extracts

the

days.

Interestingly,

possessed the ability to restrict bacterial movement

Quorum Sensing Inhibition Activity Of Selected


Traditional Chinese Medicinal (TCM) Herbs To Inhibit Biofilm Formation

VOYAGER 2014

29

through semi-solid or aqueous mediums. Presence of


inhibition decreases probability of bacterial adhesion
onto membrane surfaces in MBRs as was observed in
the modified microtitre plate assays, reducing bacterial
concentration on membrane surfaces to less than that
of biofilm formation threshold.
Consequently, since membrane biofouling consitututes
of the formation of a biofilm on the surface, in which

(a)

pores become blocked, thereby restricting the smooth


flow of water, the use of QQ into an MBR will prove to
be beneficial in reducing such fouling occurences
(Yeon et al., 2009). As biofilm initiated fouling
dominates at sub-critical flux conditions in an MBR (Ng
and Ng, 2010), the aid of QQ to inhibit swimming
motilities of bacteria, will allow the bacteria to remain
suspended within the mixed liquor.
Inhibition of Swarming Motility
The swarming motility of bacteria is the ability of the

(b)

bacteria to congregate and grow to reach a certain


concentration upon adhesion onto a liquid-liquid

Quorum Sensing Inhibition Activity Of Selected


Traditional Chinese Medicinal (TCM) Herbs To Inhibit Biofilm Formation

VOYAGER 2014

30

interface or solid-liquid interface. At this stage, QS


occurs at its peak and EPS is secreted, forming the
basis of the biofilm.
From Figure 4, Rhizoma Curcumae Longae was able to
inhibit the swarming motility of E. coli only at Day 1, but
this effect could not be sustained through Days 2 and
3.

While the effectiveness of Asini Corii Colla was

(c)

marginally ineffective, given adequate time, on Day 2

Figure 4. Effects of herbs on swarming of E. coli. Error bars represent

and Day 3, Asini Corii Colla was able to inhibit the

standard error of data; (a) Day 1, (b) Day 2, (c) Day 3.

swarming motility of E. coli of up to 15%. As for the


remaining herbs, they showed to be ineffective in

CONCLUSIONS

preventing bacterial swarming, evident by the increase


of area of bacterial colony. The promotion of bacterial

All TCM herb extracts tested in this research exhibited

swarming in E. coli could be due to the compounds

varying abilities in targeting the issue of biofouling.

exhibiting effects that amplifies the production of

Most evidently, Rhizoma Curcumae Longae, Asini Corii

autoinducers, or increases bacterial movement along

Colla and Persicae semen showed potential in the

the interface by acting as an energy source for the

combating and prevention of biofilms in Gram positive

bacteria.

and Gram negative bacteria on a whole. Time-based


analysis, however, showed discrepancies in the effects

Swarming motility comes as the second yet most

of the various extracts over time, and have to be

crucial step in biofilm formation. Antagonising the

analysed further. Different herb extracts displays

process of swarming prevents the basic structure of

varying effectiveness against membrane biofouling, as

biofilm from forming, as well as succeeding expansions

shown by the membrane titre assay. By analysing the

of the biofilm layer. Extracts of Rhizoma Curcumae

in-depth processes in which biofilm prevention occurs,

Longae and Asini Corii Colla both display varying

the individual effectiveness of each herb can be

effectiveness of swarming inhibition over time. By

deduced and more accurately justified. A combination

inhibiting swarming motility of E. coli, bacteria is

of extracts can also be derived, maximising the

unable to communicate via signalling molecules at high

potential of each herb against the various motilities, to

threshold values, preventing a series of physiological

further combat the issue of membrane biofouling to its

chain reactions and subsequent biofilm formations.

best ability.

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essential oils. BMC Complementary and Alternative
Medicine, 6(39).

Quorum Sensing Inhibition Activity Of Selected


Traditional Chinese Medicinal (TCM) Herbs To Inhibit Biofilm Formation

Gallert C. and Winter J. 2005 Bacterial metabolism in

What is an important quality you think


research students should possess?

passionate will they be dedicated, putting in effort


constantly, and seeing the project through to
completion, and this will build up to some discovery,
be it small or large. This discovery may not have a
significant impact on the world as we know it, but if
they pursue their passion in the future, this initial
research experience will definitely be beneficial, and
they will continue to push the boundaries of science.
Some might see spending countless hours in a lab as
dry and dull, but for those who have a passion for

What inspired you to take up research?


My research project involved the study of the
metabolic pathways of FAT10, a ubiquitin-like protein
which was found by the National Cancer Centre
Laboratories to be involved in the disruption of
cellular checkpoints of cell division. This was a topic
that extended what I learnt as a biology student in
class about cancer, and therefore brought real-life
implications and applications of cancer studies into
my experiences. Through the course of the project I
was inspired by how the seemingly mundane tasks of

research, they see meaning in what they do.

transferring extremely small amounts of substances

In what ways have Hwa Chong supported


you?

results leading to the contribution to potentially

I would like to thank Hwa Chong for its amazing


teachers, as without my mentor's help, my research
team would not have had the opportunities we have
had, and grown so much as researchers and as
people. The laboratory staff would also tirelessly help
all

the

research

groups

refine

our

laboratory

techniques. Hwa Chong has invested intelligently into


grooming young student researchers, by having a
well-equipped Science Research Centre, and by
providing us with the opportunities for participating
in science fairs and international competitions. Hwa
Chong is the wind beneath our wings.

with a micropipette was vital in obtaining accurate


groundbreaking fields for my mentors and other
researchers. The process of the scientific method is a
meticulous and noble oneit seems almost too easy
to overlook the fact that mundane processes
contribute significantly to humanity.

Chiang Yan Li 13S7B


Research not only broadens our knowledge in
science and bolsters our confidence in this subject; it
trains our resilience as well. I did a research project at
the Weizmann Institute in Israel as part of the
International Summer Science Institute Programme. I
met

instrumental,

empirical,

and

theoretical

challenges at almost every step of the journey. Words

Tan Jiefeng 14S7B

of encouragement by my mentor kept me going and


provided me with the courage to present my project

Hwa Chong has invested


intelligently into grooming young
student researchers, by having a
well-equipped Science Research
Centre, and by providing us with the
opportunities for participating in
science fairs and international
competitions. Hwa Chong is the
wind beneath our wings.

in front of my peers. I hope that my newfound spirit


of resilience will act as the vital scaffold for my
eventual success. I believe that scientific research is
just like the inexplicable black-holeit is mysterious,
elusive, and we never know what we will pick up and
learn from it. But one thing is for certain: it is certainly
attractive!

Yi Kui Shuai 14S7B

15
VOYAGER 2014

students should possess is passion. Only if they are

Students Voices

I think that the one important quality research

GREEN SYNTHESIS OF
SILVER NANOPARTICLES
USING LALANG EXTRACT
1. INTRODUCTION
Lee Wei Hao, Joel 13S7C
Phua Yue Jun 13S6F
Tan Kwan Wei Kevin 13A16
Hwa Chong Institution

The application of nanoparticles, usually ranging from 1


to 100 nanometers (nm), is an emerging area of
nanoscience

and

nanotechnology.

Nanoparticles

exhibit completely new or improved properties based


on specific characteristics such as size, distribution and
morphology. Silver nanoparticles have been widely

Research Mentor
Mrs Sow-Peh Yoke Keow
Hwa Chong Institution

used in the field of catalysis, micro-electronics (Jain et


al, 2009), biological systems (Roy & Barik, 2010) and
medicine due to their antimicrobial and antibacterial
effects (Kamal et al, 2010).

Singapore Science and Engineering Fair,

Silver nanoparticles are synthesized by the reduction

Gold Award

of silver ions from Ag+ to Ag0 (Roy & Barik,

2010)

using reducing agents. The current methods of


synthesizing silver nanoparticles include chemical and

Green Synthesis Of Silver Nanoparticles Using Lalang Extract

VOYAGER 2014

32

National representative at the


Intel International Science and
Engineering Fair

photochemical reactions in reverse micelles, thermal


decomposition of silver compounds

in organic

solvents (Jain et al, 2009; Singh et al, 2010), radiation


chemical reduction and the chemical reduction of
silver ions in aqueous metal salt solutions (Forough &
Farhadi, 2010) with different external reducing agents
(Kamal et al, 2010). However, these chemical routes
can be extremely expensive and require the use of
toxic chemicals (Kamal et al, 2010). Hence there is a
need to explore alternative green chemistry routes,
where natural products are used for the synthesis of
silver nanoparticles.
The use of environmentally benign and organic
materials for the synthesis of silver nanoparticles
offers

several

benefits

of

eco-friendliness

and

compatibility for pharmaceutical and other biomedical


applications as they do not use toxic chemicals.
Chemical synthesis methods lead to the presence of
some toxic chemical adsorbed on the surface that may
have adverse effect in the medical applications (Jain et
al, 2009). Green synthesis, on the other hand, is
environment friendly, cost effective, easily scaled up
for large scale synthesis and does not require the use
of high pressure, energy, temperature and toxic
chemicals.

These

biologically

synthesized

silver

nanoparticles are also toxic against multi drug resistant


human pathogens (Kamal et al, 2010; Jain et al, 2009),
so they can be effectively used in the medical industry.
Chemical and physical methods are replaced with
green alternatives, using microorganisms, enzymes,

fungus and plants or plant extracts (Forough &

3 hours. The colloidal silver nanoparticles obtained

Farhadi,

was characterised using a UV-VIS spectrophotometer

2010)

for

the

biological

methods

of

synthesizing nanoparticles. Among these possible

(Shimadzu UV-1800). Triplicates were performed.

natural products, plants or plant extracts are preferred


over the microorganism routes as there is no need to
maintain

microbial

cultures,

which

can

be

2.2.3 Purification of silver nanoparticles

troublesome and arduous (Kamal et al, 2010).


The reaction mixture was centrifuged at 8000 rpm to
Thus this project aims to make use of organic

obtain the solid silver nanoparticles. Excess extract

extracts derived from lalang (Imperata cylindrical),

was decanted and the silver nanoparticles were

a weed, to synthesize silver nanoparticles. Lalang is

purified by repeated centrifugation at 8000 rpm with

a weed found commonly in tropical countries such

deionised water. The silver nanoparticles were dried

as Singapore and Malaysia. It is a novel weed to study

with a vacuum concentrator (Christ, RVC 2-25 CD).

as thus far, there have been no reports on the use of

The size of the solid silver nanoparticles was

this weed to synthesize silver nanoparticles. The silver

determined by a JEOL JSM-5200 Scanning Electron

nanoparticles synthesized were characterized using

Microscope (SEM).

UV-VIS Spectroscopy and its size determined by SEM


(Scanning electron microscope). Its anti-bacterial

2.2.4 Antibacterial Activity

property against Staphylococcus epidermidis (S.


Escherichia.

Coli

(E.coli)

and

The antibacterial properties of both the colloidal and

antifungal property against Aspergillus niger (A.niger)

solid

were also studied.

S.epidermidis, a common gram positive bacterium

silver

nanoparticles

were

studied

against

commonly found on human skin and mucosa and


E.coli, a common gram negative bacteria commonly

Hypothesis
The

solid

synthesized

and

colloidal

silver

lalang

extract

using

nanoparticles
will

exhibit

found in intestinal and digestive tracks of mammal


were tested.

antibacterial property against S. epidermidis and E.


coli and antifungal property against A. niger.

2.2.4.1 Colloidal silver nanoparticles

2. MATERIALS AND METHOD

Well diffusion test was carried out. The bacteria were


first grown in a nutrient broth for 24 hours/37C
before use. The overnight culture was swabbed onto

2.1 Materials

the Mueller-Hinton agar plates. 4 wells of 0.6cm in


Lalang

Road

diameter were created on each plate and they were

(Singapore). S.epidermidis and E.coli were obtained

was

obtained

from

Sembawang

labeled A (which contained the lalang colloidal silver

from Microbiologics while A.niger was obtained from

nanoparticles), B (which contained the lalang extract),

Carolina. Silver nitrate was obtained from GCE

+ve control (which contained 10% bleach solution)

Laboratory Chemicals.

and ve control (which contained sterile water). They


were all filled with 75l of respective extracts or

2.2 Methods

control. After 24 hours of incubation at 37C, the

2.2.1 Preparation of extracts

zones of inhibition around each plate were measured.

