Chemical Management Plan

University of the Philippines Baguio
Gov. Pack Road, Baguio City, Philippines 2600
CHEMICAL MANAGEMENT PLAN

Chemical Control Order (CCO)
Cyanide and Cyanide Compounds
Table of Contents
SECTION 1. STRUCTURE AND RESPONSIBILITIES / ACCOUNTABILITIES 1

SECTION 2. BRIEF DESCRIPTION OF THE CHEMICALS 2
SECTION 3. TRANSPORT 2
SECTION 4. STORAGE 3
SECTION 5. HANDLING 4
SECTION 6. DISPOSAL 6
SECTION 7. SUBSTITUTION AND PHASE OUT PLAN 7
SECTION 8. CONTINGENCY PLAN 7
SECTION 9. RECORDS 9
Appendix 1. Chemical Containers 11
Appendix 2. Laboratory Experiments 14
SECTION 1. STRUCTURE AND RESPONSIBILITIES / ACCOUNTABILITIES
All staff, teachers, students, and contractors who purchase, use, store, or dispose of chemicals
or controlled substances on behalf of the University of the Philippines Baguio are required to
undertake their responsibilities in line with the university’s rules and guidelines.
Table 1: Roles and responsibilities related to chemical management
Role Responsibility
Chancellor Responsible for all the activities conducted in the university
Pollution Control Implements and maintains measures consistent with the university’s
Managing Head chemical management plan, ensuring compliance with legislative
requirements
Pollution Control Officer Implements and maintains measures consistent with the university’s
chemical management plan, ensuring compliance with legislative
requirements; responsible for application of necessary permits from the
Environmental Management Bureau
Disaster Risk Reduction Ensure emergency planning is undertaken
Management Committee
(DRRMC)
Occupational Safety and Provide advice, issue safety bulletins, and support to areas in relation
Health (OSH) Committee to safety and health
Laboratory Technicians Maintain inventory of chemicals and regularly inspect storage area;
report all incidents
Faculty Members Ensure class compliance with the university’s health and safety
requirements, and chemical guidelines; report all incidents; provide
advice and guidance on appropriate management of chemical and
biological materials
Research Staff To maintain inventory of chemicals and comply with the university’s
health and safety requirements, and chemical guidelines; report all
incidents; provide advice and guidance on appropriate management of
biological materials and chemicals
Individual workers/ To comply with the university’s health and safety requirements, and
students chemical guidelines; report all incidents
UP Baguio
Chancellor
Pollution
Control
(blank)
Managing
Head
Pollution OSH
DRRMC
Control Officer Committee
Laboratory Faculty Research

Technicians Members Staff
Figure 1: Organization chart related to chemical management
Chemical Management Plan | Cyanide and Cyanide Compounds

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SECTION 2. BRIEF DESCRIPTION OF THE CHEMICALS
2.1 Physical and chemical properties:

Potassium cyanide Potassium ferrocyanide Sodium cyanide
Physical state Powder solid Powder solid Solid
Color White Yellow White
Odor Bitter almonds Odorless Bitter almond
Odor Threshold No data available No information available No information available
pH 11 - 12 9.5 11 - 12
Melting point/Range 634 °C 70 °C 562 °C
Initial boiling point and
1625 °C No information available 1497 °C
boiling range
Flash point No information available No information available No information available
Evaporation rate Not applicable Not applicable Not applicable
Upper flammability or
No data available No data available No data available
explosive limits
Lower flammability or
No data available No data available No data available
explosive limits
Vapor pressure No data available Negligible 1 HPa @ 817 °C
Vapor density Not applicable Not applicable Not applicable
Density 1.52 No information available No information available
Specific Gravity 1.52 1.850 No information available
Bulk Density No data available No information available 750 – 950 kg/m3
Water solubility 400 g/L Soluble No information available
Partition coefficient:
No information available No data available No data available
n-octanol/water
Autoignition temperature Not applicable Not applicable No information available
Decomposition
No data available > 70 °C No information available
temperature
Viscosity Not applicable Not applicable Not applicable
2.2 Chemical Use:
All these chemicals are mainly for laboratory use:
2.2.1 Class experiments

• Stability of proteins
• Enzymatic catalysis
2.2.2 Other approved research projects
2.3 Process description where chemicals are used:
Use in Research
Scheduling of
Procurement of Laboratory/
Storage Withdrawal from Disposal
Chemical Classroom
Storage
Experiments
Figure 2: Chemical use process flow
SECTION 3. TRANSPORT
3.1 Transport of Chemicals from the Receiving Area to Storage Area/ Stockroom
3.1.1 Upon arrival of the chemicals in the university, the chemicals should have
proper labelling (in accordance with GHS) and should be accompanied with
proper documents, Safety Data Sheet (SDS), and other appropriate permits.

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This will undergo inspection by the designated property inspector before
releasing the chemical to the end user.
3.1.2 The use of trolleys or carts is the recommended method of transport for
chemicals from the receiving area to the storage area.
3.2 Transport of Chemicals from the Storage Area to the Research Laboratory / Laboratory
Classroom
3.2.1 The use of a chemical cart is the recommended method of transport for
chemicals.
3.3 Any worker with probable exposure to the chemical should use personal protective
equipment (PPE) such as apron/lab gown, industrial gloves, safety shoes, and
appropriate clothing such as sleeved shirts and long pants.
3.4 The possibility of incompatible materials contacting one another, as a result of

container failure while being transported shall be evaluated. It shall be ensured that
such materials can be conveyed in a manner which will not allow chemical interaction.
SECTION 4. STORAGE
4.1 Stockroom
4.1.1 Safety Data Sheets and chemical inventory are available and accessible at
the stockroom.
4.1.2 The stockroom is properly ventilated and maintained.
4.1.3 Incompatible chemicals are not stored together. Chemicals are segregated
based on hazards category and compatibility of their physical and chemical
characteristics.
4.1.3.1 Chemicals with temperature requirements shall be kept in a

controlled temperature environment such as refrigerators.
4.1.3.2 Flammable chemicals will be stored in the flammables cabinet.
4.1.4 Chemicals and shelves are properly labelled.
4.1.5 Stockroom is kept locked when not in use. Only authorized persons have
access to the stockroom. No student or unauthorized faculty will be allowed
in the storage area unsupervised.
4.1.6 Chemical spill kits, PPE, eyewash, and fire safety kits are readily available.
Procedures shall be established to deal with clean up and safe disposal of
spillages.
4.1.7 Regular inspection and monitoring schedules of person in charge.
4.1.7.1 Leaking or damaged containers shall be removed and

transferred to a safe area for repacking or disposal.
4.1.7.2 Labels shall be reattached or replaced as necessary.

