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Table of Contents:
FHSB 1214 Biology I
FHSC 1214 Cell Biology
Experiment Description Page
Introduction Exercise 1: Writing of Lab Reports Exercise 2:
Notes on Biological Drawings
6
Practical 1 Cell Biology Studies I
Practical 1 Biological molecules I
Identification of Biochemical in Their Pure Form
21
Practical 2 Cell Biology Studies II
Practical 2 Biological molecules II
Investigation of Action of Saliva and HCl in Two Carbohydrate
Solutions
30
Practical 3 Cell Biology Studies III
Practical 3 Enzyme studies I (Experiment 1) Optional: Practical
3 Enzyme studies I (Experiment 2)
Investigation of the Effects of Catalase Concentration on
Hydrogen Peroxide Synthesis of Starch Using an Enzyme Extracted
from Potato Tuber
34
37
Practical 4 Cell Biology Studies IV
Practical 4 Enzyme studies II
Investigation of the Enzymatic Effects of Materials on Hydrogen
Peroxide
39
Practical 7 Cell Biology Studies VII
Practical 5 Cell studies I
Exercise 1: The Microscope and Its Uses Exercise 2: On-site
Assessment Exercise 3: Preparation of Wet Mount Exercise 4:
Preparation of Microscopic Slides Exercise 5: Measurement with a
Microscope Exercise 6: Observation of Starch Grains (Additional
practice tasks if time permits) Exercise 7: Observation of Hair
(Additional practice tasks if time permits)
43 49 50 52
58
64
65
- Practical 6 Cell studies II
Extraction of Cell Organelles by Differential Centrifugation
66
Practical 5 Cell Biology Studies V
Practical 7 Cell studies III
Determination of Solute Potential of Potato Cell Sap
73
Practical 6 Cell Biology Studies VI
Practical 8 Cell studies IV
Effects of Various Treatments on Pieces of Stained Potato
Cells
83
Practical 8 Cell Biology Studies VIII
Practical 9 Energetics I
Respiration of Germinating Beans
87
Practical 9 Cell Biology Studies IX
- Exercise 1: Microscopic Examination of Cells at Various Stages
of Plant Mitosis Exercise 2: Reproductive Tissues in Plants
(Histology of Plant - Lily Reproductive Structures) [Meiosis]
92
99
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FHSB 1214 Biology I
FHSC 1214 Cell Biology
Experiment Description Page
Practical 10 Cell Biology Studies X
- Exercise 1: Mitosis and Meiosis Modelling Exercise 2: DNA
Replication Modelling
110 112
- Practical 10 Energetics II
Respiration of Yeast
114
Appendix : Case study
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E-lab manuals: towards saving the earth More and more people all
over the world and in Malaysia are getting involved in saving the
planet electric cars, wind and solar energy, and paper-less
projects. What can YOU do?
Beginning January 2011, CFS PJ Biology has piloted test e-lab
manuals in an attempt to join global conservation efforts to save
the planet by saving trees. Less paper means more trees will be
left standing to absorb carbon dioxide and reduce the greenhouse
effect. Dont you & your future loved ones deserve to enjoy a
cooler planet? Another advantage is that full-colour biology
pictures will be accessible to students for the first time.
Important rules on tests and lab assignments Summary:
No MCs or any valid reasons accepted for late/missed assignments
and tests.
It is the students responsibility to submit another assignment
in-lieu or sit for a replacement test when announced (on different
topic).
The lecturer will NOT remind students to submit late/missed
assignments nor attend replacement tests. (Treating you as
adults.)
Details:
Unacceptable: MCs, valid reasons (chicken pox, met with an
accident, menstrual cramps, stomach ache, dog died etc, non-valid
reasons (forgetfulness; lateness; server/ IT problems). This is to
be fair to everyone as fake MCs and liars are present in
Malaysia.
If a student fails to submit an assignment or misses a test, the
lecturer will NOT remind you to submit a new assignment nor to sit
for a replacement test. The replacement test will be announced to
everyone in general and not to individual absentees. Those who are
supposed to attend must turn up and will not be reminded. It will
be conducted at the end of the semester on a different topic
(usually more difficult) when all students are so busy with tests
and assignments.
It is the responsibility of the student to choose one other
assignment to be submitted later in the semester when all students
are so busy with tests and assignments.
If a submission is done online, a minimum of 7 days are given to
submit your assignment. As such, no excuses will be entertained if
theres a server/ IT failure or technical problems with your UTAR
account. Hence, you have an option to submit your assignment on day
1 to be safe, or on day 7 to be stupid. If youre late, a link for
Late submissions is available if you dont mind getting a 50%
discount, or you may choose to submit another assignment which the
lecturer will not bother to remind you about.
I acknowledge reading the above & agree to be bound by terms
therein. Your signature:
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How YOU can do well in BIOLOGY
Follow the 4As and you can expect As. ttitude
Attend ALL lectures, tutorials and practicals on time without
fail.
Be attentive in class and revise your notes after class while
the topic is still fresh in your mind. Why waste time re-reading
2-3 months later?
Do your assignments faithfully as they carry marks for the
finals.
Come prepared for lessons (i.e. read up beforehand).
Read up beforehand before attending lectures so that you wont be
lost and wasted hours of your life week after week.
Why stress yourself out if you can avoid it? Do NOT count on
last minute revision for tests and examinations, as it will be too
late to catch up and seek help in areas where you may find
confusing or unclear of.
Why panic before exams because you cant find this or that? Keep
separate files for lecture, tutorial and practical. File up the
respective notes systematically so that you do not lose them along
the semester.
Do you expect the lecturer/ tutor to be available all the time
to answer your questions? It is YOUR responsibility to take the
initiative to clear your doubts or satisfy your curiosity to
understand certain scientific phenomena by reading up on the
relevant topics.
ttendance for lectures, tutorials and practicals
Lectures, tutorials and practicals carry marks that count
towards your finals.
You are expected to be present at ALL lectures, tutorials and
practicals.
Absence from any lesson must be accompanied by a photocopy of
your medical certificate presented to your lecturer/ tutor at your
next meeting.
If you know in advance that you will not be able to attend the
practical for a particular week, you are expected to inform your
tutor latest by the Friday before the affected week.
A
A
Based on a true story A professor at the National University of
Singapore recounts how on one occasion a student consulted him days
before the exam. Student: Prof, could you explain this page to me
please? Professor: What dont you understand about this page?
Student: EVERYTHING. Professor: But I already went through this
during lecture. Student: Oh, I didnt attend most of the lectures
actually. As for the next page, could you explain this page to me
please? ... and this page too and that too Prof: Im sorry, I cant
help you. Student: (Hmmmph, HES so selfish. Hey, I paid to study
here!) What do YOU think?
If the student failed, whose fault was it?
Was this student clever in skipping lectures?
Was it fair for the student to make demands on the lecturers
precious time to answer his questions?
How would the student have benefited himself if he looked up
books and other sources of information for himself first?
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ssignments
Use proper A4 foolscap for all handwritten assignments.
Write neatly and legibly in blue or black ink. Your tutor
reserves the absolute right to reject your assignment and ask you
to re-do the assignment should he/she consider it to be below the
expected quality.
Submit your assignment on time. Late submissions may entail mark
deduction or not be graded at all.
ssessments
ALL academic tests and examinations help prepare you better for
the finals.
As such, to sit for them all is not only compulsory, but
beneficial. After sitting for one, youll just want to sit for
another, and another, and another
Absence from tests and examinations MUST be covered by a medical
certificate, or will be considered to have failed the tests.
A
A
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Introduction Exercise 1 Writing of Lab Reports
hy should I bother writing lab reports in the correct way? The
Foundation Programme is designed to prepare you for undergraduate
studies at UTAR which will require the writing of lab reports all
years generally. At the end of your third
year, you may have an opportunity to work on scientific projects
which will culminate in an official scientific report. Depending on
the quality of your report, the golden chance remains of publishing
your report in a scientific journal. Such recognition may open
doors of opportunity (e.g., strengthen application for scholarships
and further studies etc.). Science professors are evaluated in most
parts of the world by the papers they write. Format of a lab report
Your lab report should be preceded by a cover page which contains
the following:
Name
Partners name Group
Date
Program
Unit code
Unit description
Year and semester of study
Title of lab report
Lecturers name
W
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Example:
Your lab report should contain the following sections:
Title
Objective
Apparatus, materials and methods (if your assignment is
submitted online, this step may be omitted)
Observations and/or results with discussion
Conclusion The following guidelines on report writing are those
required by the actual internationally-recognized scientific
community. The text in quotation marks in the following section is
taken from Warren D. Dolphin of Iowa State University. Credit has
been given to the author by citing the source. This is good
practice as opposed to plagiarism, in which copied material is
claimed as the possession of the copyist. 1 Apparatus, materials
and methods As the name implies, the materials and methods used in
the experiments should be reported in this section. The difficulty
in writing this section is to provide enough detail for the reader
to understand the experiment without overwhelming him or her. When
procedures from a lab book or another report are followed exactly,
simply cite the work, noting that details can be found in that
particular source. However, it is still necessary to describe
special pieces of equipment and the general theory of the assays
used. This can usually be done in a short paragraph, possibly along
with a drawing of the
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experimental apparatus. Generally, this section attempts to
answer the following questions:
1. What materials were used? 2. How were they used? 3. Where and
when was the work done? (This question is most important in
field
studies.)