Lalang leaves were cut into small pieces. 25 g of

2.2.4.2 Solid silver nanoparticles

leaves were boiled in 100 ml of deionised water for 15


minutes. The mixture was centrifuged and filtered to

Total plate count method was carried out. The

obtain aqueous extract. The extract was stored at 4C.

bacteria were first grown in a nutrient broth for 24


hours/37C before use. The absorbance of the

2.2.2 Synthesis of silver nanoparticles

overnight culture was measured at 600nm. 100l of


the overnight culture was transferred to 10ml of

2.5 ml of lalang extract and 2.5 ml of deionised water

Nutrient Broth tubes. The tubes were labelled as

were added to 45 ml of 1mM aqueous silver nitrate. A

sample and control respectively. 0.01 g of solid silver

control was set up using 47.5ml of deionised water

nanoparticles was washed twice with 70% ethanol

with 2.5ml of lalang extract. The flasks were shaken for

followed by sterile water (2 times) and dried. The

33
VOYAGER 2014

and

Green Synthesis Of Silver Nanoparticles Using Lalang Extract

epidermidis)

1.6

broth and sonicated to obtain a homogeneous

1.4

mixture. To the sample tubes was added 1 ml of silver


nanoparticle suspension while to the control tubes
was added the same volume of nutrient broth. The
tubes were incubated at an incubator shaker for 24hr.
After 24 hr, the overnight broth culture was diluted
and 100l was spread onto the Nutrient Agar plates by
using

an

L-shaped

spreader.

The

plates

Absorbance

nanoparticles were then suspended in 5 ml of nutrient

1.2
1
0.8
0.6
0.4
0.2

were

incubated at 37C in an incubator for 24 hr. The

400

colonies were counted.


2.2.5

Antifungal

Assay

of

colloidal

500

600

700

800

Wavelength/nm

Figure 2. UV-VIS spectrum of colloidal lalang silver nanoparticles

silver

Figure 2 shows the UV-VIS spectrum of colloidal silver

nanoparticles

nanoparticles synthesized. A peak was observed at


silver

around 450 nm, which is typical of absorption for

nanoparticles was tested on the fungus, A. niger.

metallic silver nanoparticles, due to the Surface

Culture media was prepared using Potato Dextrose

Plasmon Resonance (SPR), confirming that silver

Agar. The agar was cooled at a 50C water-bath for at

nanoparticles have been successfully synthesized.

The

antifungal

property

of

the

colloidal

least 15 minutes and 5 ml of colloidal silver


nanoparticles were added and mixed. The agar was
poured onto petri dishes. A.niger was grown in Potato

Green Synthesis Of Silver Nanoparticles Using Lalang Extract

VOYAGER 2014

34

Dextrose Broth at room temperature for 24 hours. A


loopful of overnight culture was placed onto the

Figure 3. SEM image of

middle of the agar and the plate incubated at room

lalang silver nanoparticles

temperature for 5 days. The diameter of growth of the


fungus was measured on the 3rd and 5th day.

3. RESULTS AND DISCUSSION


Figure 3 shows the SEM image of the silver
nanoparticles synthesized by lalang extract. The size

3.1 Synthesis of silver nanoparticles

of silver nanoparticles ranges from 40 to 50nm. The


On shaking for three hours, the flask which contains

silver nanoparticles synthesized are spherical in

lalang extract and aqueous silver nitrate turned brown

shape. The size of the silver nanoparticles is smaller

(Figure 1), indicating the reduction of Ag(I) to Ag(0).

than that synthesized from weeds from India which

The formation of brown colouration is due to the

have a size ranging from 100-400 nm (Roy et al, 2010)

excitation of surface plasmon vibrations in the silver

and similar to that synthesized from aloe vera (Zhang

metal nanoparticles.

et al, 2010).

Silver

3.2

nitrate

Antibacterial

+
lalang
extract

Control
(without silver

of

colloidal

silver

(+) 10% Bleach solution

( - ) Sterilized water

(A) Colloidal lalang silver nano

(B) Lalang extract

nitrate)
Figure 1. Colour change of extract before and after
adding silver nitrate

Activity

nanoparticles

A
Figure 4. Zone of inhibition for
S.epidermidis

Figure 5. Zone of inhibition for


E.coli

Figures 4 and 5 show the presence of zones of

Figures 6 and Figure 7 show that the lalang solid silver

inhibition for both S.epidermidis and E.coli in the well

nanoparticles exhibit antibacterial properties against

filled

nanoparticles,

the two bacteria tested upon. For S.epidermidis,

indicating that the colloidal silver nanoparticles have

with

colloidal

lalang

silver

the lalang solid silver nanoparticles result in a drastic

antibacterial property against both bacteria. The

drop in the bacteria colonies, from 5 x 107 colonies in

inhibition zone for S.epidermidis was 1.25 cm while

control plate to 1 x 104 colonies in the plate containing

that for E.coli was 1.33 cm. In contrast, sterile water

silver nanoparticles, amounting to close to 100%

and lalang extract show no zone of inhibition. Lalang

reduction of bacteria colonies.

extract was tested to confirm that the antibacterial

reduction of bacteria colonies was around 48%, with

properties were in fact from the silver nanoparticles

the reduction of bacteria colonies from 84 x 106

synthesized from lalang extract and not from the

colonies in control plate to 43.5 x 106 colonies in the

lalang extract itself.

plate containing silver nanoparticles.

According to Marambio-Jones and Hoek (2010), silver


nanoparticles

exert

bactericidal

bind to the negatively charged cell membrane. Also


the silver nanoparticles could bind to thiol containing
by

permeability.

chelation,
This

in

which

turn

enhances

results

in

3.4 Antifungal Activity

activity

predominantly through the release of silver ions which

protein

For E.coli, the

Lalang
extract

Lalang
Silver nano

Control

cell

efflux

of

phosphate, leakage of cellular contents, disruption of


DNA replication and eventually cell death.
properties

of

solid

35

silver

nanoparticles
Figure 8. Photograph showing diameter of growth of A.niger in different
media on 3rd day

50
10

40

99.99%

30
20
10
0

control

Silver nano

Figure 6. Total plate count for S.epidermidis

Diameter of gowth/cm

Bacterial count x106/(CFU/ml)

60

7.9

5.6

6
4

2.5

2
0

7.7

2.5
0.5

No. of days
3rd Day
Control
Lalang silver nano

5th Day
Lalang extract

Figure 9. Diameter of growth of A.niger for a period of 5 days

Bacterial count x 106/ (CFU/ml)

From figure 8 and 9, it can be deduced that the lalang


100

84

silver nanoparticles are able to slow down the growth

90

of A.niger. As can be seen from Figure 8, there is little

80

48.2%

70
60

43.5

50

growth for the plate with the silver nanoparticles


added on the 3rd day, and this is in stark contrast with
the significant growth observed for the control plate
and the plate with lalang extract. Figure 9 shows that

40

on the 5th day, the diameter of growth for the plate

30

containing lalang silver nanoparticles was still smaller

20

than both the control plate and the plate containing

10

lalang extract.

control

Silver nano

Figure 7. Total plate count for E.coli

VOYAGER 2014

Antibacterial

Green Synthesis Of Silver Nanoparticles Using Lalang Extract

3.3

4. CONCLUSIONS AND RECOMMENDATIONS

International

Journal

of

Nanotechnology

and

Applications, 4, 95-101. Retrieved February 22, 2011 from


Silver nanoparticles can be synthesized from lalang

http://www.ripublication.com/ijna/ijnav4n2_4.pdf

extract. The presence of silver nanoparticles was


confirmed by UV-VIS spectrophotometry whereby a

Singh, A., Jain, D., Upadhyay, M.K., Khandelwal, N. &

peak was observed at 450 nm. The particle size of the

Verma,

silver nanoparticles synthesized range from 40 nm to

nanoparticles using argemone mexicana leaf extract

50nm. Silver nanoparticles synthesized from lalang

and evaluation of their anti microbial activities. Digest

extract were found to be effective against the bacteria

Journal

Staphylococcus epidermidis and Escherichia coli as

483-489.

well as against the fungus Aspergillus niger. As lalang

http://www.chalcogen.infim.ro/483_Singh.pdf

H.N.

of

(2010).

Green

Nanomaterials
Retrieved

synthesis

and

of

silver

Biostructures,

February

22,

2011

5,

from

is a common and fast growing weed in Singapore


which is unexploited, the use of it as a starting

Zhang, Y., Yang, D., Kong Y., Wang X., Pandoli, O., Gao.,

material for the large scale synthesis of silver

G. (2010). Synergetic antibacterial effects of silver

nanoparticles is viable, economical, eco- friendly and

nanoparticles@ aloe vera prepared via a green

sustainable.

method. Nano Biomed Eng, 267-274

This scope of this study can be widened by


investigating more types of bacteria. The lalang silver
nanoparticles can also be incorporated into everyday
products such as soap, cosmetics or socks and the
antibacterial properties of these products can be
evaluated.

Green Synthesis Of Silver Nanoparticles Using Lalang Extract

VOYAGER 2014

36

REFERENCES
Forough, Mehrdad & Farhadi, Khalil (2010). Biological
and green synthesis of silver nanoparticles. Turkish J.
Eng. Env. Sci., 34, 1-7. Retrieved February 22, 2011 from
http://journals.tubitak.gov.tr/engineering/issues/muh10-34-4/muh-34-4-7-1005-30.pdf
Jain, D., Diama, H. Kumar, Kachhwaha, S. & Kothari,
S.L.

(2009).

nanoparticles

Synthesis
using

of

plant-mediated

papaya

fruit

silver

extract

and

evaluation of their anti microbial activities. Digest


Journal

of

Nanomaterials

and

Biostructures,

4,

557-563. Retrieved February 22, 2011 from


http://www.chalcogen.infim.ro/557_Jain.pdf
Kamal, S.S.K., Sahoo, P.K., Vimala, J., Premkumar, M.,
Ram, S. & Durai, L. (2010). A Novel Green Chemical
Route for Synthesis of Silver Nanoparticles Using
Camellia Sinensis. Acta Chim. Slov., 57, 808-812.
Retrieved February 22, 2011 from
http://acta.chem-soc.si/57/57-4-808.pdf
Marambio-Jones, C., Hoek, E. M.V. A review of the
antibacterial effects of silver nanomaterials and
potential implications for human health and the
environment, J Nanopart Res. 2010;12:1531-1551
Roy, N. & Barik, A. (2010). Green Synthesis of Silver
nanoparticles from the Unexploited Weed Resources.

DETERMINING THE AMINO ACIDS


INVOLVED IN INHIBITION OF
PDE4B BY STRUCTURALLY-DIVERSE
COMPOUNDS
1 INTRODUCTION
Cai Anni 11S7B
Hwa Chong Institution
External Research Mentor
Professor Charles S. Hoffman
Boston College
Centre for Excellence in Education,
Research Science Institute, MIT, USA
Top prize for written report and oral
presentation

Phosphodiesterases (PDEs) are a class of related


enzymes

that

degrade

adenosine

35-cyclic

monophosphate (cAMP) by selectively catalyzing the


hydrolysis of the 3 cyclic phosphate bond of
cAMP[4].
Figure 1 : PDE Action On cAMP.
PDEs catalyze the hydrolysis of 3 cyclic phosphate
bond (shown in bond), thereby converting cAMP to
AMP.

Singapore Science and Engineering Fair,


Silver Award

transduction

pathways,

including

production

of

cortisol and metabolism of lipids. Spontaneous


degradation

of

cAMP

by

PDEs

allows

cAMP-mediated cellular transduction pathway to be


quickly

terminated

once

the

initial

chemical

messenger is no longer present, so that the cell may


respond to a fresh or repeated signal. This feedback
loop forms a basis for the tight regulation of
metabolic pathways in response to a fluctuating
environment [5].
Human PDEs are categorized into 11 families based on
substrate

specificity,

regulatory

properties,

and

homologies in primary sequence. The various PDEs


are encoded by 21 genes, which is transcribed into
more than 100 isoforms of the enzyme via alternative
splicing. In the official PDE nomenclature, the Arabic
numeral specifies the PDE family to which the enzyme
belongs, while the capitalized letter represents the
gene as reported in Genbank (NIH genetic sequence
database) [4]. Each family has unique substrate
selectivity: only the PDE4, PDE7, and PDE8 families
act specifically on cAMP; others act on both cGMP (a
second messenger similar to cAMP), or on cGMP and
cAMP [11].
It has been postulated that some PDEs function solely
to prevent

clouds of cyclic nucleotides from

spreading to inappropriate regions of the cell while

37
VOYAGER 2014

key second messenger involvedin many cellular signal

Determining The Amino Acids Involved In Inhibition Of


PDE4B By Structurally-diverse Compoundseditorial

PDEs are physiologically important because cAMP is a

others regulate local access to specific cAMP/cGMP

functions.

receptors.

unique

family-specific, but isoform-specific. As such, drugs

distributions of PDE isoforms, and the nature and

can be made to target specific disease pathways by

localization of these PDEs are likely to be a major

selectively

regulator of the local concentration of cAMP/cGMP in

involved. This can potentially minimize side-effects.