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4.2 Chemical Storage in Laboratories
4.2.1 Chemical storage cabinets are the recommended method of storage for
chemicals in the laboratory. Chemicals and shelves are properly labelled.
Cabinets must be always locked when not in use.
4.2.2 Incompatible chemicals are not stored together. Chemicals are segregated
based on hazards category and compatibility of their physical and chemical
characteristics.
4.2.3 Chemical spill kits, PPE, eyewash, and fire safety kits are readily available.
Procedures shall be established to deal with clean up and safe disposal of
spillages.
SECTION 5. HANDLING
5.1 Work Instructions
5.1.1 A laboratory manual will be provided to each student. The laboratory manual
contains the procedures and work instructions for all experiments. Research
methodology and protocols will be provided to all research staff.
5.2 Laboratory Safety Guidelines
5.2.1 Wear the prescribed PPE. No student is allowed to enter the laboratory and
carry out experiments without donning the complete prescribed Personnel
Protective Equipment
5.2.1.1 Laboratory safety goggles must be always worn in the laboratory

whenever experiments are being performed. Do not wear
contact lenses. Corrosive fumes and chemicals could get into
your eyes. Wearing contact lenses would prevent effective
flushing of the eyes, in case of an accident.
5.2.1.2 Closed shoes should always be worn in the laboratory as

protection against possible chemical spills and broken glass.
5.2.1.3 The prescribed laboratory gowns must be worn at all times when
experiments are being performed.
5.2.1.4 Protective face mask is recommended when handling chemicals

in the form of fine powder or in the presence of chemical fumes.
5.2.1.5 As an additional safety measure, long hair and loose clothing

must be properly secured to avoid interference in your work.
5.2.1.6 Avoid wearing jewelry in the laboratory.
5.2.2 Keep food, drinks, and gum out of the laboratory. Most chemicals in the
laboratory are poisonous. Eating, drinking, and chewing gum in the
laboratory are strictly prohibited.

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5.2.3 No smoking in the laboratory. Smoking is strictly prohibited, as some of the
laboratory chemicals are flammable. Flammable chemicals like organic
solvents must only be used under the fume hood.
5.2.4 Do not work alone in the laboratory. Perform experiments only during the
scheduled laboratory period and only under the supervision of your
laboratory instructor.
5.2.5 Do not perform unauthorized experiments. Only scheduled experiments

must be carried out during your laboratory period. Moreover, no additional
experiment procedures or deviations from the experiment procedures is
allowed, without the permission of the laboratory instructor.
5.2.6 No loitering in the laboratory. Do not play games or engage in horseplay in

the laboratory. Do not run around in the laboratory.
5.2.7 Keep all mobile phones and other gadgets in your bag.
5.2.8 Do not leave an on-going experiment set-up unattended.
5.2.9 Do not leave a lighted Bunsen burner unattended.
5.2.10 Read the labels. Carefully read the labels of chemical reagent containers,
check the chemical formula for correctness and check for appropriate
concentrations.
5.2.11 Handle all chemicals with extreme caution. Know the hazards associated
with the chemicals as well as their physical and chemical properties.
Examine the Safety Data Sheet (SDS) to know where to find specific
information on the hazards, toxicology, immediate first aid, and correct
disposal of the substance.
5.2.12 Keep knowledge of the exits, safety showers, eyewash, fire blankets and
extinguishers.
5.3 Hazards identification of Cyanide and Cyanide Compounds
5.3.1 Physical Hazards
5.3.1.1 Corrosive to metals
5.3.2 Health hazards
5.3.2.1 Acute oral toxicity

5.3.2.2 Acute dermal toxicity
5.3.2.3 Acute inhalation toxicity – dusts and mists
5.3.2.4 Specific target organ toxicity (repeated exposure)
5.3.3 Environmental hazards
5.3.3.1 Acute aquatic toxicity

5.3.3.2 Chronic aquatic toxicity

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5.4 PPE required when handling Cyanide and Cyanide Compounds
5.4.1 Eye/face protection: Wear appropriate chemical safety goggles
5.4.2 Hand protection: Protective gloves (Butyl rubber)
5.4.2.1 Inspect gloves before use.

5.4.2.2 Ensure gloves are suitable for the task: chemical compatibility,
dexterity, operational conditions, user susceptibility
5.4.2.3 Remove gloves with care avoiding skin contamination
5.4.3 Skin and body protection: Long sleeved clothing
5.4.4 Respiratory protection necessary
5.5 Standards for the area where the chemicals will be used
5.5.1 Packages and reagent bottles should not be opened in the actual storage
cabinet, shelf, or immediate storage area. This is to avoid the risks resulting
from handling obstructions and close proximity to other packages which may
lead to accidental escape of chemicals, vapors, or dust during transfers and
possible reaction with other substances.
5.5.2 The laboratory or working area must have proper ventilation. When handling
hazardous chemicals where toxic vapors or dust are generated, such as
these cyanide compounds, the use of a fume hood is a must.
5.5.3 Safe practices must be always utilized. Ensure that safety showers are close
to workstation location.
5.6 Maintenance of the process area
5.6.1 Make sure all the working areas are clean after the experiments, stoppers of
reagent bottles are replaced, and electronic materials are unplugged.
5.6.2 Dispose chemical wastes in appropriate waste containers.
SECTION 6. DISPOSAL
6.1 The university is a registered Hazardous Waste Generator. All hazardous wastes must
be correctly handled, stored, and labelled. These wastes are collected and stored in
the university’s accumulation area until the wastes are picked up by a DENR-
accredited transporter to be delivered to a DENR-accredited TSD facility for treatment
and disposal. Disposal must be done at least once a year.
6.2 PPE must always be used when handling hazardous wastes.
6.3 Chemical Wastes
6.1.1 Chemical wastes must be correctly handled, stored, and labelled.
6.1.2 4.0-L or 1-gallon bottles and 20.0-L carboy containers are allowed as liquid
waste containers and should only be filled up to 2/3 capacity.

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6.1.3 Chemical waste should not be allowed to accumulate and must only be
mixed with compatible waste.
6.1.4 Laboratory technicians and research staff shall coordinate with the Pollution
Control Office for the transfer of wastes from storage to accumulation area
of the university. This must be done at least once every semester.
6.4 Expired Chemicals
6.2.1 Expired chemicals shall be disposed of unless appropriate justification can

be made to keep it.
6.2.2 Chemicals with unknown expiration date will be disposed of after 5 years
unless appropriate justification can be made to keep it.
6.5 Contaminated items
6.3.1 Empty chemical containers must be properly stored in a designated area

until these will be transferred to the accumulation area.
6.3.2 Contaminated broken glasswares must be disposed of in designated bins

which are properly labelled.
6.3.3 Used spill kit items must be disposed of in designated bins which are
properly labelled.
SECTION 7. SUBSTITUTION AND PHASE OUT PLAN
7.1 Phaseout plan to be done within five (5) years to allow time for Research and
Development to develop the technology to properly adapt the new chemical to the
process
7.2 Consultation with faculty members on the phaseout plan and possible substitution of
these cyanide compounds.
7.3 Discussion with other Constituent Units on their current practices and phaseout plan
for cyanide compounds.
SECTION 8. CONTINGENCY PLAN
8.1 Hazardous material spills/leaks
8.1.1 When handling wastes, the following protective personal equipment (PPE)
should always be worn: laboratory gloves, goggles, mask and protective
clothing, whenever necessary.
8.1.2 Research project leaders and laboratory instructors should plan and prepare
for spill response as follows:
8.1.2.1 Make an emergency guide and post its chart in conspicuous

areas.
8.1.2.2 Make sure lab personnel read and understand the chemical spill
response procedures.