2 Observations and/or results with discussion Results The
results section should summarize the data from the experiments
without discussing their implications. The data should be organized
into tables, figures, graphs, photographs, and so on. But data
included in a table should not be duplicated in a figure or graph.
All figures and tables should have descriptive titles and should
include a legend explaining any symbols, abbreviations, or special
methods used. Figures and tables should be numbered separately and
should be referred to in the text by number, for example:
Figure 1 shows that the activity decreased after five
minutes.
The activity decreased after five minutes (fig. 1). Figures and
tables should be self-explanatory; that is, the reader should be
able to understand them without referring to the text. All columns
and rows in tables and axes in figures should be labelled. This
section of your report should concentrate on general trends and
differences and not on trivial details. Many authors organize and
write the results section before the rest of the report. 2.1
Recording Qualitative Data Qualitative experiments include those
that require observations of non-quantifiable data such as
observations of colour, slides and whole specimens. Below are
guidelines on reporting a segment of qualitative experiments.
Liquid in container: Be careful to distinguish accurately among
solution, suspension, emulsion etc. Often, mixture is a safe
descriptive term to employ. It is your responsibility to look up
the definitions as studied in secondary school.
KI solution was added to the starch suspension
emulsion of lipid droplets in water Amount of light penetrating
solution Be careful to distinguish accurately among transparent,
translucent and opaque. It is your responsibility to look up the
definitions as studied in secondary school.
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Colour Some descriptions of colour are unacceptable as they are
ambiguous.
Light/pale brown, instead of beige
Murky/ cloudy white, instead of milky If theres a change in
colouration, you may choose to report as follows.
The initial blue colouration of the mixture turns green, then
yellow and may finally appear brick red.
If the transition cannot be easily seen, at least state the
initial and final colours. If there is no change, one must state
the colour (e.g., it remained blue). It is incomplete to only
report there was no colour change without at least recording the
initial colour. Precipitate One should comment on the precipitate
colour and relative quantity. To do so, the mixture must be left to
settle.
Colour of precipitate - green, yellow, brick red precipitate
Amount of precipitate - a little, moderate amount, abundant
Example: When describing observations involving Benedicts test, one
should report that when one shakes the test tube containing
Benedicts solution and precipitate, the entire mixture will take
the colour of the precipitate. This colour upon shaking is recorded
and also the amount of light penetrating solution (transparent/
translucent/ opaque).
Moderate amount of brick red precipitate suspended in solution,
which bore a tinge of blue. Solution was opaque.
Note: Particles cannot be regarded as precipitate. (e.g.
groundnut particles in water.) 2.2 Recording Quantitative Data
Quantitative experiments include those that require observations of
quantifiable data such as time, quantity, weight, etc. Tabulation
and graphing
There are two categories of data normally used in reporting
quantitative results raw data and processed data. Raw data refers
to the readings obtained from measurements (e.g., length, weight,
height, quantity, etc.).
The table must be accompanied by the following features:
Informative table title
Gridlines
Columns/ rows with appropriate headings and units (units and
calculations should not be in the table body)
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All processed data related to and required for plotting graph
must be shown in the table. E.g. Averages, rate of yeast
respiration in terms of no. of bubbles formed per minute.
Precision and decimal places: One must express data according to
the precision afforded by the instrument. E.g., if the instrument
can weigh an item as light as 0.1 g, then do not record it as 0.10
g, so as to correctly reflect the precision of the instrument. Note
that the decimal places in the table must be the same for the same
unit of measurement, and reflect the precision of the instrument.
If a measurement unit is converted to percentage or any other unit,
one is not bound by the precision of the instrument. However, the
recording should maintain a consistent and reasonable use of the
number of decimals (e.g., avoid too many decimals 88.8888888 %).
Note that the table and graph below feature such consistency of
decimal places.
Precision of processed data can be presented in the following
manner:
Averages calculated need not follow the decimal places of the
raw data.
Processed data involving summation and/ or subtraction should
follow decimal places of the raw data.
Decimals arising from processed data involving multiplication
and/ or division should be reasonable (e.g., not unnecessarily
long).
Sample table: Title: Mass of precipitate of standards at various
concentrations of glucose solutions.
Precipitate mass (g)
Glucose concentration (%)
Reading 1 Reading 2 Reading 3 Average
4 0.1 18.6 18.4 18.7
2 8.2 9.3 9.0 8.8
1 5.2 4.5 4.8 4.8
0.5 2.3 1.8 2.1 2.1
0.1 0.4 0.3 0.4 0.4
Graph
Plot a graph that will show the trend of the investigation.
Include the following in the plotting of graph:
Informative title
x-axis : labelled, including units (independent variables)
y-axis : labelled, including units (dependent variables)
appropriate scale used
points plotted
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Shape of graph can only be drawn using pencil, blue and black
ink pen
points plotted according to table of data
best fit line/ curve Sample graph:
Note: The line of the plot does not go beyond the concentrations
used (no extrapolation
of points). Hence, one should not extrapolate otherwise it is a
claim that a certain y value is predicted for a certain
concentration.
Avoid clashing headings with clashing units (e.g., headings with
two different units but both have gram in their units gram eggs vs.
gram nutrients per gram plain feed)
Mass of eggs laid in a week (g)
Amount of nutrients (g/ g plain feed)
0.30
0.25 0.20 0.15 0.10 0.00
Mean 78.0 74.0 69.3 62.7 59. 7 58.0
2.3 What if I dont obtain desired results? For the purpose of
your UTAR lab report, if you dont obtain the desired results, just
record them as they are. By right, you should repeat it however,
you may be constrained by a limited amount of supplied solutions in
the UTAR lab and time. Hence, if your repeats involve consuming
more solutions, please ask your tutor first. You may put a footnote
concerning the expected results. In your discussion, be sure to
explain the possible reasons for the anomaly.
Average mass of precipitate of standards at various
concentrations of glucose solutions
0
2
4
6
8
10
12
14
16
18
20
0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0
Concentration of glucose solution (%)
Ave. precipitate mass (g)
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3 Discussion This section should not just be a restatement of
the results but should emphasize interpretation of the data,
relating them to existing theory and knowledge. Speculation is
appropriate, if it is so identified. Suggestions for the
improvement of techniques or experimental design may also be
included here. In writing this section, you should explain the
logic that allows you to accept or reject your original hypotheses.
You should also be able to suggest future experiments that might
clarify areas of doubt in your results. 3.1 General Comments on
Style 1. All scientific names (genus and species) must be
italicized. Underlining indicates
italics in a typed paper. 2. Use the metric system of
measurements. Abbreviations of units are used without a
following period. 3. Be aware that the word data is plural while
datum is singular. This affects the
choice of a correct verb. The word species is used both as a
singular and as a plural.
4. Numbers should be written as numerals when they are greater
than ten or when
they are associated with measurements
6 mm or 2 g
two explanations of six factors.
When one list includes numbers over and under ten, all numbers
in the list may be expressed as numerals; for example,
17 sunfish, 13 bass, and 2 trout.
Never start a sentence with numerals. Spell all numbers
beginning sentences.
5. Be sure to divide paragraphs correctly and to use starting
and ending sentences that indicate the purpose of the paragraph. A
report or a section of a report should not be one long
paragraph.
6. Every sentence must have a subject and a verb.
7. Avoid using the first person, I or we, in writing. Keep your
writing impersonal, in the
third person. Instead of saying, "We weighed the frogs and put
them in a glass jar," write, "The frogs were weighed and put in a
glass jar."
8. Avoid the use of slang and the overuse of contractions.
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9. Be consistent in the use of tense throughout a paragraph--do
not switch between past and present. It is best to use past
tense.
10. Be sure that pronouns refer to antecedents. For example, in
the statement,
"Sometimes cecropia caterpillars are in cherry trees but they
are hard to find." Does "they" refer to caterpillars or trees?
After writing a report, read it over, watching especially for
lack of precision and for ambiguity. Each sentence should present a
clear message. The following examples illustrate lack of
precision:
"The sample was incubated in mixture A minus B plus C."
Does the mixture lack both B and C or lack B and contain C?
"Protection against Carcinogenesis by Antioxidants"
The title leaves the reader wondering whether antioxidants
protect from or cause cancer.