Different

cell

types

express

Some

pathways

inhibiting

the

are

not

specific

PDE

merely

isoform

the cell [4]. Tissue-specific expression of various PDEs


is

physiological

To this end, PDE inhibitor drugs have gained

functions. For example, PDE1B is implicated in allergic

intimately

mainstream acceptance. The erectile dys- function

reactions

for

drug, sildenafil (Viagra), is a selective PDE5 inhibitor

regulating IL-13 in PHA or anti-CD3/CD28-activated

which prevents cGMP from being degraded, thereby

human T-lymphocytes. PDE6, predominantly found in

prolonging the signal for smooth muscle relaxation.

rod cells of the photo-receptor layer of the retina,

The

functions in photo-transduction by reducing the

cavernosum optimizes physiological conditions for

steady-state concentration of cGMP in response to

penile erection [24]. Sildenafil is also used to treat

light stimulus, thereby leading to the closure of cyclic

pulmonary arterial hypertension [16] and altitude

nucleotide-gated

ion

sickness

channels.

consequent

as

linked
PDE1B

The

to

different

mRNA

is

channel

responsible

(CNG)
cell

cationic

membrane

consequent

[19]

vasodilation

based

on

the

in

the

same

corpus

mechanism

(vasodilation).

hyper-polarization sends an electric signal through


optical nerves to the brain, which is interpreted as a

Inspired by the pharmacological success of sildenafil,

visual stimulus [18].

drug companies are increasingly looking towards


archetypal PDE4 in- hibitor, was originally created by

38

recent years as it was found to influence a number of

Schering AG as a potential antidepressant and also

autoimmune diseases. One of the earliest to be

shows promise for treating inflammatory diseases

characterized, the PDE4 family was originally named

associated with PDE4B activity, ranging from chronic

for its selective resistance to rolipram (see Appendix

obstructive pulmonary disease to asthma, arthritis

for structure) and is characterized by cAMP substrate

and psoriasis. However, rolipram induces emetic

selectivity and low Km values. It comprises 4 isoforms

side-effects which are thought to be

which are sub-categorized into >20 variants that

non-specific inhibition of PDE4D in the brain [15]. Our

differ in their N-termini, where the PDE4 regulatory

current research is focused on understanding PDE4B

domains and phosphorylation sites are located [4].

inhibitor-specificity by determining the amino acids

Studies in knockout mice indicate that each individual

involved in inhibition of PDE4B through a novel

PDE4 gene plays a non-redundant regulatory role that

screening strategy outlined below.

Determining The Amino Acids Involved In Inhibition Of


PDE4B By Structurally-diverse Compoundseditorial

Much interest has been focused on the PDE4 family in

VOYAGER 2014

PDE4 inhibitors as drug candidates. Rolipram, the

caused

by

cannot be supplanted by the other 3 PDE4 transcripts


[12, 20], which agrees with the isoform-specific nature

MATERIALS AND METHODS

of many PDE-related diseases. One region of the


PDE4D gene is linked to heightened risk of ischemic

2.1

stroke[21], and absence of PDE4D3 led to progressive

PCR was carried out on plasmid pK63-N21 containing

cardiomyopathy

and accelerated heart failure in

the human PDE4B gene using the FailSafe PCR Kit

knockout mice possibly by increasing PKA-mediated

(Epicentre Biotechnologies, Madison, WI). Premix

phosphorylation of the cardiac ryanodine receptor

buffers A, B, E, F, K, L (Integrated DNA Technologies,

which rendered the ion channels leaky [14]. PDE4B,

Coralville, IA) containing 100mM Tris-HCl (pH 8.3),

the focus of this study, regulates inflammatory

100mM

responses by controlling the the production of TNF-

concentrations of MgCl2 (3-7mM) and PCR Enhancer

in both monocytes [12] and macrophages [13] in

(0-8X) were used with primers Cgs2-PCRmut-3 (3 to

response

open reading frame) 5AAA GTG TCC GAT GAG AAA

to

bacterial

lipopolysaccharide

(LPS)

stimulation.

Random mutagenesis via PCR

KCl,

400M

of

each

dNTP,

varying

AGC GTG 3 and Cgs2-PCRmut-5 (5 to ORF) 5 TCA


TAG CAT ACT TCT TCA CCA AGC 3. 30 cycles of PCR

The diversity and specificity of PDEs make them

were performed (each cycle consisted of 30s at 95C

excellent pharmacological targets. Unlike similarly

to denature DNA, 30s at 55C to anneal primers to the

ubiquitous cellular enzymes such as PKA that show no

template DNA, and 2.5 minutes at 72C for extension),

structural variance, each of the structurally different 11

after

PDE families is involved in different physiological

which

the

mutated

gene

products

were

2.2

Screening for inhibitor resistant PDE4B alleles

Schizosaccharomyces
(h leu1-32ade6-17216

pombe
was

cell culture was mixed with 10l of DMSO, BC27-5,

SP578

BC35 and BC58 respectively at 4M concentration in

transformed

DMSO. 5l of each mixture was plated on agar

following a gap-repair DMSO transformation protocol

containing YELleu medium and incubated at 30C

[2]. Yeast cells were cultured in Yeast Extract Liquid

for 2 days. Colonies were stained with iodine to

(YEL) medium [9] overnight, then subcultured into

identify colonies resistant to inhibitors.

90

cgs2-2)

strain

Edinburgh Minimal Medium (EMM)(MP Biomedicals,


Solon, OH) to target for 107 cells/ml. Cells were then

2.3 Phenotypic characterization of transformants

washed with 1X LiOAc/TE buffer ((10LiAc: 1M lithium

expressing can- didate mutant PDE4B alleles

acetate, adjusted to pH 7.5 with diluted acetic acid;

Selected colonies were transferred to liquid YELleu

10TE: 0.1M Tris-HCl, 0.01M EDTA, pH7.5) [2] and

medium and incubated at 37C for 1 day. The cell

brought to a final concentration of 2109cells/ml in

cultures were then diluted to a concentration of

this buffer. Either 200ng (first transformation) or

1107cells/ml. 10l of cell culture was mixed with 90l

50ng

StuI-linearized

of DMSO, and 40 M of compounds BC27-5, BC35 and

plasmid pKG3-N21 together with an excess of PCR

(second

transformation)

BC58, respectively to yield 100l of mixture, which is

product from each of the 6 PCR reactions and 20g

incubated at 30C for 1 day. Sample solutions from

boiled carrier DNA (Sigma Corporation, St. Louis, MO)

each cell culture were then inspected under the

were mixed with 100l of the prepared yeast cells. The

hemocytometer to determine the number of asci,

DNA-cell mixtures were incubated for 10 minutes at

zygotes and stationary phase cells. Mating efficiency

room

was calculated using the formula

temperature,

after

which

of

260l

of

40%

PEG/LiOAc/TE was added. The mixture was then

2(a + z) + s
2(a + z) + s + p

heat shocked for 5 minutes at 42C. Following


pelleting of cells (centrifugation at 16000 g for 5s),

where a = No. of asci, z = No. of zygotes, s = No. of

removal of supernatant and re-suspension in water,

spores and p = No. of stationary phase cells.

the cell solution was spread onto EMM plates


containing 3% glucose, adenine, histidine and uracil,

2.4

Plasmid rescue and DNA sequencing

but lacking leucine to select for transformed cells.

Plasmid rescue from yeast to E. coli was done using

Plates were incubated at 30C for five to seven days.

the Smash and Grab method [10]. The PDE4B2 alleles


on the plasmids were PCR-amplified as performed for

was

the initial gap-repair transformations using pre-mix

performed on colonies to identify candidates that

buffer G, purified on a Qiagen spin-column to remove

have successfully expressed the human PDE4B gene.

excess PCR primers and sent out to Eurofins MWG

Stained colonies were picked and transferred to a

Operon (Huntsville, Alabama) for DNA sequencing

384-well microtiter dish and incubated at 37C

to

using custom DNA primers Cgs2-PCRmut-5 (5 TCA

enrich for vegetative growth relative to mating and

TAG CAT ACT TCT TCA CCA AGC 3), PDE4B2-505F

meiosis.

(5TTC CTC CAT GAT GCG GTC TGT C 3), and

An

iodine-staining

assay

(see

Appendix)

PDE4B2-1165F (5CAC TGT GCA GAC CTG AGC AAC


Cells were then transferred to 50 l cultures of

3).

EMM-leucine medium in each of 4 plates: 1 control


plate containing 0.5% DMSO solution, the other 3

2.5

Structural Analysis of PDE4B

containing compounds 40 M BC27-5, BC35 and BC58

PDE4B structure (ID: 2QYL) was downloaded from

(see Appendix) dissolved in DMSO, respectively. The

RCSB PDB (http://www.pdb.org/pdb/home/home.do).

cultures from each of the compound-containing

The protein viewing software Chimera from UCSF

dishes were screened for ascus formation in the

(http://www.cgl.ucsf.edu/chimera/) was used to analyse

presence of inhibitor compounds, after which the

the mutant PDE4B enzyme. Residues flanking the

candidates were purified for plasmid rescue.

pocket were labeled, and the location of hydrophobic


and charged residues was determined relative to the

Purified single colonies were then transferred to liquid

pocket to assess its likelihood of being a ligand-binding

YELleu medium. Cell density was estimated using a

domain.

cytometer. Cell cultures were diluted to approximately


the same order of concentration (~ 107cells/ml). 10l of

39
VOYAGER 2014

mating efficiency =

addition of 43l of DMSO solution, the mixture was

Determining The Amino Acids Involved In Inhibition Of


PDE4B By Structurally-diverse Compoundseditorial

incubated for 30-60 minutes at 30C. Following

RESULTS

transformants yielded sparser colonies, allowing


colonies to be picked with greater accuracy.

3.1

Generating PDE4B mutant alleles via PCR

We generated PCR products using 6 different buffers

Frequency of stained colonies in each plate vary

designed to confer different rates and types of

among the different PCR buffers used for generating

mutations.

product

the mutant PDE4B allele. Based on visual inspection,

samples showed that the gene product was 2.1kb in

the percentage of colonies stained in each plate was

size, corresponding to that of PDE4B2. All bands were

estimated. Notwithstanding the limited precision of

distinct, which shows a large amount of product

this type of measurement, it is clear that buffers B and

formed.

E generated a much higher frequency of stained

Gel

electrophoresis

of

PCR

colonies than buffers K and L.

Figure 2: Gel Electrophoresis Image of PCR Products


a. PCR products generated using 6 different buffers were all 2.1kb in size,
corresponding to that of PDE4B. b. The plasmid cut with StuI is 21kb in
size, and smaller than the uncut circular plasmid.

The band corresponding to the cut plasmid travelled a

PCR buffer
A
B
E
F
K
L

Percentage of colonies stained


40%
60%
60%
20%
5%
10%

Table 1 : Frequency of stained colonies for different PCR buffers used

smaller distance than that of the uncut plasmid,


showing successful linearization by StuI.

Determining The Amino Acids Involved In Inhibition Of


PDE4B By Structurally-diverse Compoundseditorial

VOYAGER 2014

40

3.2

Transformation of S. pombe

Transformation of the first batch of S. pombe using


200ng of plasmid vector yielded a lawn of colonies
after 48 hours, making the process of picking
individual colonies difficult. To decrease colony
density, the second batch of cells were transformed
using only 50ng of plasmid vector. Number of
colonies was reduced 10-fold, resulting in increased

Figure 4 : Different frequency of stained colonies for different PCR


buffers
a. Left plate showed colonies expressing PCR product from buffer E, where
more than 60% of colonies were stained. b. Right plate showed PCR
products from buffer K, where less than 5% of colonies were stained.

colony separation.
3.3.2 Selecting candidates resistant to inhibitor
Mutants of interest have PDEs that function in the
presence of inhibitors and can regulate the glucose
sensing pathway. Such cells enter into a quiescent
state readily in glucose-deficient medium, presenting
with a short, shiny morphology, unlike cells that lack
PDE activity and fail to enter stationary phase [7].
Figure 3 : Comparison of colony density of Batch 1 and Batch 2

There should also be increased mating as reflected by

transformants

an abundance of zygotes, asci and spores, as cells

a. Left plate was transformed using 200ng of plasmid vector, and yielded
a dense mesh of small colonies which impeded later efforts to pick

lacking PDE activity display a severe mating defect

individual stained colonies. b. Right plate was transformed using 50ng of

[7].

plasmid vector, and yielded larger colonies with distinct separation.

3.3
3.3.1

Preliminary screening eliminated colonies showing

Screening for candidate mutants

little mating in the control plate. The remaining

Iodine Staining

The iodine staining assay identified 318 colonies from

candidates were screened for mating or quiescent

the first batch and 257 colonies from the second

cells in the inhibitor dishes. Out of 257 preliminary

batch expressing functional (though not necessarily

candidates, colonies we named A5, A6, A13, A17, A18,

wild-type) PDE genes. However, the first batch

A23, B11, B21, B29, D11, D19, D21, G4, G10, G13, G14, G17,

yielded

G18 were selected after the second screening.

dense

colonies

with

poor

separation,

compromising the accuracy with which stained


colonies

were

picked.

The

second

batch

of

Following

single-cell

purification

to

eliminate

contamination of strains, these 18 colonies underwent


iodine staining, which identified colonies A18, A23,
B21, D19, D21 as clearly inhibitor-resistant.