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8.1.2.3 If the chemicals in your lab require specific instructions not listed
in the emergency guide, establish standard operating
procedures for special conditions in your laboratory, and make
sure everyone working in the lab reads and understands the
procedures.
8.1.2.4 Assemble a spill kit tailored to clean up small spills of chemicals

commonly used in the lab, keep it fully stocked and easily
accessible and train personnel how to use its contents and when
it is safe to clean up a spill.
8.1.3 Research project/study leaders should make sure everyone working in the
laboratory knows:
8.1.3.1 The locations of fire extinguishers, eyewashes, and emergency

showers
8.1.3.2 How to operate the fire extinguisher and when it is safe to do so
8.1.3.3 How to use the eyewash and emergency shower
8.1.4 Small chemical spills are spills that can be cleaned up by lab personnel
without putting themselves or others in danger and should be handled as
follows:
8.1.4.1 Alert people in the area. Avoid breathing vapors and try to
determine what is spilled.
8.1.4.2 If someone has been splashed with chemicals, immediately flush

the affected area with water for at least 15 minutes and seek
medical attention as soon as possible.
8.1.4.3 Wear PPE and a long-sleeved lab coat during cleanup.
8.1.4.4 Confine the spill to a small area. Use a commercial kit or

absorbent material from your spill kit to absorb spilled materials.
8.1.4.5 Place the saturated absorbent in a plastic bag. Label the bag
with an HW tag and include it in the next HW collection.
8.1.4.6 Clean the spill area with water.
8.1.4.7 Replenish your spill kit supplies.
8.1.5 Moderate or large-scale spills are those that present an immediate hazard
(fire, explosion, chemical exposure, etc.) and should only be handled by
emergency responders. In case this kind of spill occurs, alert people in the
area and evacuate, closing all doors. Have someone available who is
knowledgeable about the spilled material to provide information to the
responders.
8.2 Emergency response in the laboratory

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8.2.1 General advice – Consult a physician. Show the SDS to the doctor in
attendance.
8.2.2 Fire – If your clothing catches on fire, drop and roll to put out the flames.
Immediately notify the supervisor of the incident.
8.2.3 Burns – Cool the burn using cold running water, seek medical attention and
notify the supervisor of the incident immediately. If the burn is due to an acid,
apply burn ointment that should be available from the laboratory’s first aid
kit.
8.2.4 Chemical spill on clothing – Get to the safety shower immediately and remain
there for at least 15 minutes. Remove contaminated clothing while in the
shower. Notify the immediate supervisor of the incident.
8.2.5 Chemical splashes to the eye – Immediately go to the eye wash station and
flush eyes with water for at least 15 minutes. Hold eyelids open to allow
water to reach all surfaces of the eyes and eyelids. Seek medical attention
and notify the supervisor of the incident immediately.
8.3 Natural disasters (Flood / Fire / Earthquake)
8.3.1 Posting of emergency numbers in conspicuous areas
8.3.2 Building premises shall have adequate fire, emergency or danger signs and
safety instructions.
8.3.3 Installation of proper firefighting equipment in accordance with the SDS and
compatible with the chemicals in storage.
8.3.4 Adequate emergency response procedure
8.3.5 Conducting regular safety drills and evacuation drills
SECTION 9. RECORDS
9.1 Chemical Inventory
9.1.1 Laboratory technicians and research staff must keep an inventory of the
chemicals stored in each storage location. Updating of the chemical
inventory must be done at least twice yearly to ensure all chemicals are
entered and all information is correct.
9.2 Chemical incidents and spill reporting
9.2.1 All incidents and spills involving hazardous substances must be reported to
the Pollution Control Officer.
9.2.2 The PCO will keep records of all reported incidents.
9.3 Transport and treatment of hazardous wastes
9.3.1. The PCO will keep records of all transport and treatment of hazardous
wastes

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9.4 Accidents/illness records
9.4.1 The Occupational Safety and Health (OSH) Committee maintains records of
all reported accidents and illness.
9.4.2 The Health Services Office (HSO) conducts annual physical examination for
all faculty and staff.
9.5 Safety Inspection Reports
9.5.1 Safety inspection reports and safety drills documentation are to be kept by
the DRRMC.
9.6 Trainings/Seminars
9.5.1 The PCO will keep records of all trainings and seminars on chemical
management and emergency response
9.5.2 The Human Resources Development Office (HRDO) keeps records of the
trainings and safety orientations of all faculty and staff.

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Appendix 1. Chemical Containers
Sodium cyanide
CAS Number 143-33-9
Figure 1. Chemical container of sodium cyanide

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Potassium cyanide
CAS Number 151-50-8
Figure 2. Chemical container of potassium cyanide

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Potassium ferrocyanide trihydrate
CAS Number 14459-95-1
Figure 3. Chemical container of potassium ferrocyanide trihydrate

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Appendix 2. Laboratory Experiments
Experiment 1. Extraction and Characterization of Proteins