The only way to prevent such errors is to read and think about
what you write. Learn to reread and edit your work.
Identify trends/ patterns by in words the trend shown in the
graph. Remember to make reference to the values shown on the graph.
Explain all the observations or trend obtained during the
investigation.
As temperature increases from 25 oC to 50OC, rate of yeast
respiration/ mean number of bubbles formed per 3 mins. increases
proportionately/ linearly from 7 to 28.
In summary, the discussion should be correctly applying the
theoretical concept involved in the experiment.
4 Conclusion
State the general trend obtained through the investigation and
provides a concise conclusion about the investigation. 5 Literature
Cited This section lists all articles or books cited in your
report. It is not the same as a bibliography, which simply lists
references regardless of whether they were cited in the paper. The
listing should be alphabetized by the last names of the authors.
Different journals require different formats for citing literature.
For articles: Fox, J.W. 1988. Nest-building behavior of the
catbird, Dumetella carolinensis. Journal of Ecology 47: 113-17.
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For Books: Bird, W.Z. 1990. Ecological aspects of fox
reproduction. Berlin: Guttenberg Press. For chapters in books:
Smith, C.J. 1989. Basal cell carcinomas. In Histological aspects of
cancer, ed. C.D. Wilfred, pp. 278-91. Boston: Medical Press. When
citing references in the text, do not use footnotes; instead, refer
to articles by the author's name and the date the paper was
published.
Fox in 1988 investigated the hormones on the nest-building
behavior of catbirds.
Hormones are known to influence the nest-building behavior of
catbirds (Fox, 1988).
When citing papers that have two authors, both names must be
listed. When three or more authors are involved, the Latin et al.
(et alia) meaning "and others" may be used. A paper by Smith,
Lynch, Merrill, and Beam published in 1989 would be cited in the
text as:
Smith et al. (1989) have shown that... This short form is for
text use only. In the Literature Cited, all names would be listed,
usually last name preceding initials.
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Introduction Exercise 2 Notes on Biological Drawings
______________________________________________________________________
Drawings are an aid to precise observations and for this reason
they are an important part of laboratory work. In the practical
examination, the examiner will have only your written recordings
and drawings to assess you. Therefore full recordings and neatly
labelled drawings are of great importance. Keep the following
points in mind when making drawings: 1. Use a sharp, pointed HB/2B
pencil. 2. Drawings should be as large as possible and made to fit
into the space available. 3. Attention must be given to the general
shape and proportion of the specimen.
First consider what you want to show. Then plan your drawing so
that various parts are in proportion and fit on the page. Small
marks indicating the length and breadth of the drawing are of great
help in planning, and a faint outline can be rapidly drawn to show
the relative positions of the parts.
4. The final outline should be drawn with clean firm lines (not
sketchy broken lines). Details should be put in clearly with a
sharp pencil. If important details are too small to be shown in
proportion, they can be shown in an enlarged drawing on the
side.
5. Drawings should be accurate records of your observations. The
biologist is not expected to be an artist, but to become, in some
degree, a draughtsman. Clear and accurate line drawings are
needed.
6. Shading and colouring should be avoided. It should be
possible to make the drawing perfectly clear by the judicious use
of thick and thin pencil lines and careful cross-hatching. Get into
the habit of making your drawings large and clear.
7. As important as the drawing is the labelling. This should be
done neatly in pencil and the letters printed. Each label should be
connected to the appropriate part of the drawing by a clear
guideline without arrowheads. Do not label too close to the
drawings, and never write on the drawing itself. Always make sure
that each drawing is fully labelled before you leave it. Guidelines
to the labels must be drawn with pencil and ruler and never
crossing one another.
8. Each drawing must always have a title. The title should
specify whether it is a transverse section, s longitudinal section,
whole mount, etc.
9. Magnification of drawing can be stated if necessary.
Calculate using the formula, Magnification = size of drawing___
Actual size of specimen 10. Plan diagrams of microscopic
sections should not include any cell structure.
They are outline drawings showing relative amounts and
distribution of various tissues.
11. In making high-power detailed drawings, repeated features
need not all be drawn but only a small representative portion
showing a few large accurate cells (3 or 4 adjacent cells) of each
type must be indicated.
12. It is sometimes appropriate, particularly when drawing live
specimens, to make succinct notes to the labels. These are called
annotated drawings, which are particularly valuable as they combine
a record of structure with functional observations. Annotations
must be beneath the labels.
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Example of an annotated drawing Title: Detailed drawing of a
Hydrilla leaf cell Magnification power: 10x. 40x
Note: The distinction between a plan diagram and a drawing: A
drawing is an exact and accurate representation of an object,
unlike a diagram which is a simplified outline. Warning: Memorized
textbook drawings or diagrams, bearing little likeness to the
specimens or observations will not be awarded marks.
Cell wall (made of cellulose, stained yellow)
Chloroplast (moves in cytoplasm, site of photosynthesis; stained
brownish with iodine solution)
Cytoplasm (granulated; found at periphery of cell; stained light
yellow with iodine solution)
Cell vacuole (contains salt and sugar solution; bound by
membrane tonoplast; colourless)
Nucleus (doubled membrane organelle embedded in the cytoplasm;
control center of the cell; stained orange brown with iodine
solution.)
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A tissue map or plan diagram refers to a generalized outline of
the tissue regions of a specimen. If such is requested, no detailed
drawings of individual cells are required. The illustration below
is a cross section of a leaf. The items in the square box are
detailed drawings of cells whereas those of the vascular bundle
reflect a tissue map or plan diagram.
Is the picture below a drawing or diagram?
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General Marking Scheme It is your responsibility to read the
guidelines in the introduction to this lab manual. In the past many
clever students lost marks simply because they failed to look up
instructions already provided. Would you like to be as clever as
them? This is an example of what a marking scheme may look like
for: General instructions for students:
One to two slides may be drawn [please consult your lecturer].
If two drawings are to be done, each student is allowed only 30
mins per slide for drawings to be done in the 1st half.
The second half of the practical will be used to assess students
microscope and / or identification skills on one type of slide.
Students are required to identify at least any three structures
which they choose.
For online submissions, students are required to upload pictures
of their drawings (more instructions on WBLE).
In order not to lose marks unnecessarily, please ensure that you
comply with the instructions on writing lab reports at the
beginning of this manual, including your particulars (e.g., name,
group, etc.) as stated in the instructions.
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Checklist for biological drawings This section may comprise 10
marks out of 20 marks. Any mistake will result in subtraction of 1
mark.
Please read full version for your personal success
Items Remarks from Lab Introduction
1. Appropriate & comprehensive (detailed and complete)
title
The title should specify whether it is a transverse section, s
longitudinal section, whole mount, etc.
2. Written indication of objective used (multiplied by __x
ocular lens)
3. Requirement for detailed drawing or plan diagram instruction
adhered to
A tissue map or plan diagram refers to a generalized outline of
the tissue regions of a specimen. If such is requested, no detailed
drawings of individual cells are required.
4. Correctly labelled items
5. Annotations if requested annotated drawings, which are
particularly valuable as they combine a record of structure with
functional observations.
6. Drawing is as what is seen under microscope (i.e., not
textbook-perfect picture)
Memorized textbook drawings or diagrams, bearing little likeness
to the specimens or observations will not be awarded marks.
7. Magnification if requested
Magnification = ___size of drawing Actual size of specimen 8.
Method of calculation if
requested
9. Early submission up to tutor to delete marks according to
lateness
10. Overall impression of drawings (e.g., neatness)
a. Satisfactory b. Quite good c. Good d. Very good e.
Excellent
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General guidelines for biological photo(s)
please read full version for your personal success
Items Remarks from lab introduction
1. Title - Appropriate & comprehensive (detailed and
complete)
Specify whether it is a transverse section/cross section,
longitudinal section, whole mount, etc.
2. Magnification - Written indication of objective used
(multiplied by __x ocular lens)
10x. 4x or 40x / 10x. 10x or 100x
3. Label - Correctly labelled items
Label box should not overlap the photo. Label line should point
to the specific area. Label line should not overlap each other.
4. Annotations if requested Include the structure and function
Annotation should be beneath the label.
5. Images and tidiness Photos taken must be clear.
6. Late submission - up to tutor to delete marks according to
lateness
General instructions for lecturers: Two areas of assessment:
1) Biological drawings 1st hour allocated for drawing Kindly
instruct students to draw slides according to availability (as
supplied). The manual provides options to lecturers to request 1
detailed drawing or 2 plan diagrams. Note: if students are to
submit drawing on the spot, 1 drawing should suffice. Recommended
time allocation: Briefing (5-10 mins) + Slide familiarisation &
reading (30 mins) + drawing (30 min if 1 detailed drawing; longer
if 2 plan diagrams required)
2) On-site assessment 2nd hour allocated for on-site assessment.