3.5 DNA Sequencing


To investigate the underlying molecular mechanism
for inhibition, sequencing was performed on D19.
Similar analyses are being performed for the
remaining colonies.
Sequencing analysis showed that the mutant PDE4B
recovered from colony D19 had an A to G transition
mutation at base pair 1403 of the open reading frame,
resulting in an aspartic acid to glycine substitution at
the position 468 in the primary sequence (NCBI
Reference Sequence: NM-001037339.1).
Figure 6
shows the location of the amino acid substitution in
the 3D conformation of PDE4B, obtained via
comparison of the primary sequence with the known
crystal structure of PDE4B.

Figure 5 : Result of iodine staining of 18 colonies selected after second


screening
DMSO plate generally showed dark staining. In the compound plates,

DISCUSSION

colonies A18, A23, B21, D19, D21 also showed significant staining,

compound plates.

However, D19 showed darker

staining in the BC35 plate than in the DMSO plate.


BC58 appears to be the most potent inhibitor in this
assay since, the staining was appreciably lighter for
most colonies in the BC58 plate as compared to the
DMSO plate. Inspection under the microscope also
showed that the dish containing BC58 contained the
fewest number of asci after an incubation period of
24h.
3.4
Phenotypic characterisation: quantitative
mating assay
Results of the quantitative mating assay performed on
colonies A18, A23, B21, D19, D21 is shown in Figure 6.

4.2 Mating assay


The results of the mating assay differed significantly
from those obtained via the iodine staining assay.
Random error may be a factor compromising both
assays, but each approach has its advantages and
drawbacks.
The mating assay here is done under more controlled
conditions with the yeast cell concentration fixed
across all colonies, as opposed to the solid medium
iodine staining assay, which concentrations of yeast
cells may vary 10-fold. However, the lack of distinct
morpho-logical difference between a spore and a
stationary phase cell means that counting of cells is
liable to human error.

Figure 6 : Results of mating assay


Colonies in DMSO plates generally showed the highest mating efficiency,
consistent with our prediction. A18 was an exception, as it should higher
mating efficiency in the BC35 plate than in the DMSO plate.

The results obtained from the mating assay differed


from those of the iodine staining assay. Colonies
generally showed highest mating in the DMSO plate,
with the exception of A18, which showed higher
mating in the BC35 plate. BC58 showed most potent
inhibition only for 2 of the 5 plates, while BC35 did not
show potent inhibition for most plates.

41
VOYAGER 2014

DMSO plates generally showed darker staining than

4.1 Generation of PDE4B mutants and


transformation
The different frequency of stained colonies for
different buffers may be correlated to the rate and
type of mutagenesis that the 6 buffers confer. High
rates of mutagenesis may lead to changes in the
essential amino acids of the catalytic domain, thus
rendering the PDE product non-functional. Future
research may be done to confirm whether this result is
reproducible to ascertain if the buffer delivers
consistent rates of mutation.

Determining The Amino Acids Involved In Inhibition Of


PDE4B By Structurally-diverse Compoundseditorial

suggesting that they express inhibitor-resistant mutant PDE4B alleles.

to interesting possibilities. Firstly, this D468G


mutation removes negative charge from the region,
altering the electrostatic interactions of that region
and possibly affecting binding affinities for charged
ligands. This may have caused BC35 to bind in a
different orientation, such that it becomes an
activator rather than an inhibitor of PDE4B.
Furthermore, glycine, by virtue of its small size, breaks
the local secondary structure, which may cause
drastic conformational changes in the entire pocket.
Much of the uncertainty regarding the precise effect
of this D468G mutation arises from our interpretation
of the PDE4B protein crystal structure. protein crystal
structures are inherently static, however, and cannot
Figure 7 : Model of PDE4B enzymatic structure
(Pink: Amino acid residues flanking the pocket. Orange: substituted
Gly-468. Yellow: hy- drophobic residues.) a. Pocket is flanked by 3 -helices

ological conditions. Nevertheless, all the evidence

and lined with hydrophobic residues, which provides a hydrophobic

discussed above suggest strongly that we have

environment for bound ligands. b. A close-up view of the pocket. A


disordered loop shields the entrance to the pocket, and may function as a

VOYAGER 2014
Determining The Amino Acids Involved In Inhibition Of
PDE4B By Structurally-diverse Compoundseditorial

identified a ligand binding pocket in PDE4B that

gate. The two fused rings of tryptophan protrudes and blocks entry to the

functions as an allosteric regulatory site, and is

pocket. c. Top view of the pocket. Tryptophan is located right in the center,

therefore a possible target for PDE4B inhibitor drugs.

and its bulkiness prevents access by binding ligands.

42

accurately depict enzyme interactions under physi-

The iodine staining assay is a more robust screening


test as high levels of mating must be present in a
colony for it to be stained. Results are representative
of the entire colony, unlike the mating assay which is
limited by the field of vision of the microscope.

Nevertheless, BC58 appears to be a potent inhibitor in


both the mating assay and iodine staining test, having
equal or sometimes greater effect than either BC27-5
or BC35. This contrasts with the results of in vitro
assays [6] which showed that BC58 is 10-fold less
potent than BC27-5 and BC35. It is possible that BC58
is more readily uptaken by the cells and thus show
greater effect in vivo.

transformed PDE- deficient S. pombe hosts with

4.3 Structural Analysis of PDE4B D468G Mutation


Analysis of PDE4B enzymatic structure reveals that
the substituted glycine lies at the periphery of a
pocket flanked by 3 - helices and partially covered by
a disordered loop. The presence of the disordered
loop in between highly ordered -helices meets the
requirements for a gated binding site, separate from
the substrate binding/catalytic site. This notion is
further supported by the presence of a large
tryptophan residue in the center of the pocket, which
likely regulates access to the putative allosteric
binding site by ligands. The core of the pocket is lined
with hydrophobic residues, providing a hydrophobic
environment for binding ligands. The periphery of the
pocket, however, contains several charged residues
that can form electrostatic interactions with
charged/polar groups of binding ligands.
The observation that the mutated PDE4B appears to
be activated rather than inhibited by BC35 gives rise

CONCLUSION

This study used a classic chemical genetics approach to


determine the amino acids involved in PDE4B inhibition.
We generated random mutant alleles of PDE4B,
mutated alleles and used the iodine staining assay to
screen

for

inhibitor-resistant

candidates.

DNA

sequencing of PDE4B2 mutant alleles recovered from


candidates

of

interest

revealed

an

amino

acid

substitution from aspartic acid to glycine near a pocket


directly opposite to the active site. Analysis of the
mutant PDE4B enzyme revealed strong evidence that
the pocket is a ligand-binding domain and functions as
an allosteric regulatory site. This information may be
useful in rational drug design when creating a PDE4B
specific inhibitor.

ACKNOWLEDGMENTS

I would like to express my profound gratitude to my


mentor, Professor Charles S. Hoffman of Boston College
for

his

invaluable

guidance

and

encouragement

throughout the course of this research project. In


addition, I would also like to thank Mr. Vinay Tripuraneni
for his insightful comments and suggestions for
improvement. Last but not least, I would like to thank
the Center for Excellence in Education for offering me
this opportunity, MIT for hosting this program, and the
Ministry of Education, Singapore for its generous
funding.

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[18]

Interaction between ran1+ protein kinase and cAMP

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Hoffman. (2008). Development of a fission yeast-based

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VOYAGER 2014

as Treatment for Pulmonary Arterial Hypertension.

edition. Redwood City, Calif.: Benjamin Cummings.

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S.G.

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(1979).

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A.3 Results of mating assay

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phosphodiesterase adn resolution of multiple molecular


forms of the enzyme. Adv Cyclic Nucleotide

Res.

10:69-92
[23] R.X. Xu, W.J. Rocque, M.H. Lambert, W.D. Holmes,
M.A. Luther, W.J. Rocque, M.V. Milburn, Y. Zhao, H. Ke, et
al. (2000). Atomic structure of PDE4: insights into
phosphodiesterase mechanism and specificity. Science
(Wash DC). 288:1822-1825
[24] D.J. Webb, S. Freestone, M.J. Allen, G.J. Muirhead.
(1999). Sildenafil citrate and bloodpressure-lowering

Table 2: Mating efficiencies of colonies in different compound plates

drugs: results of drug interaction studies with an

A.4

PDE4B enzymatic structure

organic nitrate and a calcium antagonist. Am. J. Cardiol.


83 (5A): 21C-28C

A APPENDIX
A.1 Inhibitor compounds

Determining The Amino Acids Involved In Inhibition Of


PDE4B By Structurally-diverse Compoundseditorial

VOYAGER 2014

44

Figure 8 : Known Inhibitors of Rolipram


Rolipram is the archetypal PDE4 inhibitor, for which the family was initially
named. BC27-5, BC35 and BC58 were previously identified by the
Hoffman lab in an in vitro assay as potent PDE4 inhibitors.

A.2

Iodine staining assay

This assay is used to identify colonies expressing a


functional PDE. Small iodine crystals are spread on a
plate. Each colony plate is inverted and placed on top of
the iodine plate, covering the latter. Following 2 minutes
of iodine vapor staining, the colony plate is removed.
Colonies with high rates of mating are stained.
The basis for this assay lies in a cAMP-mediated
glucose sensing pathway that controls mating. In a
glucose-deficient

environment,

no

new

cAMP

is

Figure 9: Additional PDE4B enzymatic structures


Different views of the PDE4B enzyme. Pink: Residues flanking the pocket
are colored pink. Orange: Substituted Gly-468. Different panels show the
ribbon structure (left), ribbon structure (right).

synthesized, so cells with functional PDE will have low


cAMP levels, which induces mating. Cells undergoing
mating have high levels of glycogen, which stains with
iodine. In contrast, dysfunctional PDE cannot degrade
cAMP, and cellular cAMP levels is persistently high in
the absence of glucose, so no mating occurs.

Figure 10: Additional PDE4B enzymatic structures


Yellow: Hydrophobic residues Different panels show the ribbon structure
(left), ribbon structure and stick structure (center) and stick structure
only (right).

Fall Detection System based on


Kinect Sensor using Novel Detection
and Posture Recognition Algorithm
1 INTRODUCTION
Lee Choon Kiat 12S68
Hwa Chong Institution

The progressive aging of population has become a


major social challenge for countries around the world.
As more elderly begin living with health problems and

External Research Mentor


Lee Vwen Yen
Institute for Infocomm Research, Agency
for Science, Technology and Research

are home-alone, they require increasing assistive


support in daily activities. For the elderly, involuntary
falls are frequent. A visit to Lee Ah Mooi Old Folks
Home showed that among the 100 elderly residing in
the home, about 3 fall cases occur in every quarter of
the year, while annual statistics show that one in every
three adults age 65 and older in the USA have recently
suffered a fall [1]. Falls cause a loss in quality of life for
the fallen elderly and can be more dangerous due to
the fact that the victim can easily lose consciousness
and thus become unable to seek help if they are home
alone, which is detrimental to their long-term health if
order to avoid this scenario, fall detection systems
that

are

capable

of

identifying

and

notifying

caregivers when the elderly fall are very much


essential in the bid to provide assistive services to the
elderly.
Current fall detection systems have fall detection rates
of 70% to 80% [3], which might be unconvincing to
some

consumers

when

making

decisions

on

purchasing such systems. There are privacy concerns


when vision-based fall detection systems are used,
although there are situations where it is acceptable
for visual sensors to be deployed, where the risk of
falls being undetected heavily outweighs the loss of
privacy.

Furthermore,

detection

systems

current

with

vision-based

heavy

dependence

fall
on

complex video processing also limit the systems


effectiveness when deployed in real time [3].
We aim to develop and test a fall detection system
that utilizes the Microsoft Kinect sensor [4] and our
novel

fall

detection

algorithm

to

determine

automatically if a fall has occurred. The Kinect sensor


has an RGB camera and depth sensor which provides
full-body 3D motion capture and facial recognition
[5]. When compared to other commercially available
depth sensors, Kinects low cost makes it a good
choice for developing inexpensive, vision-based fall
detection systems that appeals to the consumer
market and industry. Relative lack of research into

45
VOYAGER 2014

the accident is serious and undetected [2]. Thus, in

Fall Detection System based on Kinect Sensor using Novel


Detection and Posture Recognition Algorithm

This research was presented at the 2013


International Conference On Smart homes
and health Telematics (ICOST)

Kinect-based fall detection systems also provides us


with

much

room

in

conducting

research

and

development within this field.


We propose our fall detection system which is
optimized using current preliminary datasets and
when used in conjunction with our novel posture
recognition

algorithm,

help

us

to

achieve

significantly higher specificity rate of fall detection, up


to 90% accuracy rate in an actual test with a real
human subject.