Experiment 3. Enzymatic Catalysis

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EXPERIMENT 1
EXTRACTION AND CHARACTERIZATION OF PROTEINS
Jöns Jacob Berzelius discovered proteins in 1838. They are one of the classes of biological
macromolecules, alongside polysaccharides and nucleic acids that make up the primary constituents
of living things.
Proteins are high molecular weight organic compounds with very complex structures containing
may different kinds of simple substances called amino acids joined by peptide bonds. In addition, the
side chains of various amino acids in the polypeptide may interact, intrachain or interchain, resulting
to many different physical and chemical properties of proteins. Their functions range from the
enzymes that carry out the numerous metabolic processes of the cell to structural components that
provides cells their structures and organization.
Protein extraction involves a series of processes intended to isolate a single type of protein
from a complex mixture. Protein purification is vital for the characterization of the function, structure
and interactions of the protein of interest. The starting material is usually a biological tissue or a
microbial culture. The main goal in protein extraction is to abstract the biomolecule efficiently, causing
little degradation. Secondary but equally important goals are to preserve the proteins as much as
possible and to create an accurate representation of all proteins present in the original tissue. A variety
of protein extraction methods have been developed. Common techniques used in these methods
include grinding, homogenizing, centrifugation, salting out, dialysis, and ultrafiltration. The choice of
technique/s depends on the type of tissues and cell materials, how the sample has been treated, and
the proteins that are to be extracted. Similarly, the reagent to be used for protein extraction is also
dependent on the aforementioned factors.
One of the most useful parameters used in the study of proteins in solution is the concentration
of protein as this is necessary in subsequent quantitative determination of the activity of the protein.
The concentrations of protein in biological extracts can be estimated using spectrophotometric
methods. Bradford assay is a method based on the binding of Coomassie brilliant blue dye to proteins
under acidic conditions. The protein-dye complex causes a shift in the dye’s wavelength of maximum
absorption from 465 nm to 595 nm. By comparing the absorbance of a solution containing an unknown
amount of the protein–dye complex to the absorbance values of some standard solutions having
known concentrations, the concentration of protein in the sample can be estimated. The Bradford
assay has been found useful in determining protein concentrations in the range 1-20 μg/ml. The
Warburg-Christian method, on the other hand, is more suitable for semi-quantitative analysis of
biological samples and is generally applied in detecting protein concentrations in the range of 20-
3000 μg/ml. The method makes use of the maximum absorbance of tyrosine and tryptophan residues
at 280 nm for estimating total protein content and the strong absorption of nucleic acids at 260 nm.
To be biologically active, proteins must adopt specific folded three-dimensional tertiary
structures. Yet the genetic information for the protein specifies only the primary structure or the linear
sequence of amino acids in the polypeptide backbone. Many types of intermolecular forces are
believed to contribute in varying magnitudes to give the proteins’ stability. These forces can be long
range (ion-ion, ion dipole, or dipole-dipole) or short range (van der Waals repulsive and attractive
forces). The interactions can be local (between adjacent amino acids in the linear sequence) or
nonlocal (between sequences separated in the linear sequence but brought close together in 3D
space). Clues as to what stabilizes the tertiary structure of a native protein can be gained by subjecting
proteins to agents that unfold or denature a protein.
Denaturation is defined as the loss of natural conformation of proteins and nucleic acids. It
involves rupture and possible destruction of the secondary, tertiary, and quaternary structure of
Page 1 of 12
Chem 40.1 – Elementary Biochemistry Laboratory

Experiment 1 - Extraction and Characterization of Proteins
proteins. Denaturation of proteins does not affect the primary structure as peptide bonds are not
hydrolyzed in the process. When proteins are denatured, they assume random strand structure and
lose their biological activities. Denaturation is a cooperative process in the sense that the biopolymer
becomes increasingly more susceptible to denaturation once the process begins.
There are two approaches in defining the concept of denaturation: (1) molecular, in terms of
actual structural changes taking place on the molecule and (2) operational, in terms changes in
measurable properties.
The denaturation process can be achieved by physical methods such as increasing
temperature, changing pH, using high pressure, or carrying out ultrasonic homogenization. Chemical
methods can also be utilized such as the use of denaturants (i.e., urea, guanidine hydrochloride, beta-
mercaptoethanol, dithiothreitol), inorganic salts (i.e., lithium bromide, potassium thiocyanate, sodium
iodide), organic solvents (i.e., formamide, dimethylformamide, dichloroacetic and trichloroacetic acids
and their salts), and detergents (i.e., SDS).
Color reactions are useful tools in studying protein chemistry. Some of these are general tests
for proteins such as the Biuret and Ninhydrin tests. Other color reactions like xanthoproteic, Hopkins-
Cole, Millon’s and Cystine tests are indicative of the presence of specific functional groups.
LEARNING OUTCOMES
Upon completion of the experiment, the student should be able to:
1. perform isolation of casein from milk and albumin from egg;
2. describe the methods employed for protein extraction;
3. apply spectrophotometric methods in quantifying extracted proteins;
4. examine the effect of denaturants on a protein’s stability:
5. analyze the effects of protein denaturants by using color reaction tests; and
6. determine the amino acids present in different proteins using color reactions.
MATERIALS
Part I Part II Part III

1.00 M acetic acid 0.9% NaCl 1% standard protein solutions
saturated (NH4)2SO4 0.01 M NaOH 0.50% CuSO4
10% acetic acid 1% standard casein 3 M NaOH
95% ethanol 1% standard BSA Bradford Hopkins-Cole reagent
acetone reagent Concentrated H2SO4
50-mL graduated cylinders test tubes (12) 10% NaOH
stirring rod cuvettes 0.90 % NaCl solution
serological pipets UV-Vis spectrophotometer 1% CH3COOH
250-mL beakers extracts from Part I 2% CH3COOH
250-mL Erlenmeyer flask concentrated HNO3
cheesecloth saturated picric acid solution
droppers 10% TCA (trichloroacetic acid)
watch glass 5% potassium ferrocyanide
vials with lid saturated (NH4)2SO4
filter paper hot plate
funnel litmus
centrifuge test tubes
analytical balance pipette
top-loading balance extracts from Part I
hot plate with magnetic stirrer
refrigerator
fume hood
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PROCEDURES
Part I. EXTRACTION OF PROTEINS
A. Albumin from Egg

1. Separate the egg white from the yolk of one medium-sized egg and obtain 30.0 mL in a 50-
mL graduated cylinder.
2. Stir the egg white and add 3.00 mL of 1.00 M acetic acid dropwise.
3. Filter the mixture through cheesecloth (use stirring rod to speed up passage of filtrate) and
squeeze the cheesecloth to break the membranes. Collect the filtrate in a 250-mL beaker.
4. Add an equal volume of saturated (NH4)2SO4 solution to the filtrate.
5. Allow the mixture to stand for 30 minutes.
6. Centrifuge the mixture at 6000 rpm for 5 minutes. Discard the precipitate.
7. Transfer the supernatant into a 250-mL Erlenmeyer flask.
8. Add saturated (NH4)2SO4 to the clear yellow supernatant while continuously stirring until
turbidity persists.
9. Store the mixture in the refrigerator for at least 30 minutes.
10. Centrifuge the mixture and discard the supernatant. Transfer the residue in a pre-weighed
watch glass.
11. Air dry the albumin extract under the hood.
12. Determine the weight of the dry precipitate using weighing by difference. Have the crude
extract checked by the instructor.
13. Place the precipitate in a vial with a tight lid, label and store in the refrigerator for use in Part
II.
B. Casein from Milk