On-the-spot identification of 3 structures (10 marks); 2-3 students
assessed within 3 minutes. Drawing (10 marks) Recommended time
allocation: On-site assessment (50 min).
General instructions for lab reports:
Prepare your answers on your own A4 sheets of paper.
You are not required to re-write questions.
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Practical 1 (FHSB 1214 Biology I & FHSC 1214 Cell Biology)
Identification of Biochemical in Their Pure Form
______________________________________________________________________
Objective: To identify the components of the solution in its pure
form with various food tests and state the justifications.
Important notice: Any heating that has to be done in the following
tests should be carried out in a water bath at 95oC. Direct heating
of test-tubes should not be taken place. Apparatus &
Equipments: Test tubes Test tube rack Water bath, 95oC Spatula Test
tube holder Materials:
Iodine
1 M hydrochloric acid
Sudan III
Starch solution
Corn oil
Egg albumin
1% copper sulphate solution
DCPIP (dichlorophenolindophenol) solution
Ascorbic acid (or vitamin C tablet, or lemon juice)
1% sucrose solution (Analar sucrose must be used to avoid
contamination with a reducing sugar Benedicts reagent
1 M sodium hydroxide (or potassium hydroxide or sodium hydrogen
carbonate)
Millons reagent 1% glucose solution
Absolute ethanol
Introduction The nutrients in the food you eat supply your body
with energy for growth and repair. These principle substances
include carbohydrates, proteins, fats, minerals and vitamins. We
can test for the presence of these important compounds in food by
using chemical reagents that react in predictable ways in the
presence of these nutrients. Please refer to the notes given above
on:
How to record qualitative data. (Marks will be awarded based on
proper recording.)
What to do if you dont obtain the desired results.
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Flowchart Students will be allowed to proceed with the
experiment only if they have come into the laboratory with a
flowchart of the days experiment. Procedures: The following tests
are to be done in pairs unless otherwise specified. Part 1:
Identification of Carbohydrates Test for reducing sugars The
reducing sugars include all monosaccharide, such as glucose and
fructose, and some disaccharides, such as maltose and lactose, use
0.1 1% sugar solutions. Common tests for reducing sugars include
Benedicts test (described below) and Fehlings test (not done here).
See basis of test below for explanation of the following
reaction:
Benedicts test for reducing sugars:
Procedure* Basis of test Observation
Reducing sugar test
Add 2 cm3 of any one solution of the reducing sugar provided to
test-tube. Add an equal volume of Benedicts solution. Using a
test-tube holder, shake and heat at a high temperature for one
minute (a water bath is provided), shaking continuously to minimize
spitting.
Benedicts solution contains copper sulphate. Reducing sugars
reduce soluble alkaline blue copper sulphate containing copper (II)
ions, Cu2+ to insoluble red-brown copper oxide containing copper
(I). The latter is seen as a precipitate.
[Note: report after shaking and after contents settle down; see
introduction pg. 9]
*: Please do NOT remove measuring cylinder or any other item
from the stations provided. Observe and report characteristics of
tube contents before and after precipitate settles to bottom of
tube, taking note of liquid, colour and precipitate. Test for
non-reducing sugars The most common non-reducing sugar is sucrose,
a disaccharide. If reducing sugars have been shown to be absent
(negative result in above test), a brick-red precipitate in
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the test below indicates the presence of a non-reducing sugar.
If reducing sugars have been shown to be present, a heavier
precipitate will be observed in the following test than with the
reducing test if non-reducing sugar is also present. The proper
procedure for testing for an unknown carbohydrate sample for
non-reducing sugars involves:
First test for reducing sugars: Benedicts test on the unknown
fresh sample Why is this step necessary? What results will one get
which will cause this step to be called a negative test?
Second test for reducing sugars: Benedicts test on the
acid-hydrolysed unknown sample What results will one get which will
cause this step to be called a positive test?
Procedure* Basis of test Observation
Non-reducing sugar test
Add 2 cm3 of fresh sucrose solution to a test tube. Add 1 cm3
O.1 M hydrochloric acid. Using a test-tube holder, heat at a high
temperature for one minute. Carefully neutralize with equal volume
(1 cm3) of 1 M sodium hydroxide or sodium hydrogen carbonate or
potassium hydroxide. (Care is required because effervescence
occurs.) Finally, add an equal volume (4 cm3 ) of Benedicts
solution to the acid-hydrolysed sugar solution. Using a test-tube
holder, shake continuously to minimize spitting when heating at a
high temperature for one minute (a water bath is provided).
A polysaccharide or disaccharide can be hydrolyzed to smaller
component constituents by boiling with O.1 M hydrochloric acid.
Sucrose is hydrolyzed to glucose and fructose, both of which are
reducing sugars and give the reducing sugar result with the
Benedicts test.
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Additional Information The mixture is likely to bump violently
during heating and extra care should therefore be taken. The test
is semi-quantitative, that is, a rough estimation of the amount of
reducing sugar present will be possible. The final precipitate will
appear green to yellow to orange to red-brown with increasing
amounts to reducing sugar. The initial yellow colour blends with
the blue of the copper sulphate solution to give the green
colouration. Is the precipitate that of reducing sugar or copper
oxide?
*: Please do NOT remove measuring cylinder or any other item
from the stations provided. Observe and report characteristics of
tube contents before and after precipitate settles to bottom of
tube, taking note of liquid, colour and precipitate. Test for
starch Starch is only slightly soluble in water, in which it forms
a colloidal suspension. It can be tested as a mainly solid in
suspension.
Procedure* Basis of test Observation
Iodine test
***Note: The starch prepared for you is already cooked starch.
Add a few drops of 1% cooked starch solution on a white tile. Add a
few drops of I2/KI solution (iodine). Be sure to mix them together
on the tile with an object such as your pen cover.
A polyiodide complex is formed with starch.
*: Please do NOT remove measuring cylinder or any other item
from the stations provided.
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Part 2: Identification of Lipids Lipids include oils (such as
corn oil and olive oil), fats and waxes.
Procedure* Basis of test Observation
Sudan III
Sudan lll is a red dye. Add 2 cm3 of oil to 2 cm3 of distilled
water in a test-tube. Add a few drops of Sudan III and shake.
Fat globules are stained red and are less dense than water.
Emulsion test
Add 2 cm3 fat or oil to a Test-tube containing 2 cm3 of absolute
ethanol. Dissolve the lipid by shaking vigorously. Add an 4 cm3
volume of cold (or tap) water.
Lipids are immiscible with water. Adding water to a solution of
the lipid in alcohol results in an emulsion if tiny lipid droplets
in the water which reflect light and give a white, translucent
appearance.
[***Note: report after shaking and after contents settle
down]
*: Please do NOT remove measuring cylinder or any other item
from the stations provided.
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Part 3: Identification of Proteins A suitable protein for these
tests is egg albumen.
Procedure* Basis of test Observation
Millons Test
Add 2 cm3 protein (albumin) solution or suspension to a
test-tube. Add 1 cm3 Millons reagent. Using a test-tube holder,
heat at a high temperature for one minute (a water bath is
provided). Millons reagent is poisonous: be extremely careful!
Millons reagent contains mercury acidified with nitric acid,
giving mercury (II) nitrate and nitrite. The amino acid tyrosine
contains a phenol group which reacts to give a red mercury (II)
complex. This is a reaction given by all phenolics and is not
specific for proteins. Protein usually coagulates on boiling. Thus
appearing solid. The only common protein lacking tyrosine likely to
be used is gelatin.
Biuret Test
Add 2 cm3 (albumin) protein solution to a test tube. Add an
equal volume of 5% sodium hydroxide (or potassium hydroxide)
solution and mix. Add 2 drops of 1% copper sulphate solution and
mix. No heating is required.
A test for peptide bonds. In the presence of dilute copper
sulphate in alkaline solution, nitrogen atoms in the peptide chain
form a purple complex with copper (II) ions, Cu2+. Biuret is a
compound derived from urea which also contains the CONH group and
gives a positive result.
*: Please do NOT remove measuring cylinder or any other item
from the stations provided.
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Part 4: Identification of Vitamin C (ascorbic acid) ***Note: If
more than 5 drops of ascorbic acid are required to turn DCPIP to
colourless, please dilute the latter significantly. This test can
be conducted on a quantitative basis if required, in which case the
volumes given below must be measured accurately. A suitable source
of vitamin C is a 50/50 mix of fresh orange or lemon juice with
distilled water. Vitamin C tablets may also be purchased.
Procedure* Basis of test Observation
DCPIP test
Using 0.1% ascorbic acid solution as a standard. Add 1 cm3 of
DCPIP solution to a test-tube. ***Add the 0.1% ascorbic acid to the
DCPIP drop by drop until it becomes approximately colourless (or by
stirring gently if youre provided with a syringe needle/ glass
rod). Note the no. of drop(s) of ascorbic acid solution used.