2 FALL DETECTION SYSTEM


In this project, we aim to create a fall detection system

Fig. 2: Block diagram of our fall detection system

(Fig. 1 & 2) for use in confined areas like the bedrooms


or washrooms in single room apartments. This

3 FALL DETECTION ALGORITHM


3.1 Control Algorithm

who normally live in such apartments. In order to

Many other current fall detection algorithms are

avoid privacy concerns over the use of our system in

based on the concept that during a fall, the person will

these settings [6], the fall detection system only

experience a large initial downward velocity, ending

46

utilises the Kinects depth camera and displays only a

with the persons centre of mass on or near the floor

stickman model of the user, and eliminates the usage

[8]. Though it has been successfully used by other fall

of any actual footage, thus making it suitable to be

detection systems, this algorithm has a few flaws,

used in such regions. In addition, our system is also

chiefly among which is its sub-optimal specificity and

able to operate in situations with little or no light [7],

sensitivity, which still have not been addressed by

therefore making it possible for the Kinect to track

current research. We adopted this algorithm as a

human activity both in the day and night, something

starting point for our own fall detection algorithm

that conventional stereo video camera systems are

with modifications to address the flaws of this main

unable to do. This makes it especially useful as most

algorithm. We then implemented this fall detection

falls by the elderly happen at night when visibility is

algorithm

poor and this also improves the effectiveness of our

effectiveness of our own novel algorithm

Fall Detection System based on Kinect Sensor using Novel


Detection and Posture Recognition Algorithm

towards the detection of falls for home-alone elderly

VOYAGER 2014

constraint is in place as the project is targeted

as

control

so

as

to

assess

the

fall detection system.


3.2 Our Novel Fall Detection Algorithm
The Kinect depth camera captures a depth image of

The control fall detection algorithm described earlier

the surroundings. The raw data streams are then

was then applied to a preliminary dataset of 10,479

processed by the Microsoft Kinect SDK to return a

frames of skeletal data from the Kinect comprising 34

pipeline of information that provides our fall detection

fall and non-fall events, with the results used to

system with skeletal data of the user. The skeletal data

optimise our enhanced fall detection algorithm. The

of the user is then transmitted to our fall detection

results of this experiment are described in greater

software and subsequently passed to our algorithm as

detail in the Results section (section 4) of this paper.

described in section 3 for further processing. If a fall is

Our enhanced fall detection algorithm is based on a

detected, we would relay a signal to an alarm system

modular system with skeletal data received from the

to sound an alert.

Kinect sensor and processed by two functions

Fig. 1: Setup of our fall detection


system

(function 1 and 2). If both functions return a positive


result, a fall is tentatively indicated. After that, our
novel postural recognition algorithm is then applied to
reduce the amount of false positives returned by our
fall detection algorithm, resulting in a fall detection
algorithm that manages to achieve relatively high
sensitivity and specificity.

3.2.1 Function 1 (Checking position of users centre


of mass)
The real world coordinates of the persons hip centre

ds y (t )
dt

(roughly representative of the persons centre of

v(t )

(1)

sh (t )

sx (t ) 2 sz (t ) 2
dsh (t )
dt

vh (t )

(2)

mass), are obtained by the fall detection algorithm


after processing of the raw data from the Kinect by
our fall detection system. Our algorithm checks if the
users hip centre is within a certain threshold distance
from the floor returns a positive result if found to be
true.
4.2.2 Function 2 (Checking velocity of users centre
of mass)
Many current algorithms that make use of the centre
of mass are unable to accurately detect slow falls.
This is because the measured vertical velocity in slow
falls tends to be relatively lower than that measured
into its horizontal and vertical components

not detected. Figure 3 shows the different velocities

The different velocities of the users hip centre are

of the users hip centre joint where the vertical

given different scores for the vertical and horizontal

velocity of the slow fall did not exceed the threshold,

velocities as shown below in Table 1. Our fall

resulting in no falls being detected.

detection algorithm checks if the combined scores


for both the vertical and horizontal component
exceed our pre-set overall threshold score. As the
vertical velocity of the users hip centre was deemed
to be an important factor in determining a fall, we
weighted the algorithm accordingly. The overall
threshold score was then optimized such that a large
vertical velocity typically indicative of a fall would be
able to trigger an alert, but a large horizontal velocity
which could signify a subject moving at fast speeds
would not trigger a fall alert, thus enhancing the
specificity of our fall detection algorithm.
The table of scores shown in Table 1 was optimized
manually by studying the preliminary dataset of falls
and tweaking the parameters until a maximum fall

Fig. 3: Graph showing different velocities of the users hip centre joint for

detection rate was obtained.

both the slow fall (solid lines) and the vertical fall (dotted line).
Table 1 Table showing how the scoring system was applied for vertical

Our algorithm separates the velocity of the users hip


centre into two components. The first component is
the vertical velocity of the hip centre (Equation 1),
while the second component is the velocity of the hip

velocities

Vertical velocity
Score of users hip centre, Remarks
VV / m s-1
VV > -0.9

-0.9> VV > -1.4

Representative of a small vertical velocity that


is typical of a slow horizontal fall, but not low
enough to be triggered by everyday activities

-1.4> VV > -1.6

Representative of a moderate vertical velocity


typical of a fast horizontal fall

VV < -1.6

centre with respect to the vector sum of the


displacement of the hip centre in both the x and z
directions, thus giving us the horizontal component
of the hip centres velocity, as illustrated in Fig. 4 and
(Equation 2).

Representative of negligible vertical velocity


typical of everyday activities

Representative of a large vertical velocity


typical of a vertical fall

47
VOYAGER 2014

values are normally not exceeded and the slow fall is

Fig. 4: Diagram showing how the actual overall velocity of a joint is split

Fall Detection System based on Kinect Sensor using Novel


Detection and Posture Recognition Algorithm

during a normal fall. Therefore pre-set threshold

This new method of fall detection provides two main

In order for our postural recognition system to

benefits:
1. It is more sensitive to falls in any direction than

correctly identify these postures, two main defining

conventional fall detection systems as both the

features were identified in order to separate fallen


postures from other possible postures.

horizontal and vertical components of velocity of


the users hip centre are taken into account,

1. Feature 1: Users ankles have to be below the users

reducing the impact of the reduced vertical

hip centre.

velocity of the persons hip centre during a

!$!')!/1.! ,+/01.!/#%*/0,+/01.!/

horizontal fall.

and 3 listed in Table 2 as we observed from skeletal


data that all three postures exhibit this feature.

2. It is less prone to false positives generated by large


horizontal

movements

by

the

user

than

2. Feature 2: One of the users legs is either folded

conventional fall detection systems that determine

below his body or one knee being significantly higher

falls based on the total velocity of the users

than the other.

centroid.

!$!')!/1.! ,+/01.!/#%*/0,+/01.!%*
Table 2 for users that bend down to perform actions
such as wearing of shoes or tying of shoelaces.

4.2.3 Postural Recognition Algorithm


We observed that the starting adopted control

Fall Detection System based on Kinect Sensor using Novel


Detection and Posture Recognition Algorithm

VOYAGER 2014

48

algorithm has a high rate of false positives. Due to the

These two features were refined and checked against

multiple similarities between falls that are classified as

previously obtained skeletal data of postures to

false positive and actual real falls, most current fall

ensure their specificity.

detection

systems

between

these

have
two

difficulty

differentiating

classifications

[9].

Table 2: List of false positive postures considered

We

demonstrate this similarity with results in Figure 5


using our own data, where the monitored subjects

Posture

Classified
Fall /
Non-Fall using rule no.
Non-fall
1

Sitting on the floor


both legs folded behind

Kneeling on the floor

Non-fall

the false positives from actual real falls by checking

Squatting

Non-fall

the users posture to determine the likelihood that the

Non-fall
4 Bending down
(to wear shoes or tie shoelaces)

velocity is similar for an actual fall and an action of just


bending down. To resolve this problem, we developed
a novel postural recognition algorithm that separates

user has been involved in a fall when previous


functions 1 and 2 return positive results.

4 RESULTS
In order to test the actual effectiveness of our system,
we conducted a trial experiment that was simulated
as closely to the actual conditions where our fall
detection system would be deployed. We want to test
the robustness of our fall detection system by
conducting tests within a setting that is previously
unknown to the system. A nursing ward was recreated
within our laboratory to provide a realistic setting that
is similar to wards found in the previously mentioned
Ah Mooi Old Folks Home. A hospital bed was placed
in the corner of our ward, with PVC pipes representing
the walls of the ward and the Kinect sensor was
deployed at the top corner of our ward.
Fig. 5: Graph showing different velocities of the users hip centre joint

The nurses and caregivers at Ah Mooi Old Folks Home

for bending down (solid lines) and the an actual fall (dotted line).

have provided us with domain knowledge regarding


elderly falls and the common locations of such falls.
There are mainly three kinds of falls that frequently
occur within the Home, mainly falling after getting out

of the bed, falling after getting out of the wheelchair,

Our fall detection system is capable of processing

and a normal fall in any direction in open space. These

incoming sensor information and displaying of results

different kinds of falls were then simulated by our

concurrently without a loss in processing speed or

volunteers to replicate the kind of conditions that our

performance degradation. By connecting an SMS

fall detection system will face when deployed at the

gateway or alert module to the system, we can detect

Home. Our test volunteer was given the freedom and

falls within half a second of the fall and subsequently

not restricted in the way that he could fall. A video

alert the caregivers. The system automatically starts

camera was placed to capture his movements and

when someone enters the field of vision of the Kinect

also to provide ground truth which we could then

sensor. By producing a stickman display (Fig. 6.2) for

compare against our logged results. Fig. 6.1 shows our

display purposes which users can choose to hide, we

experimental setup, while Fig. 6.2 shows a screenshot

also managed to preserve the users privacy.

of our fall detection system user interface during the


Initial results show that most falls that include falling

experiment.

from the bed or in open spaces were accurately


identified by our fall detection system. Multiple other
kinds of non-fall events were also accurately detected
by

our

fall

detection

algorithm

and

correctly

classified, mainly due to the assistance from our novel


posture

recognition

algorithm

that

points

out

abnormalities in the data obtained through the


sensors. From our results, this recognition algorithm is
able to point out 62.5% of all non-fall events and helps

of our system and help in preventing more false


positives from being classified, thus demonstrating
the usefulness of our posture recognition algorithm in
recognizing and differentiating falls. Our system is
able to achieve an overall specificity rate of up to 90%
and our algorithms helped improve the fall detection
rate such that we could detect all falls (100%) in
Fig. 6.1 Picture showing experimental setup

certain situations, where conditions are optimal. This


includes situations where more sensor information
allows us to perform inference in order to provide a
more accurate assessment.
However, further analysis also showed that there were
occasions where the system was not able to classify
actual falls. For example, a user might fall off the chair
and the fall is undetected, due to the fall taking place
in a low position, and reduced time required by the
system to measure a significant increase in users
velocity while he is falling, hence resulting in the
system not being able to correctly detect the fall. We
have detailed data and information regarding this kind
of misclassification and also have a solution for this
problem and are currently working on correcting this
issue so as to ensure that it does not occur again in
subsequent modifications.

Fig. 6.2 Screenshot of fall detection system user interface running on


laptop

49
VOYAGER 2014

other non-fall events are then detected by other parts

Fall Detection System based on Kinect Sensor using Novel


Detection and Posture Recognition Algorithm

us in classification of the majority of actual falls. The

5 CONCLUSIONS

Manuscript submitted for publication

Our fall detection system is able to detect and identify

[9] Gadre, K. R. (2012). Fall Detection System Using

falls using the Kinect sensor within confined spaces at


a relatively high specificity rate when compared to
usage of control algorithms. The system also uses our
novel posture recognition algorithm to enhance our
detection

rate

deployment.

and

ensures

Overall,

with

robustness

during

sufficient

sensor

information while maintaining users privacy, our fall


detection system is able to automatically recognise
falls and notify healthcare services to provide timely
assistance to elderly who have fallen.

REFERENCES
[1] George, F. F. (2000). Falls in the elderly. Am Fam
Physician, 61(7), (pp. 2159-2168.)
[2] Wild, D., Nayak, U. S., & Isaacs, B. (1981). How
dangerous are falls in old people at home?. British
medical journal (Clinical research ed.), 282(6260), 266

Fall Detection System based on Kinect Sensor using Novel


Detection and Posture Recognition Algorithm

VOYAGER 2014

50
[3] Perry, J. T., Kellog, S., Vaidya, S. M., Youn, J. H., Ali,
H., & Sharif, H. (2009, December). Survey and
evaluation of real-time fall detection approaches. In
High-Capacity
Technologies

Optical

Networks

(HONET),

2009

and

6th

Enabling

International

Symposium on (pp. 158-164). IEEE.


[4] C. Rougier, J. Meunier, A. St-Arnaud and J.
Rousseau, (2007). Fall Detection from Human Shape
and Motion History using Video Surveillance, 21st IEEE
International Conference on Advanced Information
Networking

and

Applications

Workshops

(AINAW'07),875-880.
[5]

Totilo,

Stephen

(January

7,

2010).

"Natal

Recognizes 31 Body Parts, Uses Tenth of Xbox 360


"Computing Resources"". Kotaku, Gawker Media.
Retrieved November 25, 2010.
[6] Yongli, G., Yin, O. S., & Han, P. Y. State of the Art: A
Study on Fall Detection.
[7] Stone, E. E., & Skubic, M. (2011, May). Evaluation of
an inexpensive depth camera for passive in-home fall
risk assessment. In Pervasive Computing Technologies
for

Healthcare

(Pervasive

Health),

2011

5th

International Conference on (pp. 71-77). IEEE.