1. Weigh 20.0 g non-fat powdered milk in a weighing paper or measure 200 mL fresh milk
directly.
2. If sample is solid, dissolve the milk in 200 mL warm water in a 250-mL beaker.
3. Heat the milk solution to 40°C. Maintain this temperature.
4. Add 10.0 mL 10% acetic acid solution while stirring the mixture slowly and briefly. Avoid
adding excess acid.
5. Centrifuge the mixture at 3000 rpm for 10 minutes and discard the supernatant.
6. Wash the curd by resuspending it in enough cold 95% ethanol. Mix thoroughly to wash the
residue. Decant the ethanol washings. Wash the residue with ethanol repeatedly until the
ethanol washing is clear.
7. Weigh a piece of filter paper.
8. Transfer the moist residue into the pre-weighed filter paper, wash with acetone into a funnel
and air dry under the hood.
9. Determine the mass of the crude extract by weighing by difference and have it assessed by
the instructor.
10. Place the precipitate in a vial with a tight lid, label and store in the refrigerator for use in Part
II.
Part II. QUANTIFICATION OF PROTEIN CONCENTRATION BY SPECTROPHOTOMETRY
A. Sample preparation
Prepare your samples for the Warburg-Christian method and Bradford assay as follows:
1. Prepare 20.0 mL of 1% (w/v) solution of albumin in 0.9% NaCl.
2. Prepare 20.0 mL of 1% (w/v) casein solution using 0.01 M NaOH.
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NOTES:
*Reduce volumes to be prepared based on available mass of protein extract but always consult your
instructor first.
**Always store unused protein solutions in properly labeled, covered containers in the refrigerator
until Experiment 1 is completed.
B. Warburg- Christian Method

1. Measure the absorbance of the protein extracts prepared above at 280 and 260 nm. Measure
also the absorbance of the blank solution (solvent used to prepare the protein extract).
2. Compute the A280/A260 ratio and refer to the table below to estimate the % nucleic acid in the
protein extract.
Table 1.1. Ratio of the absorbance at 280 and 260 and its corresponding % nucleic acid.
A280/A260 % Nucleic Acid A280/A260 % Nucleic Acid
1.75 0.00 0.87 5.00
1.63 0.25 0.85 5.50
1.52 0.50 0.82 6.00
1.40 0.75 0.80 6.50
1.36 1.00 0.78 7.00
1.30 1.25 0.77 7.50
1.25 1.50 0.75 8.00
1.16 2.00 0.73 9.00
1.09 2.50 0.71 10.00
1.03 3.00 0.67 12.00
0.98 3.50 0.64 14.00
0.94 4.00 0.62 17.00
0.90 4.50 0.60 20.00
Estimate the purity of the protein isolate using the formula:
% purity = 100% ‒ % nucleic acid
3. Calculate the protein concentration in the crude extracts using the following equation:
Protein concentration (mg/mL) = (1.55 A280 ‒ 0.76 A260) ⨯ dilution factor (if applicable)
C. Bradford Assay
1. Set up 12 test tubes containing different amounts of protein standard and extract, distilled
water, and Bradford reagent (Refer to Table 1.2). Tube 1 will serve as blank; tubes 2 to 6 for
the construction of a standard curve for protein quantitation; tubes 7 to 12 are duplicates of
the three different concentrations of the isolates.
Table 1.2. Preparation for the Bradford assay.

Volume, mL
Test tube 1 2 3 4 5 6 7 8 9 10 11 12
Standard
0.00 0.20 0.40 0.60 0.80 1.00 --- --- --- --- --- ---
protein*
Protein
--- --- --- --- --- --- 0.30 0.30 0.50 0.50 0.70 0.70
extract
Page 4 of 12

Distilled
4.80 4.60 4.40 4.20 4.00 3.80 4.50 4.50 4.30 4.30 4.10 4.10
H2O
Bradford
0.20 0.20 0.20 0.20 0.20 0.20 0.20 0.20 0.20 0.20 0.20 0.20
reagent
*standard solution to be used should be of the same protein as that of the extract
2. Mix the contents of each tube thoroughly.

3. After 5 mins, read the absorbance of each tube at 595 nm. The absorbance readings should
be taken within 1 h after the addition of Bradford reagent.
4. Construct a calibration curve and determine the protein concentration of the crude extract.
5. If the crude extracts have absorbance readings outside the range established by the standard
curve, dilute the extracts with water and redo the absorbance measurements at 595 nm.
Consider the dilution factors when calculating for the actual protein concentration of the
extracts.
Part III. DENATURATION OF PROTEINS AND COLOR REACTIONS
A. Sample Preparation
1. Place 10.0 mL each of 1% standard protein solutions and 1% protein extracts from Part I
(casein/albumin) into separate macro-test tubes.
2. Dilute each with 10.0 mL 0.9% NaCl.
B. Denaturation
a. Heat Denaturation
1. Place 3.00 mL each of the protein solutions from section A above in labeled micro test tubes.
Heat the tubes in boiling water bath and observe.
2. Remove the tubes from the water bath and add 5 drops of 1% CH3COOH. Observe the effect.
3. Keep the solutions for the color reactions in section C. Refrigerate the solutions if they cannot
be used in the same laboratory period.
b. Using Concentrated Mineral Acids

1. To 1.00 mL of each protein solutions from section A, add concentrated HNO3 drop by drop
(shaking every after addition) until a precipitate forms. Observe the color of the precipitate.
2. Add more drops of concentrated HNO3, a drop at a time and note whether the solid that
previously formed will dissolve in excess acid or not.
c. Denaturation using Alkaloidal Reagents

1. Place 10 drops each of the protein solutions from section III-A in 3 different test tubes and
label as T1 –T3.
2. Add 3 drops of each of the following reagents:
T1 - saturated picric acid solution
T2 - 10% trichloroacetic acid (TCA)
T3 - acidify the protein solution by adding 1-2 drops of 2% acetic acid then add 3 drops
5% potassium ferrocyanide (K4[Fe(CN)6]) solution.
3. Observe precipitation if there is any. Then add excess (1-2 drops) of each reagent and observe
whether the amount of precipitate will increase or decrease.
d. Salting Out
1. Put 1.00 mL of each protein solutions from A in separate test tubes, then add 0.50 mL of
saturated (NH4)2SO4 solution and mix. Note the formation of a precipitate.
2. If there is no precipitate, add 3 -5 more drops but do not add a large excess of the salt. Observe
the separation of tiny protein aggregates, if there are any, that should float on the surface of
the mixture.
Page 5 of 12

3. Reverse the process by adding water dropwise with intermittent shaking until all evidence of
the aggregated protein has disappeared. Note the amount of water used.
C. Color Reactions of Proteins and Amino Acids