DCPIP is a blue dye which is reduced to a colourless compound by
ascorbic acid, a strong reducing agent.
Additional Information Shaking the solution would result in
oxidation of the ascorbic acid by oxygen in the air. The effects of
shaking and of boiling could be investigated.
*: Please do NOT remove measuring cylinder or any other item
from the stations provided.
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Assignments Please check with your tutor which option is
required for you. Option 1: (please refer to WBLE/ Turnitin for
instructions which may incorporate other options below) Option 2:
Skills Based Assessment: Tabulation of qualitative data
1. Tabulate your observations above for each biochemical food
test executed, according to the guidelines provide in the
introduction on writing lab reports. Note: The table in the lab
manual for this task is not presented correctly.
2. Wrong results are alright for this experiment. 3. No need to
write procedure, basis of test, discussion or conclusion. 4. You
may choose to construct one or more tables. 5. For tests involving
carbohydrates, observe and report characteristics of tube
contents before and after precipitate settles to bottom of tube,
taking note of liquid, colour and precipitate as above.
o Liquid mixture, solution, suspension, emulsion? transparent,
translucent, opaque?
o Colour state initial and final colours?
o Precipitate (if any) colour of precipitate? amount of
precipitate?
Better understanding of terms:
Mixture
Solution
Suspension Option 3: Skills Based Assessment: Critical thinking/
Discussion 1. How could you determine the concentration of ascorbic
acid in an unknown sample?
2. You are provided with three sugar solutions. First one
contains glucose, second one
is a mixture of glucose and sucrose, and lastly is sucrose
solution. (a) How could you identify each solution? (b) Supposing
that the apparatus were available, and time permitted, briefly
discuss
any further experiments you could perform to confirm your
results. 3. After carrying out Benedicts test, a student concludes
that the obtained positive results prove that glucose is present.
True or false? Provide a reason. 4. After carrying out Benedicts
test, a student identifies the coloured precipitate as reducing
sugar. True or false? Provide a reason. 5. A student pours
Benedicts solution into a tube containing a carbohydrate. No colour
change is obtained. The student concludes that the carbohydrate is
not a reducing sugar. True or false? Provide a reason.
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6. A student adds acid to a solution of sucrose followed by
neutralization and, finally, Benedicts test. A negative (instead of
positive) result is obtained. Explain why. 7. Why does sucrose
yield positive results after carrying out the non-reducing sugar
test? What are the components of sucrose? For those who have done
Sem 3 3. How would you make 100 cm3 of a 10% glucose solution? 4.
Starting with stock solutions of 10% glucose and 2% sucrose how
would you make
100 cm3 of a mixture of final concentration 1% sucrose and 1%
glucose?
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Practical 2 (FHSB 1214 Biology I & FHSC 1214 Cell Biology)
Investigation of Action of Saliva and 3 M Hydrochloric Acid in Two
Carbohydrate Solutions
______________________________________________________________________
Objective: Students are expected to state the objective of this
experiment. Apparatus & Equipments: Boiling tubes Metal test
tube racks Pipette filler Graduated glass pipette, 10ml Water bath,
37-40oC Water bath, ~90-95oC Beaker Pasteur pipette Materials:
Carbohydrate solution A Carbohydrate solution B Benedicts solution
3 M Hydrochloric acid 3 M Sodium hydroxide (or potassium hydroxide)
Flowchart Students will be allowed to proceed with the experiment
only if they have come into the laboratory with a flowchart of the
days experiment. Procedures: This experiment is to be done in
pairs. To avoid congestion, each pair should collect the following
before beginning the experiment:
8 ml NaOH
16 ml Benedicts Solution 2ml Solution A
42ml Solution B
2ml HCl
1 pipette and 1 rubber teat (to be washed with distilled water
each time before reuse)
5 ml measuring cylinder (to be washed with distilled water each
time before reuse)
Metal test tube racks (not wooden) Overview Please see tables 1
& 2 on the next page to get a rough idea of what is required in
the experiment. Can you identify in the instructions that follow,
how the tubes are to be placed under various temperatures and time
periods? Carry out your investigation as follows. 1. Prepare two
test tubes containing 2 ml solution A and 2 ml solution B
respectively.
Add 2 ml Benedicts solution to each test tube. Heat both tubes
together in the hotter (~90-95oC) water bath for two minutes.
Record the results in table 1.
2. Pipette 10 ml solution B into each of four test-tubes and,
label the tubes 1, 2, 3 and
4 respectively with labelling paper (or masking tape) near mouth
of tube. Write the initials of your group name or individuals.
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3. Place tubes 1 and 2 in a water bath of ~37o, and tubes 3 and
4 in a water bath of ~95oC (It doesnt matter how long you put it in
at this stage as no saliva or HCl have been added yet).
4. Salivate into a separate test-tube till it reaches a height
of about 1 cm - 1.5 cm.
Dilute the saliva with an approximately equal volume of
distilled water.
5. Ensure that the following two steps (5 and 6) adding of
saliva or HCL into the respective tubes (mentioned in the next
sentence and below) is done approximately at the same time. (Why is
this necessary?)
Note: for the following, please ensure that the respective tube
into which saliva is going to be dropped does NOT leave the water
bath (especially 95 oC) for more than 30 seconds! (Why is this
necessary?) 6. Use a 5 ml measuring cylinder to measure out 2 ml of
the diluted saliva prepared in
(3) and pipette 1 ml each into tubes 1 and 4. Shake the contents
of the tubes well to ensure thorough mixing.
7. Use a measuring cylinder to measure out 2 ml HCl and pipette
1 ml each into tubes
2 (already in water bath of ~37oC) and 3. Place tubes 3 and 4 in
a water bath set at 95 oC. Let tubes 1, 2 (already in water bath of
~37oC), 3 & 4 (recently in water bath of ~95oC) incubate at
their respective temperatures (see Table 2) for 35 minutes from
this moment.
8. Label 4 more new tubes (either test tubes or boiling tubes)
as follows: 1, 2, 3 and
4. After 5 minutes of incubation of tubes labelled 1 to 4
prepared previously, pour out about one-third of the total volume
of the contents from all these tubes into the respective newly
labelled test tubes (e.g., 1 into 1, 2 into 2 etc.). Ensure that
the volume in each of the tubes 1-4 is approximately the same (why
is this important?). Straightaway, place back the original tubes
(labelled 1-4) back into the respective temperatures of
incubation.
9. Neutralize the acid in each of tube labelled 2 and 3 with 2ml
of sodium hydroxide (or potassium hydroxide) (each). Shake each
tube (2 and 3) to ensure uniform mixing.
10. Remove 1ml of the solution from each tube (1 to 4) into new
tubes and label
appropriately as you wish as long as you dont get confused. To
carry out Benedicts test, add an equal volume of Benedicts solution
(1 ml) for each tube. Using a test-tube holder, shake and heat at a
high temperature for one minute (use the hotter water bath
provided), shaking continuously to minimize spitting. Record your
observations in table 2.
11. Wash the test tubes 1 to 4. After 35 minutes of incubating
tubes 1 to 4, pour out
about one-third of the total volume from test samples from all
the tubes into the respective tubes labelled 1- 4.
12. Neutralize the acid in each test tube labelled 2 and 3 with
1ml of sodium
hydroxide (or potassium hydroxide). (Why is neutralization
necessary?) Remove 1ml of solution from each tube 1 to 4 and carry
out Benedicts test with an equal
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volume of Benedicts solution (1 ml) for each tube. Remember to
heat your sample (please see previous. Record your observations in
table 2.
13. Add a few drops of fresh solution A and B separately spaced
on a white tile. On
each solution, add 1-2 drops of I2/KI solution (iodine). Be sure
to mix them together on the tile with an object such as your pen
cover. Record your observations in the table 1.
Note: no penalization for unexpected results. Please refer to
Practical 1 Exercise 1 Writing lab report. Table 1: (title)
Observations Conclusions
Solution A
Benedicts test: Iodine test:
Solution B
Benedicts test: Iodine test:
Table 2: (title)
Tube Contents Temp (C)
Benedicts TestColour Observation
After 5th min (from tubes 1 4
into 1 4)
After 35th min (from tubes 1 4 into
1 4) 1 10 ml solution B
1 ml saliva 37
2 10 ml solution B 1 ml 3 M HCl
37
3 10 ml solution B 1 ml 3 M HCl
95
4 10 ml solution B 1 ml saliva
95
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Guidelines Observations For Benedicts test and Iodine tests,
please follow lab manual guidelines for students on writing lab
report on the following:
o Liquid mixture, solution, suspension, emulsion? transparent,
translucent, opaque?
o Colour state initial and final colours?
o Precipitate colour of precipitate? amount of precipitate?