[8] Kawatsul, C., Lil, J., Chung CJ. (2012) Development
of a Fall Detection System with Microsoft Kinect,

Low

Cost

Computing

University of Missouri).

(Doctoral

dissertation,

INVESTIGATION INTO THE RELATIONSHIP


BETWEEN VIOLIN STRING TIMBRE AND
ITS HARMONIC STRUCTURE
INTRODUCTION AND HYPOTHESIS

Singapore Science and Engineering Fair,


Gold Award

Hypothesis and Assumption


We hypothesize that sound waves produced by violin
open E strings with different timbres can be matched
to different, distinctive patterns of harmonic structure.
Assumptions in this study include: sound samples
played by professional players are stable period
waves; bow stokes applied for sound samples are the
same throughout the recording process; strings in
each timbre group are most representative of that
timbre.

BACKGROUND THEORIES
Violin and Timbre
The violin is a bowed string instrument. Drawing a
bow across the string correctly causes the violin body
to resonate due to a complex system of different
couplings. (Crosmer, 2011) Its timbre or tone color can
be affected by factors including the string
composition, the instrument used, vibrations from
other strings, bow speed, bow pressure and sounding
point where the bow intersects the string. (Charles,
2010) Physically, change in technical factors will
affect the amplitude of individual component
frequencies in a complex tone, resulting in a distinct
shaped frequency spectrum that characterizes a
specific timbre.(Crosmer, 2011) As sound waves
entering human ears are interpreted by the brain,
people usually use words like rich nasal or
brilliant to describe their feeling. Different English

51
VOYAGER 2014

External Research Mentor


Professor Chan Aik Hui, Phil
Mr Tan Peng Kian
National University of Singapore

Introduction
Music is part of all cultures and traditions. Amidst the
evolution from early church hymns to modern
electronic music, people have worked towards a
deeper
understanding
of
musical
sound
characteristics so as to reproduce pleasant music.
With todays advanced signal processing technology,
much work has been done on the physical aspects
that distinguish different musical instruments.
However, the complete timbre space of a single string
instrument with distinctive rich sound the violin - is
not as well understood. Violin timbre depends on
many factors including the wood material of the violin,
bow strokes, etc. This research focuses on the
variance of violin timbre due to different violin strings
and investigates the relationship between violin
timbre descriptors and the harmonic content of
various open E string samples.

Investigation Into The Relationship Between Violin String Timbre And


Its Harmonic Structure

Yang Liuyi 12S7D


Wang Jia 12S7F
Hwa Chong Institution

descriptors for violin are mapped systematically by


the following coordinate constructed by Acoustical
Society of America (Friz, et al., 2011) (see figure 1). The
area under the dotted line represents descriptions for
pleasant violin sound, which can be further split into
3 regions in the order of harshness: rich/warm,
bright/open and penetrating/projecting. This way of
classifying string timbre will be used for later
discussion of experiment results.

At a point of discontinuity, the Fourier series


converges to the average of the two function values at
either side of the discontinuity.
In the experiment, data was Fourier analyzed by
oscilloscope software. Nonetheless, understanding
the mathematical foundation enabled us to use the
software with greater confidence.

HARMONICS
In the periodic waves produced by musical
instruments, the periodic frequencies of all
component waves are called harmonics. The first
harmonic is also called the fundamental (f0), while the
frequencies of higher harmonics (overtones fi) are
multiples of that of fundamental. (Oldham,et al.,2011)
In the trigonometric series of equations [1],

fundamental while

is

the

function

of
is

the first overtone and so on.

SPECTRAL PLOT

Investigation Into The Relationship Between Violin String Timbre And


Its Harmonic Structure

VOYAGER 2014

52

Figure 1: Acoustical Society of Americas A-MDS map of timbre descriptors


for overall violin sound quality

Fourier Analysis
Joseph Fourier showed that any complex wave can be
described as the sum of a series of simple sinusoidal
waves called partials (Campbell, 1987). (See figure 2)

When a wave is Fourier analyzed, the results can be


shown in a spectral plot. The wave spectrum is a
2-dimensional
representation
manifesting
the
amplitudes of sinusoidal waves of different frequencies
that comprise the wave. Different spikes of energy in the
spectral plot are partials. (See figure 3)

Fourier Analysis

Figure 2: Fourier analysis can break a complex wave which is the


ummation of two sine waves of different amplitude, frequencies and
phases into their components

If f(t) is a periodic function with period 2L and


fundamental angular frequency

, the

Fourier-series representation of f(t) is given by


n

[1]

The coefficients (called the Fourier coefficients of f(t)


a0, an and bn are to be evaluated and given by the
expressions
;

an

1
L

f (t ) cos n tdt

for n=1,2,;

for n=1,2,;

Figure 3: Spectral plot of the sound wave of Pirast ro Olive Gold E

NOISE CONTROL AND MINIMIZATION


Due to the high sensitivity of the microphone, it is
assumed that all sound waves are recorded. Hence, it is
pertinent to discuss sources of noise that cause errors in
recording before proceeding into data analysis. Below
are three main sources of noise involved and respective
ways to minimize their influence:
a) Initial noise is caused by the condition of recording
room, including the noise of air conditioner, human
breathing and noise outside the room. It is minimized by
using a music recording room with air conditioner off
when outside the room the noise is minimal. (See figure 5)
b) Cable noise is due to the radio waves picked up by
the conducting wire. We shortened the length of the
cable by winding up the wire.
c) Machine noise is due to the running of laptop hard
drive and oscilloscope, as well as possible radio waves
picked up by the hardware. A silent audio was played to
record machine noise A0 /dB, and amplitude of samples
without machine noise Ai/dB is calculated by the
following equation:

Ai '

Figure 4: Material used for recording of E string sound samples

Recording
The violin and microphone were placed 50 cm apart
and fixed at the same position in the room. Each E string
was tuned to 663.0 Hz and then bowed by a team
member who is an experienced violinist for recording.
Each sample includes 3 consecutive legato bows of 2
seconds as 3 trials. The bow is controlled at 2 cm away
from the bridge and maintained at a relatively constant
bow angle and speed so as to prevent variance in
timbre caused by bow strokes. (Chen, 2007) While
playing, the other 3 violin strings A, D, G were muted to

1
lg 10 Ai 10 A0
10

Figure 5: The negligible initial noise in recording

53
VOYAGER 2014

Materials
To investigate the sound quality of violin strings, we
used ten violin E strings with different composition and
the identical diameter 0.265 mm. As the sounding
frequency of a string tends to fall after it is first tuned steel stabilizes within a few minutes, synthetic strings
about 8 hours and gut strings 48 hours (Fletcher et al.,
1998), 2 days were allowed for each string to settle in a
violin from the China Conservatory of Music.
Throughout the process all materials were kept in the
same room under 25C and humidity from 30% - 50%.
Recording of open E strings took place in a music
recording room with laminated glass window and
acoustic panels to prevent excessive reverberation;
hence the sound can disperse approximately equally
throughout the room without minimal distortion.
(Campbell et al., 1987) A Neumann U87AI condenser
microphone was used due to its excellent response for
frontal sound incidence and high fidelity. Software for
sound analysis includes LeCroyWaveRunner Xi-A /
MXi-A Oscilloscope, Microsoft Excel and SciDavis, which
enable us to plot the waveform of samples and Fourier
analyze the resultant data.

prevent their influence on the timbre of E string. The


samples were saved in audio files, which were later run
through oscilloscope and computer software for
Fourier analysis.

Investigation Into The Relationship Between Violin String Timbre And


Its Harmonic Structure

METHODOLOGY

Penetrating
Projecting

1k

4k

7k

10k

13k

16k

19k

21k Freq/Hz

Figure 9: Comparison of strings with penetrating tone colour and good


projection

Discussion of Results
After grouping, patterns are observed from 3 aspects:
the strength of harmonics, the fluctuation of partials
and , the level of scattering of the amplitude data.
Figure 6: The waveform and FFT diagram of machine noise

RESULTS AND DISCUSSION

Investigation Into The Relationship Between Violin String Timbre And


Its Harmonic Structure

VOYAGER 2014

54

Interpretation of Data
After removing the effects of machine noise, we
tabulated the amplitude of various partials against
frequency for all 3 trials of each string. The average
data of 3 trials was taken to plot graphs for
comparison.
The 10 strings with varying timbres are grouped in 3
categories based on timbre descriptors provided by
string producers: rich/warm, bright/open and
projection/penetrating. The spectral diagrams of 3
groups are provided below.

Samples with a rich and warm tone colour have


relatively even distribution of energy across the
spectrum with constant fluctuations throughout the
spectrum. (See figure 7). The level scattering of
amplitude of partials is the greatest among 3 groups,
with standard deviation w from 16.5 to 16.8 (see Table
1). The brilliant tone of bright/open and
penetrating/projecting groups is likely to be
associated with high energy distributed on high
harmonics with sustained strength, which can be seen
from the plateau of spectral diagrams from 10,000
Hz to 19,000 Hz with amplitude between -50 dB to
-60 dB (see figure 8, 9). The harsh penetrating
property can be attributed to greater scattering of
amplitude as well as more fluctuations (see figure 8,
9), as P bears a slightly higher range than B.
Table 1: The standard deviation of amplitude data of different strings

Rich
Warm

1k

4k

7k

10k

13k

16k

19k

21k Freq/Hz

Figure 7: Comparison of strings with rich, warm tone colour

Bright
Open

1k

4k

7k

10k

13k

16k

19k

21k Freq/Hz

Figure 8: Comparison of strings with bright and open tone colour

Rich/Warm

<EvahP Au>

<Pir Oliv>

<D'Light>

Std Dv w

16.765

16.673

16.549

Range
16.5 16.8

Bright/Open <Chromcor> <D'heavy>

<Infd Tn>

<Lenzner>

Std Dv B

14.318

15.459

15.281

14.951

Penetrating

<Vision>

<EP Ag>

<Infd Pt>

Std Dv P

16.208

16.295

16.363

14.3 15.5

16.3 16.4

CONCLUSION AND EXTENSION


Our study has proven the hypothesis to be true, and at
the same time provided a deeper insight into patterns
of different string tone colours.
This research provides a method to build a database
of different instrumental timbre. Findings in this study,
although limited to a rather small scale, can be used in
computer generated music to create more variance in
instrumental timbre. This allows electronic music
producers to add life to music produced by
machines, which is of great practical use in
entertainment industry such as synthesis of pop music
and movie making.

We are highly indebted to our mentors Professor Chan


Aik Hui, Phil and Mr Tan Peng Kian for their inspiring
guidance and supervision throughout the project. We
would also like to express our gratitude towards our
Supervising Tutor, Dr Lim Jit Ning for his constant help
and guidance. Special thanks also go to Dr Tan
Wee-Hsin and Ms Lillian Wang for their suggestions
and feedback as professional musicians. Last but not
least, we would like to thank Hwa Chong Institution
Science Research Centre Lab staff Ms Chua and Ms
Tan for their help and facilitation for our experiments.

REFERENCES
[1] Campbell. M., Greated.C. (1987). The Musician's Guide
to Acoustics. New York: Schirmer Books. Pg 545.
[2] Chen. C.W. (2007). The Physical Modeling ofthe
Bowing Techniques for Violin. Tatung University,Taiwan.
https://docs.google.com/viewer?a=v&q=cache:clnyUna
kDC4J:ethesys.library.ttu.edu.tw/ETD-db/ETD-search/g
etfile%3FURN%3Detd-0214107-181852%26filename%3D
etd-0214107-181852.pdf+&hl=en&gl=sg&pid=bl&srcid=A
DGEESiGJvwU9YNxWDErxcGp6f20zQ7NRjuu-OW1h2
YuroJFPrSvLpTjjKXB2s1tVkYj4Nw9jbWDmMDE4JxL72
UxovPPuO9IM4v1JP-yQKiARMuoV0xU0c06soPq1ehAL
N9kzSFAs3Sf&sig=AHIEtbSAq2Z8W9F14_mgT2-kdOu
GjHaKgQ [last access 2013/01/04]
[3] Fletcher. N. H., Rossing. T. D. (1998). The Physics of
Musical Instruments, 2nd edition. New York: Springer
Science+Business Media, Inc.
[4] Fritz. C., Blackwell. A.F., Cross. I., Woodhouse. J.,
Moore. B.C. (2011). Exploring violin sound quality:
Investigating
English
timbre
descriptors
and
correlating resynthesized acoustical modifications
with perceptual properties. Universit Pierre et Marie
Curie, UMR CNRS 7190, Institut Jean Le Rond
d'Alember. 2012 Acoustical Society of America.
http://www.ncbi.nlm.nih.gov/pubmed/22280701 [last
access 2012/12/31]
[5] Charles. J. (2010). Playing Technique and Violin
Timbre: Detecting Bad Playing. Dublin Institute of
Technology.
School
of
Engineering.
http://arrow.dit.ie/cgi/viewcontent.cgi?article=1030&c
ontext=engdoc [last access 2012/12/31]