NOTE: Perform the following reactions on solutions of native proteins and on the denatured
samples from subpart B-a only. Use micro test tubes.
a. Biuret test
1. Mix 10 drops of each protein solution with 10 drops 10% NaOH. Add 1-2 drops of 0.5% CuSO4.
Note the color change, if any.
b. Xanthoproteic test
1. To 10 drops of protein sample add 10 drops concentrated HNO3.
2. Observe whether a heavy white precipitate is formed.
3. Heat carefully and observe whether the precipitate turns yellow and finally dissolves.
4. Record the color of the precipitate and/or the solution.
5. Cool and make alkaline with 10% NaOH. Record the color change, if any.
c. Cystine test
1. To 10 drops of protein sample, add 5 drops 3.00 M NaOH and 2 drops 1% Pb(NO3)2.
2. Heat the mixture in a boiling water bath for 5 mins. and note the color of precipitate formed, if
there is any.
d. Hopkins – Cole Reaction (Glyoxylic Reaction)

1. Mix 10 drops of protein solution and 5 drops Hopkins-Cole reagent.
2. Carefully add about 1.00 mL concentrated H2SO4 down the side of the tube so that there is a
minimum of mixing and a layer of H2SO4 forms under the protein-reagent layer.
3. Note the color at the junction of two liquids.
NOTE: Store in the refrigerator all prepared protein solutions and the denatured solutions from
subpart B-a. if the experiment is not completed in one lab session.
WASTE DISPOSAL
• Collect all solid wastes in a garbage bag and dispose in the trash bin.
• Discard mixtures from the two assays in the appropriate waste containers under the hood.
• Place all acid and alkaline wastes into their respective disposal jars.
• Discard hexane- or petroleum ether-containing wastes into the organic wastes jar.
• Cheesecloths may be washed with detergent. Reuse if necessary.
• Submit to the instructor all unused protein extracts in properly labeled and sealed vials.
• Discard mixtures from subparts B-a, B-b, B-d and C-d into acid waste jars.
• Discard mixtures from B-c into heavy metal disposal jars.
• Dispose all other waste solutions into the sink. Flush with ample amounts of running water.
REFERENCES
Biochemistry Laboratory Manual, 2002. UP Diliman.
Biochemistry Laboratory Manual. 2007. UP Diliman.
Boyer, R.F. 1993. Modern Experimental Biochemistry. Benjamin Cummings. Curr Protoc Mol Biol.
Nov. 2006.
Stenesh, J. 2004. Experimental Biochemistry. N.J.: Prentice Hall.
Page 6 of 12

PRE-LABORATORY QUESTIONS
Answer the following using the provided Prelab Sheets and submit at least one day before the first
day of the experiment.
1. Draw the schematic diagram for the extraction of either one of the proteins to be used.
2. Explain the principle behind the color change when Bradford reagent mixes with proteins.
3. Briefly explain (in one sentence) the general principle behind the action of the following
denaturants:
a. Heat
b. Mineral acids
c. Alkaloidal reagents
POST-LABORATORY QUESTIONS
Use these questions as guide in further understanding of the experiment and in your review for the
final examination.
1. State the purpose of each step in the extraction procedure of the protein assigned to your group.
(For separation procedures like filtration, decantation, and centrifugation, indicate what is being
removed)
2. Classify all the proteins used in this experiment based on function/s.
3. Compute the % purity and concentration of all the proteins extracted in the experiment. Show
sample calculations for one sample.
4. Compare and contrast the Warburg-Christian method and the Bradford assay as protein assay
techniques. Based on your results, explain which technique is more efficient.
5. Copy the table below and fill-out the information needed. Gather the data for other proteins from
other groups.
Observations
Protein Hopkins-
Biuret +/- Xanthoproteic +/- Cystine +/- +/-
Cole
standard
Albumin
extract
standard
Casein
extract
Based on the results above, make a list of the amino acid/s present in each protein extracted in
the experiment.
6. Give two important applications of protein denaturation related to medicine.
Page 7 of 12

DATA SHEET
EXPERIMENT 1
Part I. EXTRACTION OF PROTEINS
Protein: __________________
Fill out only the table for the protein assigned to your group.
Observations on the extraction of albumin.

Action Observations
egg white + 1.0 M CH3COOH

Filtration: Residue
Filtrate
filtrate + saturated (NH4)2SO4
After 30 minutes
Centrifugation: Residue
Supernatant
supernatant + saturated (NH4)2SO4
Supernatant
Residue
Albumin yield.
Mass, g
empty watch glass
Watch glass + albumin
albumin
Observations on the extraction of casein.

Action Observations
Heating
+ 10% CH3COOH
Supernatant
Washing with 95% EtOH
Washing with acetone
Casein yield.
Mass, g
empty filter paper
filter paper + casein
casein
Page 8 of 12

NOTES/REMARKS
Prepared by:
CHEM 40.1 Section Group
Names and signatures
Checked by:
Date:
Page 9 of 12

DATA SHEET
EXPERIMENT 1
Part II. QUANTIFICATION OF PROTEIN CONCENTRATION BY SPECTROPHOTOMETRY
Warburg-Christian assay
Albumin Casein
A280
A260
A280/A260
Bradford assay
Absorbance
Test Tube Albumin Casein
1
2
3
4
5
6
7
8
9
10
11
12
NOTES/REMARKS
Prepared by: Checked by:

CHEM 40.1 Date:
Section Group
Page 10 of 12

DATA SHEET
EXPERIMENT 1
Part III. DENATURATION OF PROTEINS AND COLOR REACTIONS
Protein: __________________
Observations on the denaturation of protein standards and extracts using heat.

Observations
Reagent/Action taken
Standard Extract
After heating
+ 1% CH3COOH
Observations on the denaturation of protein standards and extracts using concentrated

mineral acids.
Observations
Standard Extract
+ conc. HNO3
+ excess conc. HNO3
Observations on the denaturation of protein standards and extracts using alkaloidal

reagents.
Observations
Standard Extract
sample + picric acid (T1)
sample + 10% TCA (T2)
sample + 2% CH3COOH
+5% K4[Fe(CN)6] (T3)
Observations on the denaturation of protein standards and extracts via salting out.
Observations
Standard Extract
+ saturated (NH4)2SO4
+ excess saturated (NH4)2SO4
+ distilled H2O
Page 11 of 12

Observations on the color reactions of protein standards and extracts.
Observations
Standard Extract
Color reaction
Native Denatured Native Denatured
Biuret test
Xanthoproteic test
Cystine test
Hopkins-Cole test
NOTES/REMARKS
Prepared by:
CHEM 40.1 Section Group
Checked by:
Date:
Page 12 of 12