Conclusions Absence/presence of what type of carbohydrate?
Results and Discussions: 1. The results and discussion sections of
your report should not exceed 2 pages. 2. Ensure that the
guidelines for constructing tables and recording results for
this
experiment are adhered to (see introduction). 3. If youre
required to write a discussion straight-to-the-point, follow the
numbering
below (please check with your lab tutor). If your report is
full-length, write your discussion in prose form (please check with
your lab tutor). Theory to apply: Refer to relevant information
from lecture topics on biological molecules and enzymes.
Discussion should contain:
1) Name of enzyme involved 2) Specific action(s) of enzyme
involved 3) 4) Effect of HCl on substances (e.g., Solution B) 5)
Effect of temperature on substances (e.g., Solution B, saliva
content) 6) Product:
a. Identification (make suggestion(s)/ educated guesses) b.
Structure (e.g., chemical classification etc.)
7) Basis of test() used 8) Which carbohydrate is more complex, A
or B? Give a reason. 9) Conclusion: Summary of results
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Practical 3 (FHSB 1214 Biology I & FHSC 1214 Cell Biology)
Investigation of the effects of catalase concentration on hydrogen
peroxide
Objective: To investigate the effects of different catalase
concentration on the decomposition of hydrogen peroxide. Apparatus
and Materials: 5 test or boiling tubes Scalpel/ pen knife 1 beaker
(500cm3) White tile 1 beaker (250cm3) Mortar and Pestle 1 Retort
Stand (optional) Weighing boat 1 rubber bung with delivery tube 4
filter funnel and filter paper (optional) 4 test tubes or plastic
vials (if provided) Potato 1% hydrogen peroxide solution Hand-held
pipette **Caution: Hydrogen peroxide is formed continuously as a
by-product of chemical reactions in living cells; it is a very
toxic (poisonous) substance. Note to lecturer: This experiment may
be done together with Experiment 2 if the lab session is 3 h long.
Introductory instructions:
You may perform this experiment in groups of 3-5. Introduction:
Enzymes are proteinaceous molecules that speed up chemical
reactions within living systems. In this experiment, the effect of
catalase on hydrogen peroxide is investigated. Catalase is an
enzyme present in the cells of plants, animals and aerobic (oxygen
requiring) bacteria. It promotes the conversion of hydrogen
peroxide, a powerful and potentially harmful oxidizing agent, to
water and molecular oxygen.
2H2O2 + catalase 2H2O + O2
Warning: H2O2 is corrosive. For the person handling, please wear
gloves. Flowchart Students will be allowed to proceed with the
experiment only if they have come into the laboratory with a
flowchart of the days experiment. Procedures: 1. Optional: Set up
an electric water bath at 37oC. (If this is not provided, its
ok.)
2. Depending on the size of the rubber bung holding the delivery
tube, select either one
boiling or test tube and label it as tube A.
3. From the potato sample given, cut (with a pen knife/ knife/
scalpel) and weigh 5g of potato using a weighing boat so as not to
dirty the weighing balance.
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4. Cut the potato samples into smaller pieces (the smaller the
easier for you to mash) and mash the potato sample using the mortar
and pestle. Note: dont spend too much time on this it doesnt have
to be KFC mashed potato quality! Add 6 cm3 of distilled water to
the potato samples after the mashing process.
5. You can do two things: (i) separate the solid mashed potato
from the liquid in any
way you choose and pouring the liquid into a test tube; or (ii)
by filtering the mashed potato sample (with filter paper and
funnel) and collect the filtrate in a test tube or plastic vial (if
provided) [using filter paper and funnel is more
time-consuming].
6. Fill an empty test tube with tap water (see picture
below).
7. Add 5cm3 of hydrogen peroxide into Tube A using the hand-held
pipette provided.
8. Draw 1cm3 of the filtrate from the mashed potato samples and
add to Tube A.
Immediately close the test tubes with the rubber bung. Seal the
end of the delivery tube furthest away from the bung with
parafilm.
9. Set up the apparatus as shown below (if retort stand is
provided; if not just use each
others hands). Note: You need neither the water bath nor retort
stand.
10. Remove the parafilm and immediately immerse the tube in the
water bath quickly
(use a beaker for this and pour into it water from an electric
water bath) and start your watch. Count the number of gas bubbles
produced for 2 minutes and record it. After you finish, return the
water you took back to the electric water bath. [Note: water can
maintain the heat in it for quite some time.]
11. Optional, depends on time available: To get a 2nd
measurement, dispose the contents of tube A. Repeat step 7 to 10.
After you finish, return the water you took back to the electric
water bath.
12. Repeat Step 2 to 11 but with 10g of potato, then 15g and
finally 20g (optional,
depends on time available).
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13. Record the data in table 1. Your class is to record their
data on the whiteboard. Calculate the averages in order to plot
graphs.
Table 1: (title)
(What heading should you write here?)
5g 10g 15g *20g
Number of Attempt
1st 2nd *3rd 1st 2nd *3rd 1st 2nd *3rd *1st *2nd *3rd
Number of gas bubbles produced
*Optional, depends on time available. After the experiment
Please dispose of potato pieces, masking tape, parafilm etc. into
the dustbins provided. Please clean the sink, removing any potato
pieces, masking tape, parafilm etc. Assignments Please check with
your tutor which option is required for you. Option 1: (please
refer to WBLE/ Turnitin for instructions which may incorporate
other options below) Option 2: Skills-Based Assessment: Tabulation
of quantitative data (Table 1) Option 3: Skills-Based Assessment:
Graphing of quantitative data Present your graph (pasted from
Excel) of the average number of bubbles produced against potato
samples used. Use a best fit curve. To get full marks, please
observe the guidelines given on pp6-7 as well. No need to write
procedure, draw table, write a discussion or conclusion. Option 4A:
Skills-Based Assessment: Discussion Data provided to students to
discuss Write your discussion in prose form and without numbering.
Excluding your cover page, your discussion and conclusion should
NOT exceed ONE A4 page of Word document (standard/ default size).
Anything in excess will NOT be graded. Font Arial, size 11.
Margins: 1 inch from top, bottom, left and right (no need to change
if youre using the standard/ default size when MS Word opens).
Theory to apply: Refer to relevant information from lecture topics
which may or may not have been covered yet. *Option 4B:
Skills-Based Assessment: Discussion Students use own data to
discuss From the data you have collected in the practical, account
fully for the results which you have obtained. Discuss any
anomalous data/ results that you might have. Explain the trend or
pattern of the graph.
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Practical 3 Experiment 2 (FHSC 1214 Cell Biology ONLY) Synthesis
of Starch Using an Enzyme Extracted from a Potato Tuber
______________________________________________________________________
Objective: To find out which compounds the enzyme in the potato can
act on to produce starch (investigate three possible substrates:
glucose, maltose and glucose-1-phosphate). Apparatus and Materials:
Centrifuge and centrifuge tubes Glucose-1-phosphate (2%) Test tube
rack Glucose solution (2%) Pestle and mortar Maltose solution (2%)
Knife Iodine solution Labelling paper (or masking tape) Potato
tuber Test tubes Pipette White tile Procedures: Introductory
instructions:
Create a flowchart before you enter the lab in order to
understand the steps in this experiment. Show this to your tutor
before starting the experiment.
You may perform this experiment in pairs.
Take 5 ml iodine only when ready to begin the reaction.
Groups may have to take turns to centrifuge, depending on the
number of groups and holders in the centrifuge.
NOTE: After carrying out steps 1 to 2, proceed to Experiment 2.
Return to Experiment 1 only during the waiting periods of
Experiment 2.
A. Extracting the enzyme from potato tissue 1. Peel a
medium-sized half potato. Cut half of it into small cubes on a
white tile (the
smaller the easier for you to grind). Grind a few pieces of
potato cubes in a pestle and mortar with 20cm3 of water.
2. Separate the aqueous part of the extract from the solid as
best as possible. You can
do this by pouring it out while restraining the solids with your
fingers or an appropriate instrument. Divide the aqueous part of
the extract into two equal portions and pour them into two
centrifuge tubes. As far as possible, avoid letting sand and solid
matter to get into the tubes.
3. Spin the extracts in a centrifuge for ten minutes at 5000 rpm
so that the starch, cell
walls and other solid matter will settle at the bottom of the
centrifuge tubes. The starch-free liquid above the deposit, or
supernatant, should contain the enzyme.
4. Using a teat pipette, carefully, without disturbing the
deposit beneath, withdraw as
much the clear enzyme solution as possible from the centrifuge
tube. 5. To check whether this enzyme solution is starch-free,
transfer a few drops of it into a
test tube and add 2 drops of iodine solution onto it. If a blue
colour appears, then the potato extract would need to be
centrifuged again.