[7] Oldham, Guy, et al. 2011. Harmonics. In Grove


Music Online. Oxford MusicOnline,
http://0-www.oxfordmusiconline.com.library.unl.edu/
subscriber/article/grove/music/ 50023 [last access
2012/12/31]

APPENDIX
Appendix A: Information of E strings
Brand

Description of string timbre

Category

Thomastik

Pure, rapid settling and

Rich/warm

Vision

rich, complex2

Pi Infield

Perfect blend of power and

Platinum

elegance, brightness and warmth, power

Projection/

brilliant projection3
Pi Infield Tin

Full sound enhances the brighter

Bright/open

overtones

Pirastro Oliv

Complex tone with warm balance5 Rich/warm

Pirastro

Bright, loud sound with easy

Chromcor

response

Lenzner

Open and bright tone

Bright/open

DAddario

Traditional sound, rich and

Rich/warm

Kaplan Light

expressive6

Bright/open

Goldbrokat

Non-whistling Traditional sound, brilliant

Projection/

DAddario

projection, volume projection7

power

Evah Pirazzi

Nuanced rich response,

Rich/warm

Gold

and tonal complexity

Evah Pirazza

Brilliant, focused projection

Projection/

Silver

and complex overtones9

power

Kaplan Heavy
8

Thomastik-Infield Vision Titanium. http://www.thomastik-infeld.com/


strings/index.html

http://www.violinist.com/discussion/response.cfm?ID=17252

Thomastik Infield http://www.thomastik-infeld.com/strings/index.html

Pirastro Oliv http://www.pirastro.com/homeset.html

DAddario Kaplan Light http://www.daddariobowed.com/Bowed


ProductDetail.Page?ActiveID=4495&productid=173

DAddario Kaplan Heavy http://www.daddariobowed.com/Bowed


ProductDetail.Page?ActiveID=4495&productid=174

Pirastro Evah Pirazzi Gold http://www.pirastro.com/homeset.html

Pirastro Evah Pirazzi Silver http://www.pirastro.com/homeset.html

55
VOYAGER 2014

ACKNOWLEDGEMENTS

[6] Crosmer. J.P. (2011). A Comparison of Viola Strings


with Harmonic Frequency Analysis. University of
Nebraska-Lincoln.
Student
Research,
Creative
Activity, and Performance - School of Music.
http://digitalcommons.unl.edu/cgi/viewcontent.cgi?a
rticle=1032&context=musicstudent
[last
access
2012/12/31]

Investigation Into The Relationship Between Violin String Timbre And


Its Harmonic Structure

In further research, we can extend this method to the


rest of violin timbre space to study how other
technical factors affect the violin timbre.

Appendix B: Data obtained from oscilloscope to plot


graph
B1: Rich/warm
Rich/Warm
Mean

Freq/Hz

StdDv w

663

f0

1326

f1

1989

f2

2652

f3

3315

f4

3979

f5

4642

f6

5305

f7

5968

f8

6631

f9

7294

f10

7958

f11

8621

f12

9284

f13

9947

f14

10610

f15

11273

f16

11937

f17

12600

f18

13263

f19

13926

f20

14589

56

f22

VOYAGER 2014

f24

Investigation Into The Relationship Between Violin String Timbre And


Its Harmonic Structure

f21

f31

15252
15916

f23

16579
17242

f25

17905

f26

18568

f27

19232

f28

19895

f29

20558

f30

21216

<EvahP Au>

<Pir Oliv>

<D'Light>

-48.9178

-52.3219

-50.2751

16.76548

16.57306

16.54861

-24.3219

-21.3164

-26.0006

-23.9871

-27.9636

-31.1955

-39.1947

-30.6193

-39.9081

-16.6909

-27.1948

-22.9255

-41.1532

-22.8385

-19.9669

-50.6977

-29.7998

-32.5611

-23.8760

-43.0134

-32.6399

-38.7424

-51.1825

-39.0466

-30.2138

-38.0432

-31.6149

-32.7347

-35.9622

-52.8383

-34.2789

-44.5197

-39.8154

-56.4396

-50.5843

-42.3778

-46.3108

-52.8896

-46.2920

-38.8692

-52.8439

-44.4274

-43.8915

-48.3091

-58.0537

-44.2106

-55.1661

-44.0552

-47.2058

-57.6363

-46.6112

-57.7746

-60.3264

-51.4791

-53.0100

-52.6583

-50.2692

-48.0239

-54.9492

-58.2125

-46.7055

-58.2345

-64.9922

-60.2342

-72.3065

-49.9887

-55.5799

-62.8681

-58.096

-57.6032

-55.7024

-60.7513

-57.3036

-59.8076

-73.7595

-50.1053

-63.1419

-59.6102

-63.5800

-69.4399

-61.0082

-79.1951

-67.8865

-66.3197

-72.3929

-71.5053

-75.7551

-71.3881

-73.3922

-64.3089

-79.6199

-80.9946

-83.4841

-80.0351

-81.2035

-80.4374

<D'heavy>

<Infd Tn>

<Lenzner>

-46.9796

-50.1856

-48.60003

-49.78748

Freq/Hz

14.31755

15.45851

15.280554

14.951425

663

-23.2034

-19.0331

-19.5275

-22.1066

f1

1326

-26.4259

-22.1297

-19.7541

-28.0752

f2

1989

-31.5589

-25.8170

-23.4693

-25.9422

f3

2652

-28.7011

-24.7244

-21.2524

-26.7395

f4

3315

-19.9807

-21.4440

-20.5243

-31.4203

f5

3979

-49.2251

-39.4998

-36.5502

-35.0844

f6

4642

-40.7303

-41.1265

-43.2104

-37.3760

f7

5305

-33.4771

-36.7104

-52.5213

-36.4543

f8

5968

-31.3569

-43.2210

-49.4628

-42.5425

f9

6631

-39.1848

-48.9570

-35.8382

-49.8028

f10

7294

-40.9009

-50.3710

-46.0777

-41.5794

f11

7958

-36.8625

-42.7695

-39.2816

-44.4372

f12

8621

-44.5119

-57.4656

-58.3107

-41.4679

f13

9284

-54.5993

-51.0946

-55.0939

-50.4862

f14

9947

-47.9365

-50.6542

-52.6020

-54.0893

f15

10610

-52.8065

-54.6007

-49.7929

-47.9365

f16

11273

-44.3334

-51.3177

-56.8609

-52.0317

f17

11937

-45.2180

-49.4432

-56.3172

-50.0899

f18

12600

-53.8148

-51.8736

-48.3107

-61.1639

f19

13263

-51.6867

-63.6741

-51.1101

-54.2677

f20

13926

-56.1292

-65.7603

-48.8673

-51.2766

f21

14589

-47.3618

-53.3814

-53.5081

-55.9683

f22

15252

-51.3454

-50.8851

-56.0906

-48.2719

f23

15916

-49.3114

-54.7639

-58.9450

-57.6311

f24

16579

-44.302

-58.1608

-52.1780

-55.6797

f0

17242

-45.8476

-58.7123

-53.9288

-62.6674

f26

17905

-60.7032

-62.5473

-54.6861

-67.2656

f27

18568

-65.0594

-70.2071

-52.5404

-63.0912

f28

19232

-61.7979

-66.6091

-63.4110

-74.1562

f29

19895

-75.1390

-73.0750

-69.0486

-67.9333

f30

20558

-72.4676

-68.2477

-76.6602

-77.4867

f31

21216

-77.3672

-77.6626

-79.4693

-78.6780

B3: Penetratin/Projecting
Penetrating/
Projecting

<Vision>

<EP Ag>

<Infd Pt>

Mean

-49.40776

-49.0742

-49.66876

16.20762

16.294933

16.363483

-20.5397

-19.4384

-19.8627

-26.4181

-25.0842

-27.0897

-32.5140

-28.3003

-29.2218

-27.1791

-26.5525

-22.3451

-30.4252

-20.3163

-22.3451

-28.1328

-30.9450

-30.5931

-32.7161

-38.0395

-34.4001

-44.7197

-44.8233

-40.1658

-28.1940

-38.1803

-38.8145

-50.2587

-28.3347

-45.9580

-38.4081

-41.7603

-46.9780

-43.1761

-44.5780

-43.6425

-56.4784

-55.8616

-62.8411

-51.1904

-50.8337

-52.7180

-44.1314

-47.1045

-45.6277

-53.5893

-51.8078

-48.7174

-34.8218

-50.6275

-38.2764

-49.6213

-54.4555

-48.4138

-53.9245

-52.8259

-62.7310

-60.0109

-50.3794

-65.9041

-51.7180

-50.8337

-66.8417

-61.5003

-58.3602

-51.2833

-61.9840

-61.4428

-65.2969

-57.8454

-59.9077

-62.3173

-58.2500

-52.8070

-55.6011

-51.9044

-56.8744

-54.4762

-64.3513

-59.7942

-52.5863

-59.1105

-67.5981

-55.0414

-76.2884

-67.4532

-71.1726

-74.8586

-75.6530

-78.8967

-78.1596

-79.3418

-76.0006

-78.6289

-80.0603

-73.2412

StdDv p
f0
f1
f2
f3
f4
f5
f6
f7
f8
f9
f10
f11
f12
f13
f14
f15
f16
f17
f18
f19
f20
f21
f22
f23
f24
f26

<Chromcor>

Mean
StdDv B

f25

f25

B2: Bright/Open
Rich/
Warm

B2: Bright/Open

f27
f28
f29
f30
f31

Freq/Hz
663
1326
1989
2652
3315
3979
4642
5305
5968
6631
7294
7958
8621
9284
9947
10610
11273
11937
12600
13263
13926
14589
15252
15916
16579
17242
17905
18568
19232
19895
20558
21216

HISTORIC VENUS TRANSIT 2012


AND INSTRUMENTATION
BACKGROUND AND OBJECTIVES

External Research Mentor


Professor Chan Aik Hui, Phil
National University of Singapore
Venus Transit is a rare astronomical event
whereby Venus, Earth and the Sun line up
in a straight line. Historically, the size of the
Solar System was accurately determined
for the first time in 1771 based on the Venus
Transit data. This research is noteworthy
because it was the last observation in this
century. The next Venus Transit will occur
about a hundred years from now in 2117. In
this project, a formula based on Kepler's
Laws and trigonometry was derived to
determine the Astronomical Unit; the
obtained results agreed with modern
methods to within 4 percent.

Aims and objectives


Our group aims to provide a comprehensive way to
calculate the value of AU through following steps:
i. Venus transit observation: Determine the visibility
graph to select a suitable observing location.
Observe the transit and measure the four essential
contact times of Venus transit.
ii. Calculation of AU: Use trigonometry and Keplers law
to calculate Sun- Earth distance.
Estimate the degree of accuracy based on errors
involved in instrumentation and calculation.

VENUS TRANSIT INSTRUMENTATION


Determine the world visibility graph
During Venus transit, not all parts of the world can
witness this extraordinary event. Hence, it is important to
determine the function that defines the world visibility
zone. Since the diameter of the Earth is relatively
insignificant compared to the Sun-Earth distance and
the apparent diameter of the sun is much larger than
that of Venus, places that can see the sun at the time of
transit can see Venus. During the transit, the earth is near
its summer solstice and its axis is tilted around 23.5 to
the ecliptic plane. During the 6-hour transit, the transit is
visible in any parts of the world that rotated to face the
Sun.
For clearer explanation, we put the earth in a 3D
coordinate system. The rotational axis is altered to be in
the z-direction. The line AB separates the region facing

57
VOYAGER 2014

Research Mentor
Dr Lim Jit Ning
Hwa Chong Institution

Background
Venus Transit is a rare astronomical event that happens a
total of four times every 243 years (8, 121.5, 8, 105.5). It
occurs when Venus and Earth meet up at the orbital
nodes, resulting in a solar eclipse of a kind. Since the 18th
century, Venus transit has been studied by scientists to
calculate the size of solar system. In 1716, Sir Edmund
Halley proposed more accurate calculations using the
contact times of the transit based on the principle of
parallax. However, his detailed workings and
corresponding accuracy are unavailable. Therefore, our
project takes the challenge to come up with a more
comprehensive method to calculate the value of AU
based on data collected from our own observation while
restricting the method to what was known and available
in terms of astronomy and mathematics at the time of
Halley.

Historic Venus Transit 2012 And Instrumentation

Vanessa Lim Yu 12S60


Yang Shuo 12S7D
Zhang Lanqiong 12S7D
Hwa Chong Institution

and opposite to the Sun. Let R represent the radius of


the Earth, a and o represents latitude and longitude
respectively. (Detailed working can be found in the
Appendix)

VENUS TRANSIT OBSERVATION


Equipment

Figure 2.1 Earth in a


2D coordinate system

! 1.

! 3.