EXPERIMENT 3
ENZYMATIC CATALYSIS
A catalyst is a substance that lowers the activation energy required for a chemical reaction, and
therefore increases the rate of reaction without being used up in the process. An enzyme is a very large and
complex organic molecule synthesized by the cell to act as biological catalyst to speed up reactions in the
cell that would otherwise be too slow to support life.
Catalase is a heme-containing 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. Hydrogen peroxide (H2O2) is a poisonous byproduct of
metabolism that can damage cells if it is not removed. The reaction is as follows:
Figure 3.1. Pathway showing the breakdown on H2O2 by catalase.
Catalase is in a cell organelle called the peroxisome. Each catalase molecule is a tetramer of four
polypeptide chains. Each chain is composed of more than 500 amino acids. Located within this tetramer are
four porphyrin heme groups that are very much like hemoglobins, cytochromes and chlorophylls. The heme
group is responsible for catalase’s enzymatic activity. Catalase has one of the highest turnover rates for all
enzymes: one molecule of catalase can convert 6 million molecules of hydrogen peroxide to water and
oxygen each minute.
Enzyme kinetics is the study of the rate at which an enzyme acts. The rate at which an enzyme
works is influenced by several factors including the concentration of substrate, temperature, pH, salt
concentration and the presence of inhibitors or activators. Every enzyme has an optimal range for each
of these factors. Activity decreases when an enzyme is exposed to conditions that are outside the optimal
range.
The kinetic parameters KM and Vmax are commonly studied to describe the activity of enzymes. Values
of KM and Vmax are derived from the mathematical treatment of data obtained from enzyme kinetics. These
parameters appear in the Michaelis-Menten equation represented as:
where Vo = reaction rate or velocity, Vmax = maximum velocity, [S] = substrate molarity and Km =
Michaelis constant (KM) represents the substrate concentration when the reaction rate is equal to one-half
of the maximum velocity (Vmax/2). Plotting V versus [S] yields a hyperbolic curve.
In order to get a linear curve, instead of a hyperbolic curve, the Lineweaver-Burk plot is used. It is a
more convenient tool in determining KM and Vmax. It is also called the double reciprocal plot and is represented
as:
By plotting 1/V versus 1/[S], a straight line having a slope equal to KM/Vmax and a y-intercept equal to 1/ Vmax
will be obtained. By manipulating this slope and y-intercept, the values of KM and Vmax can be determined.
OBJECTIVES
Upon completion of the experiment, the student should be able to:
1. understand the general functions and activities of enzymes;
2. determine and explain the effect of several factors like substrate concentration, temperature, pH,
inhibitors and activators in the activity of enzymes;
3. explain how changes in temperature, pH, and the presence of inhibitors can affect the initial reaction
rates of enzyme-catalyzed reactions; and
4. review and apply titrimetric techniques of analysis.
MATERIALS
hot plate test tubes 30% H2O2
food processor/blender burette 0.10 M phosphate buffer pH 7
knife/cutter pipettor and pipette tips 0.005 M KCN of NaCN
Green papaya cheesecloth 6.00 N H2SO4
beakers E-flasks 0.10 M K2Cr2O7*
funnel ice
* Monitors should prepare 500 mL of this reagent for the whole class. Have your instructor check your
calculation before the preparation.
PROCEDURE
I. ENZYME KINETICS OF CATALASE
NOTE: Put all equipment, sample holder in the case of the blender and glassware to be used in the
refrigerator for 5 mins before performing parts A and B
A. Enzyme Preparation from Papaya (one set-up for the whole class)
1. Use a clean knife or cutter to peel the papaya then cut it into small pieces.
2. Weigh 150 g of cut papaya and add 150 mL of 0.10 M phosphate buffer pH 7.
3. Homogenize in blender until texture is smooth.
4. Filter the crude extract through cheesecloth.
5. Collect the filtrate and centrifuge at 5000 rpm for 10 minutes.
6. Collect supernatant and store in ice bath. Label the container as ‘’catalase extract’’.
B. Catalase Activity and Factors Affecting Enzymatic Activity

1. Set up eight test tubes as indicated in Table 3.1. Do not prepare the tubes simultaneously. Prepare
one at a time with at least 2-minutes interval.
2. Place the reagents in the tubes in precise order indicated in the table. Use micropipettes to measure
the substrate, enzyme and water. Letting the tubes stand for exactly 5 minutes is critical. Have the
H2SO4 ready to quench the reaction immediately after the 5-minute reaction time.
3. Do not add the catalase unless you are ready to proceed with titration immediately afterward.
4. Heat 1.00 mL of catalase extract in boiling water bath for 2 minutes. Do this only when you are ready
to add it to test tube 8.
Table 3.1. Set-up for the analysis of enzymatic activity of catalase.

Volume, mL
Reagent
Tube 1 Tube 2 Tube 3 Tube 4 Tube 5 Tube 6 Tube 7 Tube 8
Phosphate buffer 5.00 5.00 5.00 5.00 5.00 5.00 5.00 5.00
H2O2 0.25 0.50 1.00 1.50 2.00 0.50 0.50 0.50
H2O 1.75 1.50 1.00 0.50 --- 0.50 1.50 1.50
-3
5 x 10 M KCN (or
--- --- --- --- --- 1.00 --- ---
NaCN)
6 N H2SO4 --- --- --- --- --- --- 2.00
Catalase solution 1.00 1.00 1.00 1.00 1.00 1.00 1.00 ---
Hot catalase solution --- --- --- --- --- --- --- 1.00
o
Let all tubes stand 5 minutes at 0 C
6 N H2SO4 2.00 2.00 2.00 2.00 2.00 2.00 --- 2.00
5. Fill a buret with 0.10 M K2Cr2O7 solution. Use this reagent to titrate all reaction tubes (1 – 8) until a
distinct change to green color is obtained. Double check the end point by recording the final volume
and shaking the test tube once again to make sure that no more blue coloration is observed. Tabulate
the data (ml K2Cr2O7 used).
II. DATA ANALYSIS

A. Enzyme Kinetics of Catalase
The goal is to calculate the catalase activity (V) which is defined as the number of moles of H2O2
decomposed by the enzyme in 5 minutes per mL of catalase extract used. It can be calculated follows.
1. Calculate the number of moles (in mmol) H2O2 decomposed. It is equal to the difference of the
initial moles of H2O2 and titrated moles H2O2.
a. The initial number of moles of hydrogen peroxide in the tube before any reaction can be
calculated from the concentration of hydrogen peroxide solution used (e.g. 30% w/v, or lower
concentration if ‘agua oxinada’ from the pharmacy was used) and the volume of solution
dispensed. Convert to mmol.
b. To calculate the number of moles H2O2 titrated, consider the balanced redox reaction between
hydrogen peroxide and dichromate.
8H+ + Cr2O72- + 3H2O2 → 2Cr3+ + 3O2(g) + 7H2O
From the reaction, take note that one mole of dichromate is stoichiometrically equivalent to 3
moles of hydrogen peroxide. The titrated mmol of H2O2 can then be calculated by multiplying
the mmol of K2Cr2O7 used to this mole ratio.
b.1. The mmol of K2Cr2O7 used is equal to its molarity multiplied to its volume used to titrate a
particular test tube.
2. Finally, divide the mmol of H2O2 decomposed by 5 minutes.
B. Determination of the Km and Vmax