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B. Attempting starch synthesis 6. Label three clean test tubes
G, M and G1P, respectively. Use a separate teat pipette
and measuring cylinder in each case to place 3 cm3 of glucose
solution in the G tube, 3 cm3 of maltose solution in the M tube,
and 3cm3 of glucose-1-phosphate solution in the G1P tube.
7. To synthesise starch, pour 2 cm3 of the enzyme solution (the
liquid or supernatant
you obtained after centrifuging above) into the substrate tube
(G, M and G1P), mix well and note the time.
8. For each substrate, place 15 discrete drops of iodine
solutions on clearly labelled
piece of white tile. 9. After one minute of the reaction use a
teat pipette to place one drop of enzyme-
substrate solution onto one existing drop of iodine solution on
the white tile. Stir with a suitable object (e.g. woodsplint or
tooth pick) and record the colour produced. Repeat at intervals of
1 minute over 15 minutes, all the three tubes simultaneously.
Assignments Please check with your tutor which option is
required for you. Option 1: (please refer to WBLE/ Turnitin for
instructions which may incorporate other options below) Option 2:
Skills-Based Assessment: Discussion Discuss the following
questions: 1. Draw the structural formula of the substrates. What
features of the starch-
synthesizing substrate molecule might have been recognized by
the starch-synthesizing enzyme?
2. The synthesis of polymers such as starch requires metabolic
energy. What was the
energy source in the successful reaction? 3. The enzyme isolated
from potatoes is known as starch phosphorylase. In the intact
potato tuber it is also used to break down starch. How did
conditions in the test tube favor starch synthesis? In what
circumstances does the enzyme bring about starch synthesis in a
potato?
4. In plant leaves, starch accumulates in chloroplasts. The
synthesis of starch requires
ATP. Where do you think this ATP comes from?
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Practical 4 (FHSB 1214 Biology I & FHSC 1214 Cell Biology)
Investigation of the Enzymatic Effects of Materials on Hydrogen
Peroxide Solution
Objective: To investigates the enzymatic effect of various
materials in the hydrogen peroxide solution. Apparatus &
Equipment: Beaker Test tubes Either: water bath (95oC) or Bunsen
burner Materials: Fresh Liver Potato cubes Manganese dioxide
Hydrogen peroxide** Wood splints **Caution: Hydrogen peroxide is
formed continuously as a by-product of chemical reactions in living
cells; it is a very toxic (poisonous) substance. Flowchart Students
will be allowed to proceed with the experiment only if they have
come into the laboratory with a flowchart of the days experiment.
Procedures: Create a flowchart before you enter the lab in order to
understand the steps in this experiment. Show this to your tutor
before starting the experiment. Wear gloves when handling liver
tissue, so as not to be contaminated by any pathogen associated
with the liver tissue used. Please stick to using one pair of
gloves per person to prevent wastage. [Note: using boiling tubes
may provide better results.] 1. Label six fresh empty test or
boiling tubes 1, 2, 3, 4, 5, 6 and stand them in a rack. 2. Using a
razor blade, cut the provided liver into several pieces of roughly
0.8 cm x 0.8
cm x 0.5 cm. 3. Place one piece of liver into tube 1. 4. Boil
100 cm3 of water in a beaker. (If youre using a water bath set at
95oC, this step
is not necessary). 5. Place the second piece of liver into the
bottom of tube 2. Using a wooden splint,
gently spread the liver, without mashing it, over as wide an
area as possible of the bottom of the test or boiling tube. Place
tube 2 in the boiling or water bath (95oC) for about five
minutes.
6. Using the weighing balance, measure out two 0.5 g portions of
manganese dioxide
powder each onto a weighing boat. Pour each portion into tube 5
and tube 6.
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7. Put tube 6 in the beaker of boiling water or water bath
(95oC) for five minutes. 8. After five minutes let cool tube 2 and
6. 9. *Now put the third piece of liver into test or boiling tube
3. With the wooden stick
provided, mash it gently into a pulp.
10. Now put the third piece of liver onto a white tile. With a
mortar and pestle, mash it gently into a pulp. Scoop the pulp into
tube 3.
11. Cut potato cubes of roughly 0.8 cm x 0.8 cm x 0.5 cm. Place
one cube into a tube 4. 12. Prepare another six fresh empty test or
boiling tubes and stand them in a rack. Put 5
cm3 of hydrogen peroxide into each of them. 13. Next, quickly
add hydrogen peroxide into the test or boiling tubes 1, 2, 3, 4, 5,
and 6.
If needed, you may push down some materials with one end of the
wood splints provided.
**Step 12 and 13 are to be done quickly. 14. Using the parafilm
provided, stretch it quickly seal the mouth of the test or
boiling
tubes by stretching the film over it. In order to prevent the
parafilm from being displaced if a lot of gas is produced, secure
the parafilm covering the side of the test or boiling tube with
masking tape.
15. Leave for 20 minutes or till when you see quite a lot of gas
being produced in some test or boiling tubes as evidenced by the
bulging of parafilm from the test or boiling tube mouths.
16. Once enough gas has accumulated in some test or boiling
tubes, insert a glowing
splint (flame extinguished but glow remains) into each tube one
at a time by just penetrating the parafilm with it. You may use the
same splint.
Why is it important to test each test or boiling tube at least
without too much difference in the duration of sealing among the
tubes?
17. Record all your observations in the table. Record your
observations on each tube
immediately after the reaction has started. [Note: be sure to
use the following terms correctly: glowing splint glowed brighter,
flame rekindled, effervescence (bubbles) observed, reference to
sound, etc.]
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Table 1: (title)
Test Tube
Contents with 5 cm3
hydrogen peroxide Observations before and after using wood
splint
1
Fresh liver
2
Boiled liver (cooled)
3
Pulped liver
4 Potato cubes
5
Manganese dioxide (untreated)
6
Boiled manganese dioxide (cooled after heating)
Washing up Thoroughly wash and scrubbed all apparatus containing
liver pieces with detergent or Dettol solution provided to rid it
of unpleasant odours.
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Assignments Please check with your tutor which option is
required for you. Option 1: (please refer to WBLE/Turnitin for
instructions which may incorporate other options below) Option 2:
Skills-Based Assessment: Tabulation of qualitative data Tabulate
the results you obtained (unexpected results accepted). The results
table should not exceed 1 page. Option 3: Skills-Based Assessment:
Discussion Write your discussion in prose form and without
numbering. Excluding your cover page, your discussion and
conclusion should NOT exceed ONE A4 page of Word document
(standard/ default size). Anything in excess will NOT be graded.
Font Arial, size 11. Margins: 1 inch from top, bottom, left and
right (no need to change if youre using the standard/ default size
when MS Word opens). Theory to apply: Refer to relevant information
from lecture topics which may or may not have been covered yet.
Please be sure to address the following: 1. What is the equation of
the reaction observed? 2. What plant or animal organelle is
involved? 3. What effect does pulping the liver have upon the
reaction? Account for this. 4. What effect does boiling the liver
have upon the reaction? Account for this (include
reference to enzyme structure (bonds, molecular motion, shape,
active site). 5. What were the differences between the reactions
with fresh liver and with fresh
potato cubes? Account for these differences (include reference
to enzyme structure (bonds, molecular motion, shape, active
site)
6. What were the differences between the effects on the reaction
of boiling the liver and
heating the manganese dioxide? Account for these differences
(include reference to susceptibility (sensitivity) to heat, enzyme
shape, bonds etc).
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Practical 5 (FHSC 1214 Cell Biology); Practical 7 (FHSB 1214
Biology I) Exercise 1 The Microscope and Its Uses
Microscope usage (revision) Note to lecturer: Before any
microscope work (viewing of histological slides) commences, please
ensure students have gone through this introductory session.
Objective: To study the uses of microscope and its maintenances. To
learn microscopic techniques such as focus the object with correct
illumination under different power of magnifications. Introduction:
The microscope is a basic tool of the biologist. It is a valuable
precision optical instrument easily damaged by careless usage. It
is very important for the student to become familiar with the parts
of the microscope and the procedures in the handling of it. Treat
your microscope well and it will serve you well. Apparatus and
Materials: Binocular Microscope Cover slips Microscope slide
Newspaper (1 page) Plastic millimeter ruler Wash bottle Setting up
the Microscope: The microscope when not in use is usually kept in a
case. Remove it by grasping the handle arm while placing one hand
under the base. Set it down gently on the laboratory table and at a
reasonable distance from the table edge. Always keep the microscope
upright in the vertical position and never touch any of the lens
surfaces with the fingers since it will deposit a thin film of oil
on the glass.
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Parts of the Microscope:
Component Function
Arm For lifting and carrying the microscope.
Base To provide stability.
Body tube To house the lenses.
Eyepiece or ocular lenses
This is a set of lenses that rests loosely at the top end of the
body tube. It is obvious that if the microscope is tilted while
being carried, the lens may fall out and be ruined. The
magnification of the eyepiece (given as 10X) is printed on the
metal part of the ocular.