! 4.

x2

y2 z2

z
a

Historic Venus Transit 2012 And Instrumentation

VOYAGER 2014

58
a

sin
a

sin
1

sin

z
R

sin

tan

sin 2 66.5
sin 66.5
tan o sin 66.5

tan 2

(2.2)

x
y

tan 66.5 tan


R

tan 66.5 tan


1

(2.1)

tan 66.5 x
z
sin 1
R

R2

1. Telescope (Vixen SS08V)


2. iOptron GOTONOVA controller

(2.3)

3. Eyepieces (Vixen PL 17mm MULTI-COATED, PLSSL


32mm FULLY MULTI-COATED)

(2.4)

4. Solar filter (Thousand Oaks Optical Type 2)


Procedure
1. Assemble the telescope according to the instructions.

y
sin 2

! 2.

2. Align the SOUTH mark to face the south.


a

(2.5)

cos 2 66.5

By using the software MathCai, we managed to obtain


the graph.

3. Level the mount using the bubble at the side by


adjusting the tripod.
4. Switch on the controller. The built-in GPS would have
ascertained the location and time.
5. Select SUN as target. The telescope will automatically
rotate to the Sun and track it.
6. At intervals of 10 minutes, a video camera on a tripod
is placed at the eyepiece of the telescope to record the
transit for 2 minutes. The telescope is manually adjusted
before the recording to account for the error.

Figure 2.2 World Visibility graph

According to the graph, Singapore (1.23N, 103.92E) lies


in the region where transit starts earlier than sunrise. To
overcome this limitation, two of our group members
traveled to Shenyang, China (41.630N 123.404E) where
entire transit is visible while our international
collaborator observed in Chongqing where the entire
transit is also visible.

Distance between the trajectories

Figure 2.4 Pictures of Venus transit


Figure 3.1 Flight trajectories

Observer 1 (our group)

Observer 2 (collaborator)

Shenyang, China

Chongqing,

41.63N 123.40E

China 29.57N 106.57E

Timing of 2nd contact

06:27

06:29

Timing of 3rd contact

12:32

12:32

Duration between the

6h 5min = 365min (1min)

6h 3min = 363min (1min)

location

2nd and 3rd contact

Table 3.1 Experimental data

Hence, take the mid points of the two chords


; let be the angle between OM
andOM

PRELIMINARY CALCULATIONS

Assumptions
In our calculation, there are a few assumptions made.
(1) The distance of the observers to the sun is constant.
(2) Earth and Venus move in straight lines relative to the
Sun during the transit. (3) The orbit of Venus around the
Sun lies in the ecliptic plane.

d a2

360
Tsyn

syn

2
b

bb '
2

(3.4)

R = the apparent radius of the Sun - the apparent radius


of Venus = 32' 60.3' '

2
M aMb

The apparent velocity of Venus relative to the Sun


Relative to a rotating coordinate system, in which the
line Earth-Sun is fixed, Venus cover an orbit of in one
synodic orbit, with an angular velocity of

aa '
2

R2

Ma, Mb

2
d b sin

or 0.0000132 in radians

15.5'
2

d b cos

da

0.04538
(3.5)

Calculating the baseline

(3.1)

The synodic orbital velocity of Venus relative to the


rotating coordinate system is

vsyn

syn

d vs

(3.2)

Hence, the corresponding velocity of Venus as seen from


the Earth equals to the required apparent velocity
relative to the Sun

Vsyn
vs

d ev

syn

d vs
d ev

0.0672' ' / s

(3.3)

Figure 3.2 Earth in a 3D


coordinate system

59
VOYAGER 2014

Experimental data

Due to the movement of the observer, most transit


trajectories are curves instead of straight lines, nor are
their chords parallel. This makes it hard to obtain the
distance between the two trajectories, since it is no
longer a constant. However, the distance between the
center-points of the two chords changes only little with
the angle between them, while for all other dividing
points of the chords, the difference is larger.

Historic Venus Transit 2012 And Instrumentation

RESULT ANALYSIS

The baseline is defined as the length of projection of the


line connecting the two observers onto the plane normal
to the Earth-Sun viewing line. To calculate the baseline,
we set up a three-dimensional coordinate system, with
the x-y plane on the equator and the x-axis pointing
towards the Sun.
e is the unit vector perpendicular to the plane of ecliptic

sin sin

0.0338

sin cos

0.385

cos

Computing the AU through solar parallax

Figure 3.3 Relative positions of Earth, Venus and the Sun

(3.6)

0.922

Parallax of Venus ( v ) and the parallax of the Sun (


can be computed by

Where and are the right ascension of the Sun and


obliquity of the ecliptic respectively. Points P and Q
represent the two observers,
cos
R cos

OP

cos H p
sin H p
sin p

Px
Py
Pz

PQ

Historic Venus Transit 2012 And Instrumentation

VOYAGER 2014

60

OQ

cos

cos H q

qx

R cos

sin H q

qy

sin

OQ OP

px

qx

py

qy

pz

qz

(3.7)

PQ1

PQ 2

(3.9)

0.171

px )ex (qy

py )ey (qz

pz )ez

635.9km

b2

PQ 2 e (qx

px )ex (q y

p y )ey (qz

pz )ez

844.5km

Finding the ratio

d vs
d ev

740.2km

(3.10)
(3.11)

(3.12)

by Keplers Third Law

Keplers Third Law states that, the square of the orbital


period of a planet is directly proportional to the cube of
the semi-major axis of its orbit; or simply, T 2
r3

d es
d vs
d vs
d ev

Te
Tv

2
3

d vs
d es d vs

365.256
224.701
1
d es
d vs

2
3

b
d es

b
A

1.35968

1
2.78025
1 1.35968 1

(3.15)

(3.13)

d es
d ev

1
A

d vs
d ev

d vs
d ev

(3.16)

Sub in the quantities we have obtained from the


previous calculations, Sun-Earth distance becomes

d es

740.2
2.78025 1.56 108 km
0.0000132

(3.17)

Estimation of errors
Error of the computed AU comes mainly from the
measured duration of Venus transit, the distance
between trajectories and the accuracy of baseline.
Based on we have derived earlier, defining q as
d vs
b
b
Hence,
d
q
es

PQ1 e (qx

b1 b2
2

(3.8)

b1

d es

qz

0.134
R 0.077

b
d ev

A and
B are angular distances of the chords seen by
observers A and B respectively. By geometry,

Due to the non-linear movement of the observers, the


length of the baseline changes throughout the entire
transit. Hence, a good choice would be the arithmetic
mean of the lengths of the baseline at the start and the
end of the transit. Let R represent the radius of the
Earth= 6371km
0.231
R 0.130
0.171

d d es

b d q

d ev

q d b

bq

d q

d b

dvs
des

(4.1)
b

d q

(4.2)
Length of transit trajectory
The Length
of transit trajectory in front of the solar
disk can be determined by
t
(4.3)
Where is the apparent velocity of Venus relative to the
sun and is the duration of Venus transit.
The absolute error is
(4.4)
d
wdt td

tdt

Distance between trajectories


The apparent distance
of the trajectory to the center
of the solar disk with apparent radius R is calculated
with length of transit trajectory

R2

(3.14)

d
d

RdR
RdR

)2

d
4
d
4

(4.5)

Error of the ratio q


By Keplers Third Law, let k
d es
d ev

d es
d es d ev

1
d es
1
d vs

d (k )
k

2 1
d (Te )
3 Te

dq

1 k

2
3

d (Te )
Te

d es
d vs
1
k 1

Te
Tv

2
3

8. Moar, E. (2004). Venus in Transit. Princeton, US:


Princeton University Press.

Hence,

(4.6)

1
d (Tv )
Tv

(4.7)

d (Tv )
Tv

(4.8)

9. Roberts, C., & Cooper, M. (2010, April 13). The Scale of


the Solar System: Re-enacting the Transit of Venus
Observations 5 - 6 June 2012. Retrieved 3rd March, 2012, from
http://www.fig.net/pub/fig2010/papers/ts05f/ts05f_roberts
_cooper_3851.pdf
10. Sim, C. (2010) Chapter 6 Energy and Matter
Retrieved. 12th July, 2012, from
http://www.physicalgeography.net/fundamentals/6h.ht
ml

CONCLUSION

12. Van Roode, S. (2012) Venus curved trajectory.


Retrieved 3rd March, 2012, from
https://docs.google.com/document/d/1iWm8hFB9oeL
GETh07GhHWBcoozBGskpJLlyfu3uVqSg/edit

REFERENCES

A1. Physical quantities

1. Adam and Michael Fremont High School (1996) Right


Ascension and Declination. Retrieved 3rd March, 2012, from
http://library.thinkquest.org/2595/reference/ra.html

The radius of the Earth


Sideral period (T)

Earth
6371km
365.256 days

Venus
224.701 days

Table 2.3
2. Anonymous (2012) Solar Time. Retrieved 12th July, 2012,
from http://en.wikipedia.org/wiki/Solar_time
3. The Astronomical Society of Singapore (2012). TASOS
Transit of Venus Observation at Science Center Singapore
on 6th June 2012. Retrieved 8th August, 2012 from
http://tasos.org.sg/

The apparent diameter of the Sun

32

The apparent diameter of Venus

60.3

Right ascension of the Sun on 6th, June, 2012 ( )

5059

Obliquity of the ecliptic on 6th, June, 2012 ( )

224436

A2. Detailed derivation of the World Visibility curve


At the time of transit, the earth is near its summer solstice.
Hence, the earth is tilted in such a way that the northern

4. Blatter, H. (2003) Venus Transit 2004. Retrieved 3rd


March, 2012, from
http://eclipse.astroinfo.org/transit/venus/project2004/pu
b/Blatter.etal.eng.200306.pdf
5. Google Maps (2012) Google Maps. Retrieved 10th Nov,
2012, from https://maps.google.com/maps?hl=en&tab=wl
6. Hoffmann, S. (2006) A beginners Guide to Computer
Imaging and Printing Using Digital Cameras, Scanners and
Home Photo printers Retrieved 3rd March, 2012, from
http://www.sphoto.com/homedd/pixels&imagesize.html
7. Krisciunas, K. (2012) Right ascension and declination of
the
Sun.
Retrieved
3rd
March,
2012,
from
http://people.physics.tamu.edu/krisciunas/ra_dec_sun.ht
ml

hemisphere experiences polar day. As shown in figure 1, the


shaded region is covered in sunlight. Hence, during the 6-hour
transit, any parts of the world that enters the shaded region due
to earths self-rotation can see Venus traveling across the sun.

N66.5
!
Sun

Sunshine

VOYAGER 2014

61

APPENDIX

Historic Venus Transit 2012 And Instrumentation

In conclusion, this project gives us an invaluable learning


opportunity to observe the last Venus transit of the
century and further our understanding on calculation of
the AU in the earlier days. In preparation, the world
visibility curve that we derived, enabled us to determine
desirable location for observation. We also managed to
figure out a simple yet comprehensive way to calculate
the value of AU from Venus transit. Based on the data
from our observation and our collaborators, we
obtained a result within an error of 4.28%.

11. Teets, D. A. (December 2003). Transits of Venus and


the Astronomical Unit. Mathematics Magazine, Vol. 76,
No. 5, p. 335-348

For clearer explanation, we put the earth in a 3D


coordinate system. The rotational axis is altered to be in
the z-direction. The shaded area is the plane that
separates visible and not visible zone of the sun.

x2

y2 z2

R2

sin

z
R
tan 66.5 tan
R

sin 1 ( tan 66.5 tan

sin

66.5

B
In a 2D x-z plane, the shaded area can be represented by
the line AB. Since the rotational axis tilts 23.5 from the
vertical during the orbit, the angle between line AB and
x-axis is 66.5.

62

tan 66.5 x

(A2.2)

From (A2.1) and (A2.2)

y 2 sec 2 66.5 x 2
z
sin 1
a
R

R2

(A2.3)
(A2.4)

where a represents latitude.


From (A2.2) and (A2.3)

R
sin
tan 66.5

(A2.5)

From (A2.3) and (A2.5),

y
From (A2.5) and (A2.6),
x
Since o tan 1 , where

R 1

(sin a ) 2
sin 2 66.5

(A2.6)

represents longitude

y tan

(A2.7)

From (A2.2),

tan 66.5 tan

(A2.8)

tan 66.5 tan

tan 2

sin

tan
tan 2

) 1

sin 2 a
sin 2 66.5

sin 2 66.5

sin 66.5
sin 2 a )

(sin 2 66.5
1

VOYAGER 2014

(A2.1)

where R is the radius of the earth.

Historic Venus Transit 2012 And Instrumentation

sin

o
o

sin 66.5
cos 2 66.5

sin 2

cos 2 66.5 sin 2

ACKNOWLEDGEMENTS
We are deeply grateful to our principal, Dr. Hon Chiew Weng and deputy principals
Mrs. Chin-Leow Bee Kuan and Mr. Chan Kwok Leong for their strong support in our
students science research programmes. We would also like to give special thanks to
our Dean of Research Studies, Mrs. Tan-Tiang Ai Chin, who has given us her strong
support throughout the production.
The extent to which science research has been developed at Hwa Chong has been
possible due to the support of our external research mentors and teacher mentors.
They teach, guide and provide resources for our learning journeys.
We are also indebted to the schools laboratory staff for staying back in school late
during crucial periods; offering their invaluable support and giving our students the
extra time required to complete their research projects.

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