1. Use the [H2O2] and catalase activity data for each tube to get the Michaelis-Menten plot. You may
use Excel for plotting and adding a logarithmic trendline.
2. From the data in IIA, calculate 1/V and 1/initial [H2O2].
3. Obtain the Lineweaver-Burke plot. Alternatively, you may use the Eadie-Hofstee plot.
4. Determine Km and Vmax from the graph in Lineweaver-Burke plot.
5. Carry out the data analysis and compare your results to all other groups in the class. Explain any
significant differences in the Km and Vmax values, if there are any.
6. Using the catalase activity calculated for test tubes 2, 6, 7, and 8, conclude how different factors
affect the activity of catalase.
WASTE DISPOSAL
1. Collect all solid wastes in a garbage bag and dispose in the trash bin.
2. Discard mixtures from all tubes except tube 6 into acid into acid waste jars.
3. Discard mixture from tube 6 into cyanide disposal jars.
4. Dispose all other solutions into the sink. Flush with ample amounts of running water.
REFERENCES
Biochemistry Laboratory Manual. 2007. Quezon City: UP Diliman,
Mathews, C.K., K.E. Van Holde and K.G. Ahern. 2000. Biochemistry. CA, USA: Benjamin/Cummings.
PRE-LABORATORY QUESTIONS
1. Draw the schematic diagram for the isolation of catalase from green papaya.
2. Show, using a graph (reaction rate vs. factor) the general effects affecting the activity of enzymes.
Explain briefly.
3. What are the types of enzyme inhibition? Compare and contrast.
GUIDE FOR THE FORMAL REPORT

NOTE: Incorporate the following on the Results and Discussion part.
1. Define the following:

a. Lineweaver-Burke plot
b. Eadie-Hofstee plot
c. Michaelis-Menten plot
d. Kmax
e. Vmax
2. Discuss the principle behind the extraction of catalase.
3. For the analysis of catalase activity, tabulate your results as follows. Report values up to the 4th
decimal place.
Tube Tube Tube Tube Tube Tube Tube Tube

Parameter
1 2 3 4 5 6 7 8
Initial mmol H2O2
[H2O2], M
Volume of K2Cr2O7, mL
mmol K2Cr2O7
Final mmol H2O2
mmol H2O2 decomposed
Catalase activity, mmol/min//mL
1/V
1/[H2O2]
Concentration of H2O2 used ____________
Molarity of K2Cr2o7 used ____________
3a. Use data in test tube 1 to show sample calculations (place in appendix)
3b. Discuss the principle behind the titration and calculation of catalase activity.
3c. Use data from test tubes 1-5 to construct a Lineweaver-Burke plot. Determine KM and Vmax.
Interpret.
3d. Use data from test tubes 2, 6, 7, and 8 to discuss the factors affecting catalase activity.
EXPERIMENT 3
ENZYMATIC CATALYSIS
DATA SHEET
Table 3.1. Data on the titration of remaining H2O2 after activity of catalase using K2Cr2O7.
Volume K2Cr2O7 used, mL
Test tube
Trial 1 Trial 2 Trial 3 Average
1
8
NOTE: Perform one trial per group. Exchange data from other groups to have trials 2 and 3. Use the average
volume for the succeeding calculations.
Prepared by:
CHEM 40.1 Section _______

Group ______
Names and signature*
__________________________________
__________________________________
__________________________________
__________________________________
*those who are present only
Checked by:
__________________________________
Instructor
Date: _____________________________

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CHEMICAL MANAGEMENT PLAN

SECTION 1. STRUCTURE AND RESPONSIBILITIES / ACCOUNTABILITIES 1

Table 1: Roles and responsibilities related to chemical management

Laboratory Faculty Research

Figure 1: Organization chart related to chemical management

Chemical Management Plan | Cyanide and Cyanide Compounds

2.1 Physical and chemical properties:

2.2 Chemical Use:

All these chemicals are mainly for laboratory use:

2.2.1 Class experiments

2.2.2 Other approved research projects

2.3 Process description where chemicals are used:

Figure 2: Chemical use process flow

Chemical Management Plan | Cyanide and Cyanide Compounds

3.4 The possibility of incompatible materials contacting one another, as a result of

4.1.2 The stockroom is properly ventilated and maintained.

4.1.3.1 Chemicals with temperature requirements shall be kept in a

4.1.3.2 Flammable chemicals will be stored in the flammables cabinet.

4.1.4 Chemicals and shelves are properly labelled.

4.1.7 Regular inspection and monitoring schedules of person in charge.

4.1.7.1 Leaking or damaged containers shall be removed and

4.1.7.2 Labels shall be reattached or replaced as necessary.

Chemical Management Plan | Cyanide and Cyanide Compounds

5.1 Work Instructions

5.2 Laboratory Safety Guidelines

5.2.1.1 Laboratory safety goggles must be always worn in the laboratory

5.2.1.2 Closed shoes should always be worn in the laboratory as

5.2.1.4 Protective face mask is recommended when handling chemicals

5.2.1.5 As an additional safety measure, long hair and loose clothing

5.2.1.6 Avoid wearing jewelry in the laboratory.

Chemical Management Plan | Cyanide and Cyanide Compounds

5.2.5 Do not perform unauthorized experiments. Only scheduled experiments

5.2.6 No loitering in the laboratory. Do not play games or engage in horseplay in

5.2.8 Do not leave an on-going experiment set-up unattended.

5.2.9 Do not leave a lighted Bunsen burner unattended.

5.3 Hazards identification of Cyanide and Cyanide Compounds

5.3.1 Physical Hazards

5.3.1.1 Corrosive to metals

5.3.2 Health hazards

5.3.2.1 Acute oral toxicity

5.3.3 Environmental hazards

5.3.3.1 Acute aquatic toxicity

Chemical Management Plan | Cyanide and Cyanide Compounds

5.4.1 Eye/face protection: Wear appropriate chemical safety goggles

5.4.2 Hand protection: Protective gloves (Butyl rubber)

5.4.2.1 Inspect gloves before use.

5.4.3 Skin and body protection: Long sleeved clothing

5.4.4 Respiratory protection necessary

5.6 Maintenance of the process area

5.6.2 Dispose chemical wastes in appropriate waste containers.

6.2 PPE must always be used when handling hazardous wastes.

6.3 Chemical Wastes

6.1.1 Chemical wastes must be correctly handled, stored, and labelled.

Chemical Management Plan | Cyanide and Cyanide Compounds

6.4 Expired Chemicals

6.2.1 Expired chemicals shall be disposed of unless appropriate justification can