Revolving nosepiece
Located at the lower end of the body tube, it carries 3
objectives of different lengths. Rotating this part changes the
magnification of the objectives.
Objective lenses
They are of different magnifications with the following visible
properties:
Objectives Magnification Length Lens opening
Scanning lens 4x Shortest Widest
Low power lens 10x short wide
High power lens 60x longest narrowest
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Component Function
Focusing adjustments
These comprise two knobs located on either side of the
microscope which are used to change the distance between the object
being viewed and the objective lens. Changing the distance
determines the focus. For the object to be viewed in focus under
high magnification, the lens must be much closer to the object than
when it is under low magnification.
Coarse adjustment Made by the large knob beside the body tube
for focusing under low power magnification.
Fine adjustment Made by the small knob, which is for focusing
under high power magnification and accurate focusing.
Precautions when using the focusing adjustments: Turn both
adjustment knobs at the same time. Do not overturn the adjustment
knobs (i.e. do not force them to
go beyond their limits) Do not use the coarse adjustment knobs
when focussing under
the 60x objective lens.
Stage This is the platform for slides and specimens to be viewed
under the microscope.
Mechanical stage
This movable portion of the stage is attached to the specimen
holder and allows the slide to be moved in different directions to
facilitate viewing.
Specimen holder
This holds the glass slide in place.
Vertical feed knob
Rotating this moves the glass slide in the vertical
direction.
Horizontal feed knob
This moves the glass slide in the horizontal direction.
Condenser Located just beneath the stage of the microscope, it
incorporates a lens which collects light on the stage to bear on
the object.
Built-in light source
This is situated below the iris-diaphragm to provide light for
illuminating the object. It can be switched on or off.
Brightness adjustment knob
This provides adjustment to the illumination brightness.
Main switch This ensures that power is turned on or off.
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Preliminaries before Use: 1. Use the coarse adjustment to raise
the body tube so that the objective can clear
the stage when the revolving nosepiece is turned. 2. Turn the
nosepiece until the scanning objective is in-line with the
eyepiece. You
should hear a soft click or else feel a distinct falling into
place as the objective moves into position. If not, the field of
view is totally dark or an illuminated crescent instead of a
complete circle.
3. Turn the diaphragm to its largest opening. 4. Look into the
eyepiece and make a final adjustment to the light adjustment
knob
so that the field of view (i.e., the lit circle which you see)
is evenly illuminated. Any glare should be removed by adjusting the
diaphragm.
5. Should either of the lenses appear dirty, wipe it gently with
a piece of special
lens paper. Use a circular motion with very light finger
pressure. You should never use any other type of paper or cloth.
Discard the lens paper after use.
6. The microscope is now ready for use. 7. If youre using a
binocular compound light microscope like the diagram above,
position it so that the stage faces you. 8. Connect the
microscope to the power supply and turn on the built-in light. 9.
Ensure that the microscope stage is at its lowest position. This
will prevent breaking
of slides and lenses by mistake when adjusting the objectives by
moving the stage with the coarse adjustment knob.
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Using a higher power objective 1. Great care must be taken when
using higher power objectives. DO NOT focus the
high power objectives with the coarse adjustment knob. 2. Most
microscopes have parfocal objectives. This means that if one
switches from
viewing a specimen in sharp focus under a lower power objective
to a higher one, the object should automatically come approximately
into focus. Only some slight further focussing with the fine
adjustment knob is required to see the specimen clearly. Therefore,
if youre using the higher power objectives, do not use the coarse
knob to refine focus or youll risk breaking the slide and
lenses.
If the objectives are not parfocal, adjust the stage such that
it is about 1cm from the low-power objective. Change to the
high-power objective and then adjust the stage with the coarse
adjustment knob until it is about 1mm away from the objective. This
is determined by looking from the side of the microscope. Using the
fine adjustment knob and looking through the eyepiece now, slowly
bring the object into focus. Repeat the procedure carefully if the
first attempt at finding an object under high-power magnification
is unsuccessful.
3. When changing from one objective to another, you will hear a
click when the
objective is set in position. 4. You are now ready to switch
from the scanning objective to a higher power objective
after obtaining a sharp focus of the object. 5. If required,
adjust the fine adjustment knob to see the specimen clearly. 6. If
youre going to switch to the next higher power objective, look from
the side of the
microscope and move the revolving nosepiece slowly till that
higher power objective clicks into position. Be careful that it
does not touch the slide (normally it shouldnt unless the specimen
is too thick and also covered by a thin cover slip).
7. Take care that the lower end of the high power objective does
not touch the cover
slip. If this happens, you must repeat the whole procedure
focusing again, starting with the scanning objective.
Trouble-shooting Below are some common problems associated with
not being able to find and focus on an object under high-power
magnification.
Is the objective lens in position?
Is the cover slip on the slide facing upwards?
Is the object in the centre of the stage?
Are the lenses clean and free from dirt and moisture?
Is the condenser adjusted and focused?
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Oil Immersion: If your microscope comes with a 100 x objective,
please do NOT use it. Used the improper way, it will break. If you
require a particularly high magnification, immersion oil may be
used. Fluid with the same refractive index as the objective lens is
placed between a special objective lens and the cover slip so that
it touches both. The fluid permits a larger cone of light rays to
enter the objective from the specimen, and this increases the
resolving power obtainable. Microscope Care: Like all laboratory
instruments, the microscope needs proper care for best service.
Observe the following: 1. Turn the resolving nosepiece until the
scanning objective is in position. 2. Adjust the boy tube so that
the lower end of the objective is about 1 cm above the
stage. 3. Ensure that the stage surface is clean and dry. 4.
Return the microscope in an upright position to its storage case.
Activity: Manipulation Skill practice task Note to lecturer: this
activity may be graded. Any mistake will result in subtraction of 1
mark.
Microscope manipulation checklist Observed
Yes No
Skill: Manipulation
1. Position compound light microscope so that the stage faces
you.
2. Ensure that the microscope stage is at its lowest
position.
3. Position the specimen holder such that it is roughly in the
middle of the stage and not at either left or right extremes.
4. Secure the slide in position correctly with the specimen
holder
5. Ensure that the scanning objective is first employed.
6. Ensure that the field of view is a complete circle and not
totally dark or an illuminated crescent.
7. Both eyes open and used to look through the eyepieces.
8. Adjust the brightness adjustment knob to give the right
amount of light for viewing the object details clearly (i.e.,
instead of either too dark or too bright, obscuring the objects
finer details).
9. Focus on image accurately and sharply by using the coarse and
fine adjustment knobs.
10. When using the next higher power objective, look from the
side of the microscope to ensure that it does not touch the
slide.
11. When using higher power objectives (e.g., 40 X onwards),
only the fine adjustment knob is used (i.e., not the coarse
adjustment knob).
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Practical 5 (FHSC 1214 Cell Biology); Practical 7 (FHSB 1214
Biology I) Exercise 2 On-site Assessment
Each student will be assessed on-the-spot identification of 3
structures within certain minutes (10 marks) (The duration will be
decided by the tutor). This section may comprise 10 marks out of 20
marks. Any mistake will result in subtraction of 1 mark. Checklist
for on-site slide structure identification
Observed
Yes No
Skill: Manipulation
12. Position compound light microscope so that the stage faces
you and
ensure that the microscope stage is at its lowest position.
13. Position the specimen holder such that it is roughly in the
middle of
the stage and not at either left or right extremes.
14. Ensure that the scanning objective is first employed.
15. Ensure that the field of view is a complete circle and not
totally dark
or an illuminated crescent.
16. Both eyes open and used to look through the eyepieces.
17. Adjust the brightness adjustment knob to give the right
amount of
light for viewing the object details clearly (i.e., instead of
either too
dark or too bright, obscuring the objects finer details).
18. When using the next higher power objective, look from the
side of
the microscope to ensure that it does not touch the slide.
19. When using higher power objectives (e.g., 40 X onwards),
only the
fine adjustment knob is used (i.e., not the coarse adjustment
knob).
20. Focus on image accurately and sharply by using the coarse
and fine
adjustment knobs.
Skill: Identification
21. Able to name the specimen from the slide or identify two -
three
structures from the slide.
Total marks
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Practical 5 (FHSC 1214 Cell Biology); Practical 7 (FHSB 1214
Biology I) Exercise 3 Preparation of Wet Mount
Materials for microscopic examination are usually placed on the
glass slide of standard size, the microscope slide. In most cases,
the materials are then covered by small thin piece of glass, the
cover slip. Both microscope slide and cover slip should be very
clean before use. Cleaning microscope slides Hold the microscope
slide by the edges between the index flinger and the thumb and dip
in water. Then wipe dry using a soft tiss